Catalogue 2020

CATALOGUE 2020

For finding your local dealer visit www.zytovision.com DISCLAIMER All information is supplied without liability. All product data are subject to changes. The information contained herein is believed to be correct and corresponds to the latest state of scientific and technical knowledge. However, no warranty is made, either ZytoVision GmbH · Fischkai 1 expressed or implied, regarding its accuracy. 27572 Bremerhaven · Germany · www.zytovision.com Dear Valued Customer,

ZytoVision GmbH is known to be an innovative German company focused on the development and production of high quality, state-of-the-art diagnostic products of prognostic, predictive and therapeutic value. We fulfil this claim by a continuous product development process in cooperation with many international clinical partners as well as strict and thorough quality controls during our production processes.

Nowadays, more and more genetic markers need to be evaluated on a patient’s sample to identify the appropriate treatment. In many cases, only small biopsy samples are available resulting in a limited number of slides on which immunohistochemistry, se- quencing, PCR, and/or in situ Hybridization (ISH) should be performed. These diagnostic requirements led us to the development of a new and innovative DistinguISH™ probe design for the simultaneous detection of different genetic markers on only one slide.

This catalogue presents our most current product portfolio of ISH probes and associated reagents, introducing many new products especially for the diagnosis of tumors of the hematopoietic and lymphoid tissues. Moreover, it includes our recently launched product line ZytoMation® for an automated FISH workflow and new lexF ISH® probes designed for a flexible FISH that allows choosing between a 1-day and a 2-day protocol. In order that treatments can be tailored with the utmost precision to the clinical profile of an individual patient, ZytoVision offers clinical trial services comprising the development of companion diagnostics.

We believe in a long-lasting relationship with our customers and support you via our worldwide network of highly qualified local distributors allowing us to respond to your needs immediately.

Always to meet your expectations is one of our major strategies. We would like to thank existing customers for their partnership, and would like to give a warm welcome to those of you who are new customers.

Sincerely,

Your ZytoVision Team Table of Contents

Page Method Introduction - ZytoLight ® 7 Probes, sorted by Chromosomes 8 ff. sorted by Gene Names 14 ff. sorted by Indication 20 ff. Product Data Sheets 26 ff. Accessories 184 f. FISH Reagents, Fluorochromes and Filter Recommendations 186 ff.

Page Method Introduction - FlexISH ® 175 Probes, sorted by Chromosomes 176 f. sorted by Gene Names 178 sorted by Indication 178 Product Data Sheets 179 ff. Accessories 184 f. FISH Reagents, Fluorochromes and Filter Recommendations 186 ff.

ZytoMation Page Method Introduction - ZytoMation ® 190 Probes, sorted by Chromosomes 191 sorted by Gene Names 192 sorted by Indication 192 Product Data Sheets 193 f. FISH Reagents, Fluorochromes and Filter Recommendations 186 ff.

Page Method Introduction - ZytoDot ® 196 - ZytoDot ® 2C TM 197 Probes, sorted by Chromosomes 198 ff. sorted by Gene Names 202 f. sorted by Indication 204 f. Product Data Sheets 206 ff. Accessories 244 f.

Page Method Introduction - ZytoFast ® 247 - ZytoFast ® PLUS 248 Probes, sorted by Virus Species 249 sorted by mRNAs 249 sorted by Indication 250 Product Data Sheets 251 ff. Accessories 256 f.

VisionArray Page Method Introduction - VisionArray ® 259 Product Data Sheets 260 ff. Accessories 263 ff.

General Information Page Product Use · Trademarks · CE Marking & ISO Certificates 266 Index 267 ff.

5 5 Page

Method Introduction - ZytoLight ® 7

Probes, sorted by Chromosomes 8 ff. sorted by Gene Names 14 ff. sorted by Indication 20 ff.

Product Data Sheets 26 ff.

Accessories 184 f.

FISH Reagents, Fluorochromes and Filter Recommendations 186 ff.

6 6 Reliable Multi-Target Detection using Fluorescence in situ Hybridization!

Introduction High Sensitivity and Specificity ZytoLight ® Kits – Convenient Solutions ZytoLight ® products are designed for ZytoLight ® FISH probes are direct labeled For making FISH analysis reliable and us- the identification of genetic aberrations using the unique ZytoLight ® Direct Label er-friendly, all ZytoLight ® FISH probes can e.g. translocations, deletions, amplifica- System II providing improved signal inten- be combined with the ZytoLight ® FISH-Tis- tions, and chromosomal aneuploidies by sity. All ZytoLight ® single copy (SPEC ™) sue Implementation Kit (Z-2028-5/-20) or Fluorescence in situ Hybridization (FISH) probes are processed by the unique the ZytoLight ® FISH-Cytology Implementa- in formalin-fixed, paraffin-embedded tissue ZytoLight ® Repeat Subtraction Technique tion Kit (Z-2099-20), for FISH analyses on sections, cell samples, blood or bone mar- resulting in advanced specificity and less cytology specimens. Both Implementation row smears, and metaphase chromosome background. No further blocking of repeti- Kits include all necessary pretreatment spreads. tive sequences is needed! solutions, wash buffers and DAPI/Dura- ZytoLight ® CEN ™ probes hybridize to Tect™-Solution and a detailed protocol to highly repetitive human satellite DNA perform successful FISH experiments. sequences of chromosomes producing sharp, bright signals specific for each individual chromosome.

The ZytoLight ® system uses direct labeled FISH probes 1, eliminating the need to detect the probes with fluorophore-coupled antibodies. The probes are detected by fluorescence microscopy using appropriate filter sets 2. Due to an exciter filter 3, full-spectrum light, emitted by the microscope lamp 4, is reduced to light of a defined wavelength that specifically excites the fluorophore of the probe. This light is reflected onto the specimen by a dichroic mirror 5. The fluorophore emits light of longer wave- lengths that passes the mirror. Finally, a barrier filter 6 reduces the emitted light to a defined wavelength that can be detected.

Protocol Overview

Pepsin Denature Apply Pretreatment Apply Probe Wash TM Analyze slides digestion & Hybridize DAPI/DuraTect

7 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 7 MLE001-1-20ILE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

1 1p36.3 ZytoLight Glioma 1p/19q Probe Set Z-2272-20 20 tests 26 ZytoLight SPEC 1p36/1q25 Dual Color Probe Z-2075-50/-200 50/200 µl 27 1p36.1 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe Z-2019-50/-200 50/200 µl 128 1p32.2 ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe Z-2276-50 50 µl 29 1p12 ZytoLight SPEC 1p12 Probe Z-2101-200 200 µl 170 f. ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe Z-2102-200 200 µl 43 1q21 ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe Z-2276-50 50 µl 29 ZytoLight SPEC MCL1/1p12 Dual Color Probe Z-2173-200 200 µl 30 ZytoLight SPEC MEF2D/BCL9 TriCheck™ Probe Z-2277-50 50 µl 31 1q22-q23.1 ZytoLight SPEC MEF2D/BCL9 TriCheck™ Probe Z-2277-50 50 µl 31 1q23.1 ZytoLight SPEC NTRK1 Dual Color Break Apart Probe Z-2167-50/-200 50/200 µl 32 1q25.2 ZytoLight SPEC ABL2 Dual Color Break Apart Probe Z-2200-50 50 µl 33 1q25.3 ZytoLight Glioma 1p/19q Probe Set Z-2272-20 20 tests 26 ZytoLight SPEC 1p36/1q25 Dual Color Probe Z-2075-50/-200 50/200 µl 27 1q32.1 ZytoLight SPEC MDM4/1p12 Dual Color Probe Z-2080-200 200 µl 34

2 2p24 ZytoLight SPEC MYCN/2q11 Dual Color Probe Z-2074-50/-200 50/200 µl 35 2p23 ZytoLight SPEC ALK/EML4 TriCheck™ Probe Z-2117-50/-200 50/200 µl 36 ZytoLight SPEC ALK Dual Color Break Apart Probe Z-2124-50/-200 50/200 µl 37 ZytoLight SPEC ALK/2q11 Dual Color Probe Z-2161-200 200 µl 38 2p21 ZytoLight SPEC EML4 Dual Color Break Apart Probe Z-2136-50 50 µl 39 ZytoLight SPEC ALK/EML4 TriCheck™ Probe Z-2117-50/-200 50/200 µl 36 2p11.2 ZytoLight SPEC IGK Dual Color Break Apart Probe Z-2288-50 50 µl 40 2q11.2 ZytoLight SPEC 2q11 Probe Z-2049-200 200 µl 170 f. ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe Z-2118-200 200 µl 44 2q34 ZytoLight SPEC ERBB4/2q11 Dual Color Probe Z-2057-200 200 µl 41 2q36 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe Z-2018-50/-200 50/200 µl 126 ZytoLight SPEC FOXO1/PAX3 TriCheck™ Probe Z-2185-50 50 µl 127

3 3p25 ZytoLight SPEC VHL/CEN 3 Dual Color Probe Z-2084-200 200 µl 42 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe Z-2102-200 200 µl 43 3p14.2 ZytoLight SPEC FHIT/CEN 3 Dual Color Probe Z-2062-200 200 µl 45 3p11.1-q11.1 ZytoLight CEN 3 Probe Z-2001-200 200 µl 170 f. ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe Z-2081-50/-200 50/200 µl 90 3q21 ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe Z-2287-50 50 µl 46 3q25.1 ZytoLight SPEC WWTR1 Dual Color Break Apart Probe Z-2212-50 50 µl 47 3q26.2 ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe Z-2287-50 50 µl 46 ZytoLight SPEC TERC/CEN 3 Dual Color Probe Z-2284-200 200 µl 48 3q26.3 ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe Z-2140-200 200 µl 49 ZytoLight SPEC SOX2/CEN 3 Dual Color Probe Z-2127-200 200 µl 50 3q27 ZytoLight SPEC BCL6 Dual Color Break Apart Probe Z-2177-50/-200 50/200 µl 51

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

8 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 8 ILE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

4 4p16.3 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe Z-2170-50/-200 50/200 µl 52 ZytoLight SPEC FGFR3/4p11 Dual Color Probe Z-2082-200 200 µl 53 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe Z-2282-50 50 µl 54 4p11 ZytoLight SPEC 4p11 Probe Z-2083-200 200 µl 170 f. 4q12 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck™ Probe Z-2209-50 50 µl 55

5 5p15.3 ZytoLight SPEC TERT Dual Color Break Apart Probe Z-2273-50 50 µl 56 ZytoLight SPEC TERT/5q31 Dual Color Probe Z-2091-50/-200 50/200 µl 57 5p13.1 ZytoLight SPEC RICTOR/5q31.1 Dual Color Probe Z-2278-200 200 µl 58 5q31.2 ZytoLight SPEC EGR1/5p15 Dual Color Probe Z-2107-50/-200 50/200 µl 59 ZytoLight SPEC EGR1/D5S23,D5S721 Dual Color Probe Z-2211-50 50 µl 60 5q32 ZytoLight SPEC CSF1R Dual Color Break Apart Probe Z-2202-50 50 µl 61 ZytoLight SPEC CSF1R/D5S23,D5S721 Dual Color Probe Z-2268-50 50 µl 62 ZytoLight SPEC NRG1/CD74 TriCheck™ Probe Z-2194-200 200 µl 79 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe Z-2197-50 50 µl 63

6 6p25 ZytoLight SPEC IRF4,DUSP22 Dual Color Break Apart Probe Z-2210-50 50 µl 64 6p24 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65 6p21.3 ZytoLight SPEC PHF1 Dual Color Break Apart Probe Z-2215-50 50 µl 66 6p21.1 ZytoLight SPEC VEGFA/CEN 6 Dual Color Probe Z-2195-200 200 µl 67 6p11.1-q11 ZytoLight CEN 6 Probe Z-2002-200 200 µl 170 f. ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe Z-2118-200 200 µl 44 6q22.1 ZytoLight SPEC ROS1 Dual Color Break Apart Probe Z-2144-50/-200 50/200 µl 68 ZytoLight SPEC ROS1/CEN 6 Dual Color Probe Z-2162-200 200 µl 69 6q23.3 ZytoLight SPEC MYB Dual Color Break Apart Probe Z-2143-50/-200 50/200 µl 70 ZytoLight SPEC MYB/CEN 6 Dual Color Probe Z-2281-50 50 µl 71 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65 6q25.1 ZytoLight SPEC ESR1/CEN 6 Dual Color Probe Z-2069-50/-200 50/200 µl 72

7 7p15.2-p15.1 ZytoLight SPEC JAZF1 Dual Color Break Apart Probe Z-2132-50 50 µl 73 7p11.2 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe Z-2033-50/-200 50/200 µl 74 7p11.1-q11.1 ZytoLight CEN 7 Probe Z-2003-200 200 µl 170 f. ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe Z-2081-50/-200 50/200 µl 90 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe Z-2102-200 200 µl 43 7q22 ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe Z-2214-50 50 µl 75 7q31.2 ZytoLight SPEC MET/CEN 7 Dual Color Probe Z-2087-50/-200 50/200 µl 76 7q34 ZytoLight SPEC BRAF Dual Color Break Apart Probe Z-2189-200 200 µl 77 ZytoLight SPEC BRAF/CEN 7 Dual Color Probe Z-2191-200 200 µl 78 7q36 ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe Z-2214-50 50 µl 75

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

9 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 9 ILE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

8 8p12 ZytoLight SPEC NRG1 Dual Color Break Apart Probe Z-2181-200 200 µl 80 ZytoLight SPEC NRG1/CD74 TriCheck™ Probe Z-2194-200 200 µl 79 8p11.2 ZytoLight SPEC FGFR1 Dual Color Break Apart Probe Z-2168-50/-200 50/200 µl 81 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe Z-2072-50/-200 50/200 µl 82 8p11.1-q11.1 ZytoLight CEN 8 Probe Z-2004-50/-200 50/200 µl 170 f. 8q21.3 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe Z-2112-50/-200 50/200 µl 83 8q24.21 ZytoLight SPEC MYC Dual Color Break Apart Probe Z-2090-50/-200 50/200 µl 84 ZytoLight SPEC MYC/CEN 8 Dual Color Probe Z-2092-50/-200 50/200 µl 85 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe Z-2105-50/-200 50/200 µl 86

9 9p24 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe Z-2179-50/-200 50/200 µl 87 ZytoLight SPEC JAK2 Dual Color Break Apart Probe Z-2294-50 50 µl 88 9p21 ZytoLight SPEC CDKN2A/CEN 9 Dual Color Probe Z-2063-50/-200 50/200 µl 89 ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe Z-2081-50/-200 50/200 µl 90 9q12 ZytoLight CEN 9 Probe Z-2067-200 200 µl 170 f. 9q21.3 ZytoLight SPEC NTRK2 Dual Color Break Apart Probe Z-2205-50/-200 50/200 µl 91 9q22.3-q31 ZytoLight SPEC NR4A3 Dual Color Break Apart Probe Z-2145-50 50 µl 92 9q34.1 ZytoLight SPEC ABL1 Dual Color Break Apart Probe Z-2199-50 50 µl 93 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe Z-2111-50/-200 50/200 µl 94 ZytoLight SPEC NUP214 Dual Color Break Apart Probe Z-2265-50 50 µl 95

10 10p11.2 ZytoLight SPEC KIF5B Dual Color Break Apart Probe Z-2131-50 50 µl 96 10p11.1-q11.1 ZytoLight CEN 10 Probe Z-2079-200 200 µl 170 f. 10q11.2 ZytoLight SPEC RET Dual Color Break Apart Probe Z-2148-50/-200 50/200 µl 97 10q23.3 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe Z-2078-50/-200 50/200 µl 98 10q26.1 ZytoLight SPEC FGFR2 Dual Color Break Apart Probe Z-2169-200 200 µl 99 ZytoLight SPEC FGFR2/CEN 10 Dual Color Probe Z-2122-200 200 µl 100

11 11p15.4 ZytoLight SPEC CARS Dual Color Break Apart Probe Z-2137-50 50 µl 101 ZytoLight SPEC NUP98 Dual Color Break Apart Probe Z-2266-50 50 µl 102 11p13 ZytoLight SPEC WT1 Dual Color Break Apart Probe Z-2142-50 50 µl 103 11p11.2 ZytoLight SPEC SPI1 Dual Color Break Apart Probe Z-2291-50 50 µl 104 11p11.11-q11 ZytoLight CEN 11 Probe Z-2005-200 200 µl 170 f. 11q13.3 ZytoLight SPEC CCND1 Dual Color Break Apart Probe Z-2108-50/-200 50/200 µl 105 ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe Z-2118-200 200 µl 44 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe Z-2125-50/-200 50/200 µl 107 11q21 ZytoLight SPEC MAML2 Dual Color Break Apart Probe Z-2014-50/-200 50/200 µl 108 11q22.2 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe Z-2146-50/-200 50/200 µl 109 11q22.3 ZytoLight SPEC ATM/CEN 11 Dual Color Probe Z-2297-50 50 µl 110 ZytoLight SPEC ATM/CEN 12 Dual Color Probe Z-2296-50 50 µl 111 ZytoLight SPEC TP53/ATM Dual Color Probe Z-2159-50/-200 50/200 µl 112 11q23.3 ZytoLight SPEC 11q gain/loss Triple Color Probe Z-2216-50 50 µl 115 ZytoLight SPEC KMT2A Dual Color Break Apart Probe Z-2193-50/-200 50/200 µl 116 11q24.3 ZytoLight SPEC 11q gain/loss Triple Color Probe Z-2216-50 50 µl 115 ZytoLight SPEC EWSR1/FLI1 TriCheck™ Probe Z-2183-50 50 µl 166

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

10 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 10 ILE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

12 12p13.3 ZytoLight SPEC ZNF384 Dual Color Break Apart Z-2275-50 50 µl 117 12p13.2 ZytoLight SPEC ETV6 Dual Color Break Apart Probe Z-2176-50/-200 50/200 µl 118 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe Z-2157-50/-200 50/200 µl 119 12p12.1 ZytoLight SPEC KRAS/CEN 12 Dual Color Probe Z-2115-200 200 µl 120 12p11.1-q11 ZytoLight CEN 12 Probe Z-2050-200 200 µl 170 f. 12q13.2 ZytoLight SPEC ERBB3/CEN 12 Dual Color Probe Z-2056-200 200 µl 121 12q13.3 ZytoLight SPEC DDIT3 Dual Color Break Apart Probe Z-2100-50/-200 50/200 µl 122 12q14 ZytoLight SPEC CDK4/CEN 12 Dual Color Probe Z-2103-50/-200 50/200 µl 123 12q15 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe Z-2013-50/-200 50/200 µl 124

13 13q12.1 ZytoLight SPEC 13q12 Probe Z-2085-200 200 µl 170 f. ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe Z-2095-50/-200 50/200 µl 170 f. ZytoLight SPEC 13/21 Dual Color Probe Z-2164-200 200 µl 170 f. ZytoLight Aneuploidy Panel 18/X/Y and 13/21 Z-2279-20 20 tests 172 ZytoLight Aneuploidy Panel X/Y and 13/18/21 Z-2104-5/-20 5/20 tests 173 13q14.1 ZytoLight SPEC FOXO1 Dual Color Break Apart Probe Z-2139-50 50 µl 125 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe Z-2018-50/-200 50/200 µl 126 ZytoLight SPEC FOXO1/PAX3 TriCheck™ Probe Z-2185-50 50 µl 127 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe Z-2019-50/-200 50/200 µl 128 13q14.2 ZytoLight SPEC D13S319/13q34/CEN 12 Triple Color Probe Z-2160-50/-200 50/200 µl 113 ZytoLight SPEC D13S319/13q34 Dual Color Probe Z-2280-50 50 µl 114 ZytoLight SPEC RB1/13q12 Dual Color Probe Z-2165-50/-200 50/200 µl 129

14 14q32.3 ZytoLight SPEC IGH Dual Color Break Apart Probe Z-2110-50/-200 50/200 µl 130 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe Z-2114-50/-200 50/200 µl 153 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe Z-2125-50/-200 50/200 µl 107 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe Z-2282-50 50 µl 54 ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe Z-2270-50 50 µl 137 ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe Z-2271-50 50 µl 138 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe Z-2105-50/-200 50/200 µl 86

15 15q14 ZytoLight SPEC NUTM1 Dual Color Break Apart Probe Z-2208-200 200 µl 131 15q24 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe Z-2113-50/-200 50/200 µl 132 15q25 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe Z-2206-50/-200 50/200 µl 133

16 16p13.3 ZytoLight SPEC CREBBP Dual Color Break Apart Probe Z-2267-50 50 µl 134 16p11.2 ZytoLight SPEC FUS Dual Color Break Apart Probe Z-2130-50 50 µl 135 16q22 ZytoLight SPEC CBFB Dual Color Break Apart Probe Z-2207-50 50 µl 136 16q23 ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe Z-2270-50 50 µl 137

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

11 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 11 ILE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

17 17p13 ZytoLight SPEC TP53/17q22 Dual Color Probe Z-2198-50 50 µl 139 ZytoLight SPEC TP53/ATM Dual Color Probe Z-2159-50/-200 50/200 µl 112 ZytoLight SPEC TP53/CEN 17 Dual Color Probe Z-2153-50/-200 50/200 µl 140 ZytoLight SPEC USP6 Dual Color Break Apart Probe Z-2151-50 50 µl 141 ZytoLight SPEC YWHAE Dual Color Break Apart Probe Z-2175-50 50 µl 142 17p11.1-q11.1 ZytoLight CEN 17 Probe Z-2006-200 200 µl 170 f. ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe Z-2081-50/-200 50/200 µl 90 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe Z-2102-200 200 µl 43 17q12 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Z-2015-50/-200 50/200 µl 143 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit Z-2020-5/-20 5/20 tests 143 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe Z-2077-50/-200 50/200 µl 144 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe Z-2190-50/-200 50/200 µl 145 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe Z-2093-50/-200 50/200 µl 146 17q21.2 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe Z-2093-50/-200 50/200 µl 146 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe Z-2113-50/-200 50/200 µl 132 17q21.3 ZytoLight SPEC COL1A1 Dual Color Break Apart Probe Z-2121-200 200 µl 147 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe Z-2116-50/-200 50/200 µl 148

18 18p11.1-q11.1 ZytoLight CEN 18 Probe Z-2007-200 200 µl 170 f. ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe Z-2095-50/-200 50/200 µl 170 f. ZytoLight Aneuploidy Panel 18/X/Y and 13/21 Z-2279-20 20 tests 172 ZytoLight Aneuploidy Panel X/Y and 13/18/21 Z-2104-5/-20 5/20 tests 173 18q11.2 ZytoLight SPEC SS18 Dual Color Break Apart Probe Z-2097-50/-200 50/200 µl 149 ZytoLight SPEC SS18/SSX1 TriCheck™ Probe Z-2184-50 50 µl 150 18q21.3 ZytoLight SPEC 18/CEN X/Y Triple Color Probe Z-2163-200 200 µl 170 f. ZytoLight SPEC BCL2 Dual Color Break Apart Probe Z-2192-50/-200 50/200 µl 151 ZytoLight SPEC BCL2/CEN 18 Dual Color Probe Z-2174-50 50 µl 152 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe Z-2114-50/-200 50/200 µl 153 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe Z-2146-50/-200 50/200 µl 109 ZytoLight SPEC MALT1 Dual Color Break Apart Probe Z-2196-50/-200 50/200 µl 154

19 19p13.3 ZytoLight Glioma 1p/19q Probe Set Z-2272-20 20 tests 26 ZytoLight SPEC 19q13/19p13 Dual Color Probe Z-2076-50/-200 50/200 µl 28 19q13.2 ZytoLight SPEC CIC Dual Color Break Apart Probe Z-2285-50 50 µl 155 19q13.3 ZytoLight Glioma 1p/19q Probe Set Z-2272-20 20 tests 26 ZytoLight SPEC 19q13/19p13 Dual Color Probe Z-2076-50/-200 50/200 µl 28 19q13.4 ZytoLight SPEC C19MC/19p13 Dual Color Probe Z-2274-50 50 µl 156

20 20q11.2 ZytoLight SPEC BCL2L1/CEN 20 Dual Color Probe Z-2171-200 200 µl 157 20q12 ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe Z-2271-50 50 µl 138 20q12-q13.1 ZytoLight SPEC PTPRT/20q11 Dual Color Probe Z-2213-50 50 µl 158

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

12 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 12 ILE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

21 21q22.1 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe Z-2112-50/-200 50/200 µl 83 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe Z-2157-50/-200 50/200 µl 119 21q22.1-q22.2 ZytoLight SPEC 21q22 Probe Z-2086-200 200 µl 170 f. ZytoLight SPEC 21/CEN X/Yq12 Triple Color Probe Z-2180-200 200 µl 170 f. ZytoLight SPEC 13/21 Dual Color Probe Z-2164-200 200 µl 170 f. ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe Z-2095-50/-200 50/200 µl 170 f. ZytoLight Aneuploidy Panel 18/X/Y and 13/21 Z-2279-20 20 tests 172 ZytoLight Aneuploidy Panel X/Y and 13/18/21 Z-2104-5/-20 5/20 tests 173 21q22.2 ZytoLight SPEC ERG Dual Color Break Apart Probe Z-2138-200 200 µl 159 ZytoLight SPEC ERG/TMPRSS2 TriCheck™ Probe Z-2135-200 200 µl 160 21q22.3 ZytoLight SPEC ERG/TMPRSS2 TriCheck™ Probe Z-2135-200 200 µl 160

22 22q11.2 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe Z-2111-50/-200 50/200 µl 94 ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe Z-2299-50 50 µl 161 ZytoLight SPEC DiGeorge Triple Color Probe Z-2289-50 50 µl 162 ZytoLight SPEC IGL Dual Color Break Apart Probe Z-2286-50 50 µl 163 ZytoLight SPEC SMARCB1/22q12 Dual Color Probe Z-2178-50 50 µl 164 22q12.2 ZytoLight SPEC EWSR1 Dual Color Break Apart Probe Z-2096-50/-200 50/200 µl 165 ZytoLight SPEC EWSR1/FLI1 TriCheck™ Probe Z-2183-50 50 µl 166 22q13.1 ZytoLight SPEC PDGFB Dual Color Break Apart Probe Z-2119-50/-200 50/200 µl 167 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe Z-2116-50/-200 50/200 µl 148 22q13.3 ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe Z-2299-50 50 µl 161

X Xp22.33 ZytoLight SPEC CRLF2 Dual Color Break Apart Probe Z-2201-50 50 µl 168 Xp11.23 ZytoLight SPEC SS18/SSX1 TriCheck™ Probe Z-2184-50 50 µl 150 ZytoLight SPEC TFE3 Dual Color Break Apart Probe Z-2109-50/-200 50/200 µl 169 Xp11.1-q11.1 ZytoLight CEN X Probe Z-2008-200 200 µl 170 f. ZytoLight CEN X/Yq12 Dual Color Probe Z-2016-50/-200 50/200 µl 170 f. ZytoLight CEN X/Y Dual Color Probe Z-2120-200 200 µl 170 f. ZytoLight SPEC 18/CEN X/Y Triple Color Probe Z-2163-200 200 µl 170 f. ZytoLight SPEC 21/CEN X/Yq12 Triple Color Probe Z-2180-200 200 µl 170 f. ZytoLight Aneuploidy Panel 18/X/Y and 13/21 Z-2279-20 20 tests 172 ZytoLight Aneuploidy Panel X/Y and 13/18/21 Z-2104-5/-20 5/20 tests 173

Y Yp11.32 ZytoLight SPEC CRLF2 Dual Color Break Apart Probe Z-2201-50 50 µl 168 Yp11.1-q11.1 ZytoLight CEN Y (DYZ3) Probe Z-2123-200 200 µl 170 f. ZytoLight CEN X/Y Dual Color Probe Z-2120-200 200 µl 170 f. ZytoLight SPEC 18/CEN X/Y Triple Color Probe Z-2163-200 200 µl 170 f. Yq12 ZytoLight CEN Yq12 Probe Z-2010-200 200 µl 170 f. ZytoLight CEN X/Yq12 Dual Color Probe Z-2016-50/-200 50/200 µl 170 f. ZytoLight SPEC 21/CEN X/Yq12 Triple Color Probe Z-2180-200 200 µl 170 f. ZytoLight Aneuploidy Panel 18/X/Y and 13/21 Z-2279-20 20 tests 172 ZytoLight Aneuploidy Panel X/Y and 13/18/21 Z-2104-5/-20 5/20 tests 173

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

13 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 13 ILE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

ABL1 ABL, c-ABL ZytoLight SPEC ABL1 Dual Color Break Apart Probe Z-2199-50 50 µl 93 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe Z-2111-50/-200 50/200 µl 94

ABL2 ARG ZytoLight SPEC ABL2 Dual Color Break Apart Probe Z-2200-50 50 µl 33

ALK CD246 ZytoLight SPEC ALK/EML4 TriCheck™ Probe Z-2117-50/-200 50/200 µl 36 ZytoLight SPEC ALK Dual Color Break Apart Probe Z-2124-50/-200 50/200 µl 37 ZytoLight SPEC ALK/2q11 Dual Color Probe Z-2161-200 200 µl 38

ATM ATA, TEL1 ZytoLight SPEC ATM/CEN 11 Dual Color Probe Z-2297-50 50 µl 110 ZytoLight SPEC ATM/CEN 12 Dual Color Probe Z-2296-50 50 µl 111 ZytoLight SPEC TP53/ATM Dual Color Probe Z-2159-50/-200 50/200 µl 112

BCL2 Bcl-2, PPP1R50 ZytoLight SPEC BCL2 Dual Color Break Apart Probe Z-2192-50/-200 50/200 µl 151 ZytoLight SPEC BCL2/CEN 18 Dual Color Probe Z-2174-50 50 µl 152 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe Z-2114-50/-200 50/200 µl 153

BCL2L1 BCLX ZytoLight SPEC BCL2L1/CEN 20 Dual Color Probe Z-2171-200 200 µl 157

BCL6 ZNF51, LAZ3 ZytoLight SPEC BCL6 Dual Color Break Apart Probe Z-2177-50/-200 50/200 µl 51

BCL9 - ZytoLight SPEC MEF2D/BCL9 TriCheck™ Probe Z-2277-50 50 µl 31

BCR ALL, BCR1 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe Z-2111-50/-200 50/200 µl 94

BIRC3 C-IAP, MALT2 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe Z-2146-50/-200 50/200 µl 109

BRAF BRAF1, NS7 ZytoLight SPEC BRAF Dual Color Break Apart Probe Z-2189-200 200 µl 77 ZytoLight SPEC BRAF/CEN 7 Dual Color Probe Z-2191-200 200 µl 78

C19MC - ZytoLight SPEC C19MC/19p13 Dual Color Probe Z-2274-50 50 µl 156

CARS CARS1 ZytoLight SPEC CARS Dual Color Break Apart Probe Z-2137-50 50 µl 101

CBFB PEBP2B ZytoLight SPEC CBFB Dual Color Break Apart Probe Z-2207-50 50 µl 136

CCND1 BCL1, PRAD1 ZytoLight SPEC CCND1 Dual Color Break Apart Probe Z-2108-50/-200 50/200 µl 105 ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe Z-2118-200 200 µl 44 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe Z-2125-50/-200 50/200 µl 107

CD274 PD-L1, PDL1 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe Z-2179-50/-200 50/200 µl 87

CD74 - ZytoLight SPEC NRG1/CD74 TriCheck™ Probe Z-2194-200 200 µl 79

CDK4 PSK-J3 ZytoLight SPEC CDK4/CEN 12 Dual Color Probe Z-2103-50/-200 50/200 µl 123

CDKN2A p16, ARF, INK4 ZytoLight SPEC CDKN2A/CEN 9 Dual Color Probe Z-2063-50/-200 50/200 µl 89 ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe Z-2081-50/-200 50/200 µl 90

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

14 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 14 ILE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

CDNK2C - ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe Z-2276-50 50 µl 29

CIC KIAA0306 ZytoLight SPEC CIC Dual Color Break Apart Probe Z-2285-50 50 µl 155

CKS1B - ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe Z-2276-50 50 µl 29

COL1A1 OI4 ZytoLight SPEC COL1A1 Dual Color Break Apart Probe Z-2121-200 200 µl 147 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe Z-2116-50/-200 50/200 µl 148

CREBBP CBP, RTS ZytoLight SPEC CREBBP Dual Color Break Apart Probe Z-2267-50 50 µl 134

CRKL - ZytoLight SPEC DiGeorge Triple Color Probe Z-2289-50 50 µl 162

CRLF2 CRL2, TSLPR ZytoLight SPEC CRLF2 Dual Color Break Apart Probe Z-2201-50 50 µl 168

CSF1R FMS ZytoLight SPEC CSF1R Dual Color Break Apart Probe Z-2202-50 50 µl 61 ZytoLight SPEC CSFR1/D5S23,D5S721 Dual Color Probe Z-2268-50 50 µl 62

CUX1 CUT ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe Z-2214-50 50 µl 75

DDIT3 CHOP, GADD153 ZytoLight SPEC DDIT3 Dual Color Break Apart Probe Z-2100-50/-200 50/200 µl 122

DLEU1 BCMS1, LEU1 ZytoLight SPEC D13S319/13q34/CEN 12 Triple Color Probe Z-2160-50/-200 50/200 µl 113 ZytoLight SPEC D13S319/13q34 Dual Color Probe Z-2280-50 50 µl 114

DUSP22 JKAP ZytoLight SPEC IRF4,DUSP22 Dual Color Break Apart Probe Z-2210-50 50 µl 64

EGFR HER1, ERBB1 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe Z-2033-50/-200 50/200 µl 74

EGR1 KROX-24 ZytoLight SPEC EGR1/5p15 Dual Color Probe Z-2107-50/-200 50/200 µl 59 ZytoLight SPEC EGR1/D5S23,D5S721 Dual Color Probe Z-2211-50 50 µl 60

EML4 ROPP120 ZytoLight SPEC EML4 Dual Color Break Apart Probe Z-2136-50 50 µl 39 ZytoLight SPEC ALK/EML4 TriCheck™ Probe Z-2117-50/-200 50/200 µl 36

ERBB2 HER2, HER-2, NEU ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Z-2015-50/-200 50/200 µl 143 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit Z-2020-5/-20 5/20 tests 143 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe Z-2077-50/-200 50/200 µl 144 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe Z-2190-50/-200 50/200 µl 145 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe Z-2093-50/-200 50/200 µl 146

ERBB3 HER3 ZytoLight SPEC ERBB3/CEN 12 Dual Color Probe Z-2056-200 200 µl 121

ERBB4 HER4, ALS19 ZytoLight SPEC ERBB4/2q11 Dual Color Probe Z-2057-200 200 µl 41

ERG erg-3, p55 ZytoLight SPEC ERG Dual Color Break Apart Probe Z-2138-200 200 µl 159 ZytoLight SPEC ERG/TMPRSS2 TriCheck™ Probe Z-2135-200 200 µl 160

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

15 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 15 ILE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

ESR1 Era, NR3A1 ZytoLight SPEC ESR1/CEN 6 Dual Color Probe Z-2069-50/-200 50/200 µl 72

ETV6 TEL ZytoLight SPEC ETV6 Dual Color Break Apart Probe Z-2176-50/-200 50/200 µl 118 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe Z-2157-50/-200 50/200 µl 119

EWSR1 EWS ZytoLight SPEC EWSR1 Dual Color Break Apart Probe Z-2096-50/-200 50/200 µl 165 ZytoLight SPEC EWSR1/FLI1 TriCheck™ Probe Z-2183-50 50 µl 166

EZH2 KMT6A ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe Z-2214-50 50 µl 75

FGFR1 FLT2, BFGFR ZytoLight SPEC FGFR1 Dual Color Break Apart Probe Z-2168-50/-200 50/200 µl 81 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe Z-2072-50/-200 50/200 µl 82

FGFR2 BEK, CD332 ZytoLight SPEC FGFR2 Dual Color Break Apart Probe Z-2169-200 200 µl 99 ZytoLight SPEC FGFR2/CEN 10 Dual Color Probe Z-2122-200 200 µl 100

FGFR3 CD333, JTK4 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe Z-2170-50/-200 50/200 µl 52 ZytoLight SPEC FGFR3/4p11 Dual Color Probe Z-2082-200 200 µl 53 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe Z-2282-50 50 µl 54

FHIT FRA3B ZytoLight SPEC FHIT/CEN 3 Dual Color Probe Z-2062-200 200 µl 45

FIP1L1 FIP1 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck™ Probe Z-2209-50 50 µl 55

FLI1 EWSR2 ZytoLight SPEC EWSR1/FLI1 TriCheck™ Probe Z-2183-50 50 µl 166

FOXO1 FKHR, FKH1 ZytoLight SPEC FOXO1 Dual Color Break Apart Probe Z-2139-50 50 µl 125 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe Z-2018-50/-200 50/200 µl 126 ZytoLight SPEC FOXO1/PAX3 TriCheck™ Probe Z-2185-50 50 µl 127 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe Z-2019-50/-200 50/200 µl 128

FUS FUS1 ZytoLight SPEC FUS Dual Color Break Apart Probe Z-2130-50 50 µl 135

GATA2 NFE1B ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe Z-2287-50 50 µl 46

HIRA TUPLE1, TUP1 ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe Z-2299-50 50 µl 161 ZytoLight SPEC DiGeorge Triple Color Probe Z-2289-50 50 µl 162

IGH IGH@ ZytoLight SPEC IGH Dual Color Break Apart Probe Z-2110-50/-200 50/200 µl 130 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe Z-2114-50/-200 50/200 µl 153 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe Z-2125-50/-200 50/200 µl 107 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe Z-2282-50 50 µl 54 ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe Z-2270-50 50 µl 137 ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe Z-2271-50 50 µl 138 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe Z-2105-50/-200 50/200 µl 86

IGK IGK@ ZytoLight SPEC IGK Dual Color Break Apart Probe Z-2288-50 50 µl 40

IGL IGL@ ZytoLight SPEC IGL Dual Color Break Apart Probe Z-2286-50 50 µl 163

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

16 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 16 ILE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

IRF4 MUM1 ZytoLight SPEC IRF4,DUSP22 Dual Color Break Apart Probe Z-2210-50 50 µl 64

JAK2 JTK10 ZytoLight SPEC JAK2 Dual Color Break Apart Probe Z-2294-50 50 µl 88

JAZF1 TIP27, ZNF802 ZytoLight SPEC JAZF1 Dual Color Break Apart Probe Z-2132-50 50 µl 73

KIF5B KNS1 ZytoLight SPEC KIF5B Dual Color Break Apart Probe Z-2131-50 50 µl 96

KMT2A MLL ZytoLight SPEC KMT2A Dual Color Break Apart Probe Z-2193-50/-200 50/200 µl 116

KRAS KRAS1 ZytoLight SPEC KRAS/CEN 12 Dual Color Probe Z-2115-200 200 µl 120

MAF - ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe Z-2270-50 50 µl 137

MAFB - ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe Z-2271-50 50 µl 138

MALT1 MLT ZytoLight SPEC MALT1 Dual Color Break Apart Probe Z-2196-50/-200 50/200 µl 154 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe Z-2146-50/-200 50/200 µl 109

MAML2 MAM3 ZytoLight SPEC MAML2 Dual Color Break Apart Probe Z-2014-50/-200 50/200 µl 108

MAPK1 PRKM2, ERK ZytoLight SPEC DiGeorge Triple Color Probe Z-2289-50 50 µl 162

MCL1 BCL2L3 ZytoLight SPEC MCL1/1p12 Dual Color Probe Z-2173-200 200 µl 30

MDM2 HDM2 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe Z-2013-50/-200 50/200 µl 124

MDM4 MDMX ZytoLight SPEC MDM4/1p12 Dual Color Probe Z-2080-200 200 µl 34

MECOM MDS1, EVI1 ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe Z-2287-50 50 µl 46

MEF2D - ZytoLight SPEC MEF2D/BCL9 TriCheck™ Probe Z-2277-50 50 µl 31

MET HGFR, RCCP2 ZytoLight SPEC MET/CEN 7 Dual Color Probe Z-2087-50/-200 50/200 µl 76

MYB c-myb ZytoLight SPEC MYB Dual Color Break Apart Probe Z-2143-50/-200 50/200 µl 70 ZytoLight SPEC MYB/CEN 6 Dual Color Probe Z-2281-50 50 µl 71 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65

MYC CMYC, bHLHe39, c-Myc ZytoLight SPEC MYC Dual Color Break Apart Probe Z-2090-50/-200 50/200 µl 84 ZytoLight SPEC MYC/CEN 8 Dual Color Probe Z-2092-50/-200 50/200 µl 85 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe Z-2105-50/-200 50/200 µl 86

MYCN NMYC, N-myc ZytoLight SPEC MYCN/2q11 Dual Color Probe Z-2074-50/-200 50/200 µl 35

NR4A3 CHN, CSMF ZytoLight SPEC NR4A3 Dual Color Break Apart Probe Z-2145-50 50 µl 92

NRG1 HGL, GGF ZytoLight SPEC NRG1 Dual Color Break Apart Probe Z-2181-200 200 µl 80 ZytoLight SPEC NRG1/CD74 TriCheck™ Probe Z-2194-200 200 µl 79

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

17 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 17 ILE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

NTRK1 MTC, TRK ZytoLight SPEC NTRK1 Dual Color Break Apart Probe Z-2167-50/-200 50/200 µl 32

NTRK2 TRKB ZytoLight SPEC NTRK2 Dual Color Break Apart Probe Z-2205-50/-200 50/200 µl 91

NTRK3 TRKC ZytoLight SPEC NTRK3 Dual Color Break Apart Probe Z-2206-50/-200 50/200 µl 133

NUP98 NUP96 ZytoLight SPEC NUP98 Dual Color Break Apart Probe Z-2266-50 50 µl 102

NUP214 CAN, CAIN ZytoLight SPEC NUP214 Dual Color Break Apart Probe Z-2265-50 50 µl 95

NUTM1 NUT ZytoLight SPEC NUTM1 Dual Color Break Apart Probe Z-2208-200 200 µl 131

PAX3 HUP2 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe Z-2018-50/-200 50/200 µl 126 ZytoLight SPEC FOXO1/PAX3 TriCheck™ Probe Z-2185-50 50 µl 127

PAX7 HUP1 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe Z-2019-50/-200 50/200 µl 128

PDCD1LG2 PD-L2, PDL2 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe Z-2179-50/-200 50/200 µl 87

PDGFB SIS, SSV ZytoLight SPEC PDGFB Dual Color Break Apart Probe Z-2119-50/-200 50/200 µl 167 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe Z-2116-50/-200 50/200 µl 148

PDGFRA GAS9 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck™ Probe Z-2209-50 50 µl 55

PDGFRB JTK12, PDGFR1 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe Z-2197-50 50 µl 63

PHF1 MTF2L2, PCL1 ZytoLight SPEC PHF1 Dual Color Break Apart Probe Z-2215-50 50 µl 66

PIK3CA PI3K ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe Z-2140-200 200 µl 49

PML MYL, RNF71 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe Z-2113-50/-200 50/200 µl 132

PTEN MMAC1, TEP1 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe Z-2078-50/-200 50/200 µl 98

PTPRT KIAA0283 ZytoLight SPEC PTPRT/20q11 Dual Color Probe Z-2213-50 50 µl 158

RARA NR1B1, RAR ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe Z-2113-50/-200 50/200 µl 132

RB1 PPP1R130 ZytoLight SPEC RB1/13q12 Dual Color Probe Z-2165-50/-200 50/200 µl 129

RET HSCR1, CDHF12 ZytoLight SPEC RET Dual Color Break Apart Probe Z-2148-50/-200 50/200 µl 97

RICTOR AVO3, KIAA1999 ZytoLight SPEC RICTOR/5q31.1 Dual Color Probe Z-2278-200 200 µl 58

ROS1 MCF3, ROS ZytoLight SPEC ROS1 Dual Color Break Apart Probe Z-2144-50/-200 50/200 µl 68 ZytoLight SPEC ROS1/CEN 6 Dual Color Probe Z-2162-200 200 µl 69

RREB1 HNT ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

18 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 18 ILE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

RUNX1 AML1, AMLCR1 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe Z-2112-50/-200 50/200 µl 83 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe Z-2157-50/-200 50/200 µl 119

RUNX1T1 ETO, CDR, MTG8 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe Z-2112-50/-200 50/200 µl 83

SHANK3 prosap2 ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe Z-2299-50 50 µl 161

SMARCB1 BAF47 ZytoLight SPEC SMARCB1/22q12 Dual Color Probe Z-2178-50 50 µl 164

SOX2 ANOP3 ZytoLight SPEC SOX2/CEN 3 Dual Color Probe Z-2127-200 200 µl 50

SPI1 PU.1, SPI-A ZytoLight SPEC SPI1 Dual Color Break Apart Probe Z-2291-50 50 µl 104

SS18 SYT, SSXT ZytoLight SPEC SS18 Dual Color Break Apart Probe Z-2097-50/-200 50/200 µl 149 ZytoLight SPEC SS18/SSX1 TriCheck™ Probe Z-2184-50 50 µl 150

SSX1 - ZytoLight SPEC SS18/SSX1 TriCheck™ Probe Z-2184-50 50 µl 150

TERC hTERC, TRC3 ZytoLight SPEC TERC/CEN 3 Dual Color Probe Z-2284-200 200 µl 48

TERT EST2, TCS1 ZytoLight SPEC TERT Dual Color Break Apart Probe Z-2273-50 50 µl 56 ZytoLight SPEC TERT/5q31 Dual Color Probe Z-2091-50/-200 50/200 µl 57

TFE3 TFEA ZytoLight SPEC TFE3 Dual Color Break Apart Probe Z-2109-50/-200 50/200 µl 169

TMPRSS2 PRSS10 ZytoLight SPEC ERG/TMPRSS2 TriCheck™ Probe Z-2135-200 200 µl 160

TOP2A TOP2 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe Z-2093-50/-200 50/200 µl 146

TP53 LSF1, TRP53 ZytoLight SPEC TP53/17q22 Dual Color Probe Z-2198-50 50 µl 139 ZytoLight SPEC TP53/ATM Dual Color Probe Z-2159-50/-200 50/200 µl 112 ZytoLight SPEC TP53/CEN 17 Dual Color Probe Z-2153-50/-200 50/200 µl 140

USP6 Tre-2, TRE17 ZytoLight SPEC USP6 Dual Color Break Apart Probe Z-2151-50 50 µl 141

VEGFA VEGF, VPF ZytoLight SPEC VEGFA/CEN 6 Dual Color Probe Z-2195-200 200 µl 67

VHL VHL1 ZytoLight SPEC VHL/CEN 3 Dual Color Probe Z-2084-200 200 µl 42 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe Z-2102-200 200 µl 43

WT1 AWT1 ZytoLight SPEC WT1 Dual Color Break Apart Probe Z-2142-50 50 µl 103

WWTR1 TAZ ZytoLight SPEC WWTR1 Dual Color Break Apart Probe Z-2212-50 50 µl 47

YWHAE 14-3-3 epsilon ZytoLight SPEC YWHAE Dual Color Break Apart Probe Z-2175-50 50 µl 142

ZNF384 CIZ ZytoLight SPEC ZNF384 Dual Color Break Apart Probe Z-2275-50 50 µl 117

The Gene Index list includes only those probes directed against DNA sequences assigned to known genes. It does not contain probes directed against other genomic sequences as e.g. repetitive satellite DNA sequences. For a complete overview of all ZytoLight ® probes, please refer to the Chromosome Index.

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

19 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 19 ILE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Solid Tumors Brain and Neural Tumors ZytoLight Glioma 1p/19q Probe Set Z-2272-20 20 tests 26 ZytoLight SPEC 1p36/1q25 Dual Color Probe Z-2075-50/-200 50/200 µl 27 ZytoLight SPEC 19q13/19p13 Dual Color Probe Z-2076-50/-200 50/200 µl 28 ZytoLight SPEC C19MC/19p13 Dual Color Probe Z-2274-50 50 µl 156 ZytoLight SPEC CDKN2A/CEN 9 Dual Color Probe Z-2063-50/-200 50/200 µl 89 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe Z-2033-50/-200 50/200 µl 74 ZytoLight SPEC MET/CEN 7 Dual Color Probe Z-2087-50/-200 50/200 µl 76 ZytoLight SPEC MYCN/2q11 Dual Color Probe Z-2074-50/-200 50/200 µl 35 ZytoLight SPEC NTRK2 Dual Color Break Apart Probe Z-2205-50/-200 50/200 µl 91 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe Z-2078-50/-200 50/200 µl 98 ZytoLight SPEC TERT Dual Color Break Apart Probe Z-2273-50 50 µl 56 ZytoLight SPEC TP53/17q22 Dual Color Probe Z-2198-50 50 µl 139

Breast Cancer ZytoLight SPEC BCL2L1/CEN 20 Dual Color Probe Z-2171-200 200 µl 157 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe Z-2033-50/-200 50/200 µl 74 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Z-2015-50/-200 50/200 µl 143 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit Z-2020-5/-20 5/20 tests 143 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe Z-2077-50/-200 50/200 µl 144 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe Z-2190-50/-200 50/200 µl 145 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe Z-2093-50/-200 50/200 µl 146 ZytoLight SPEC ERBB3/CEN 12 Dual Color Probe Z-2056-200 200 µl 121 ZytoLight SPEC ERBB4/2q11 Dual Color Probe Z-2057-200 200 µl 41 ZytoLight SPEC ESR1/CEN 6 Dual Color Probe Z-2069-50/-200 50/200 µl 72 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe Z-2072-50/-200 50/200 µl 82 ZytoLight SPEC FGFR2/CEN 10 Dual Color Probe Z-2122-200 200 µl 100 ZytoLight SPEC MCL1/1p12 Dual Color Probe Z-2173-200 200 µl 30 ZytoLight SPEC MYC/CEN 8 Dual Color Probe Z-2092-50/-200 50/200 µl 85 ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe Z-2140-200 200 µl 49 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65 ZytoLight SPEC VEGFA/CEN 6 Dual Color Probe Z-2195-200 200 µl 67

Cervical Cancer ZytoLight SPEC MYC/CEN 8 Dual Color Probe Z-2092-50/-200 50/200 µl 85 ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe Z-2140-200 200 µl 49 ZytoLight SPEC TERC/CEN 3 Dual Color Probe Z-2284-200 200 µl 48 ZytoLight SPEC TERT/5q31 Dual Color Probe Z-2091-50/-200 50/200 µl 57

Gastrointestinal Cancer ZytoLight SPEC BRAF Dual Color Break Apart Probe Z-2189-200 200 µl 77 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Z-2015-50/-200 50/200 µl 143 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit Z-2020-5/-20 5/20 tests 143 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe Z-2077-50/-200 50/200 µl 144 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe Z-2190-50/-200 50/200 µl 145 ZytoLight SPEC KRAS/CEN 12 Dual Color Probe Z-2115-200 200 µl 120 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe Z-2013-50/-200 50/200 µl 124 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

20 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 20 ILE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Lung Cancer ZytoLight SPEC ALK/EML4 TriCheck™ Probe Z-2117-50/-200 50/200 µl 36 ZytoLight SPEC ALK Dual Color Break Apart Probe Z-2124-50/-200 50/200 µl 37 ZytoLight SPEC ALK/2q11 Dual Color Probe Z-2161-200 200 µl 38 ZytoLight SPEC BRAF/CEN 7 Dual Color Probe Z-2191-200 200 µl 78 ZytoLight SPEC CARS Dual Color Break Apart Probe Z-2137-50 50 µl 101 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe Z-2179-50/-200 50/200 µl 87 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe Z-2033-50/-200 50/200 µl 74 ZytoLight SPEC EML4 Dual Color Break Apart Probe Z-2136-50 50 µl 39 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Z-2015-50/-200 50/200 µl 143 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit Z-2020-5/-20 5/20 tests 143 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe Z-2077-50/-200 50/200 µl 144 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe Z-2190-50/-200 50/200 µl 145 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe Z-2072-50/-200 50/200 µl 82 ZytoLight SPEC FGFR2 Dual Color Break Apart Probe Z-2169-200 200 µl 99 ZytoLight SPEC FGFR2/CEN 10 Dual Color Probe Z-2122-200 200 µl 100 ZytoLight SPEC FGFR3/4p11 Dual Color Probe Z-2082-200 200 µl 53 ZytoLight SPEC KIF5B Dual Color Break Apart Probe Z-2131-50 50 µl 96 ZytoLight SPEC KRAS/CEN 12 Dual Color Probe Z-2115-200 200 µl 120 ZytoLight SPEC MET/CEN 7 Dual Color Probe Z-2087-50/-200 50/200 µl 76 ZytoLight SPEC NRG1 Dual Color Break Apart Probe Z-2181-200 200 µl 80 ZytoLight SPEC NRG1/CD74 TriCheck™ Probe Z-2194-200 200 µl 79 ZytoLight SPEC NTRK1 Dual Color Break Apart Probe Z-2167-50/-200 50/200 µl 32 ZytoLight SPEC NTRK2 Dual Color Break Apart Probe Z-2205-50/-200 50/200 µl 91 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe Z-2206-50/-200 50/200 µl 133 ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe Z-2140-200 200 µl 49 ZytoLight SPEC RET Dual Color Break Apart Probe Z-2148-50/-200 50/200 µl 97 ZytoLight SPEC RICTOR/5q31.1 Dual Color Probe Z-2278-200 200 µl 58 ZytoLight SPEC ROS1 Dual Color Break Apart Probe Z-2144-50/-200 50/200 µl 68 ZytoLight SPEC ROS1/CEN 6 Dual Color Probe Z-2162-200 200 µl 69 ZytoLight SPEC SOX2/CEN 3 Dual Color Probe Z-2127-200 200 µl 50

Prostate Cancer ZytoLight SPEC ERG Dual Color Break Apart Probe Z-2138-200 200 µl 159 ZytoLight SPEC ERG/TMPRSS2 TriCheck™ Probe Z-2135-200 200 µl 160 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe Z-2078-50/-200 50/200 µl 98 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe Z-2152-50/-200 50/200 µl 65

Renal Cell Carcinoma ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe Z-2118-200 200 µl 44 ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe Z-2081-50/-200 50/200 µl 90 ZytoLight SPEC FHIT/CEN 3 Dual Color Probe Z-2062-200 200 µl 45 ZytoLight SPEC TFE3 Dual Color Break Apart Probe Z-2109-50/-200 50/200 µl 169 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe Z-2102-200 200 µl 43 ZytoLight SPEC VHL/CEN 3 Dual Color Probe Z-2084-200 200 µl 42

Salivary Gland Tumors ZytoLight SPEC ETV6 Dual Color Break Apart Probe Z-2176-50/-200 50/200 µl 118 ZytoLight SPEC EWSR1 Dual Color Break Apart Probe Z-2096-50/-200 50/200 µl 165 ZytoLight SPEC MAML2 Dual Color Break Apart Probe Z-2014-50/-200 50/200 µl 108 ZytoLight SPEC MYB Dual Color Break Apart Probe Z-2143-50/-200 50/200 µl 70 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe Z-2206-50/-200 50/200 µl 133 ZytoLight SPEC NUTM1 Dual Color Break Apart Probe Z-2208-200 200 µl 131 ZytoLight SPEC WT1 Dual Color Break Apart Probe Z-2142-50 50 µl 103

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

21 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 21 ILE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Sarcomas ZytoLight SPEC ALK Dual Color Break Apart Probe Z-2124-50/-200 50/200 µl 37 ZytoLight SPEC CDK4/CEN 12 Dual Color Probe Z-2103-50/-200 50/200 µl 123 ZytoLight SPEC CIC Dual Color Break Apart Probe Z-2285-50 50 µl 155 ZytoLight SPEC COL1A1 Dual Color Break Apart Probe Z-2121-200 200 µl 147 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe Z-2116-50/-200 50/200 µl 148 ZytoLight SPEC DDIT3 Dual Color Break Apart Probe Z-2100-50/-200 50/200 µl 122 ZytoLight SPEC ETV6 Dual Color Break Apart Probe Z-2176-50/-200 50/200 µl 118 ZytoLight SPEC EWSR1 Dual Color Break Apart Probe Z-2096-50/-200 50/200 µl 165 ZytoLight SPEC EWSR1/FLI1 TriCheck™ Probe Z-2183-50 50 µl 166 ZytoLight SPEC FOXO1 Dual Color Break Apart Probe Z-2139-50 50 µl 125 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe Z-2018-50/-200 50/200 µl 126 ZytoLight SPEC FOXO1/PAX3 TriCheck™ Probe Z-2185-50 50 µl 127 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe Z-2019-50/-200 50/200 µl 128 ZytoLight SPEC FUS Dual Color Break Apart Probe Z-2130-50 50 µl 135 ZytoLight SPEC JAZF1 Dual Color Break Apart Probe Z-2132-50 50 µl 73 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe Z-2013-50/-200 50/200 µl 124 ZytoLight SPEC MDM4/1p12 Dual Color Probe Z-2080-200 200 µl 34 ZytoLight SPEC MYC/CEN 8 Dual Color Probe Z-2092-50/-200 50/200 µl 85 ZytoLight SPEC NR4A3 Dual Color Break Apart Probe Z-2145-50 50 µl 92 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe Z-2206-50/-200 50/200 µl 133 ZytoLight SPEC PDGFB Dual Color Break Apart Probe Z-2119-50/-200 50/200 µl 167 ZytoLight SPEC PHF1 Dual Color Break Apart Probe Z-2215-50 50 µl 66 ZytoLight SPEC SMARCB1/22q12 Dual Color Probe Z-2178-50 50 µl 164 ZytoLight SPEC SS18 Dual Color Break Apart Probe Z-2097-50/-200 50/200 µl 149 ZytoLight SPEC SS18/SSX1 TriCheck™ Probe Z-2184-50 50 µl 150 ZytoLight SPEC TFE3 Dual Color Break Apart Probe Z-2109-50/-200 50/200 µl 169 ZytoLight SPEC USP6 Dual Color Break Apart Probe Z-2151-50 50 µl 141 ZytoLight SPEC VEGFA/CEN 6 Dual Color Probe Z-2195-200 200 µl 67 ZytoLight SPEC WT1 Dual Color Break Apart Probe Z-2142-50 50 µl 103 ZytoLight SPEC WWTR1 Dual Color Break Apart Probe Z-2212-50 50 µl 47 ZytoLight SPEC YWHAE Dual Color Break Apart Probe Z-2175-50 50 µl 142

Hematology Specific Probes Acute Lymphoblastic Leukemia (ALL) ZytoLight SPEC ABL1 Dual Color Break Apart Probe Z-2199-50 50 µl 93 ZytoLight SPEC ABL2 Dual Color Break Apart Probe Z-2200-50 50 µl 33 ZytoLight SPEC CRLF2 Dual Color Break Apart Probe Z-2201-50 50 µl 168 ZytoLight SPEC CSF1R Dual Color Break Apart Probe Z-2202-50 50 µl 61 ZytoLight SPEC ETV6 Dual Color Break Apart Probe Z-2176-50/-200 50/200 µl 118 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe Z-2157-50/-200 50/200 µl 119 ZytoLight SPEC JAK2 Dual Color Break Apart Probe Z-2294-50 50 µl 88 ZytoLight SPEC KMT2A Dual Color Break Apart Probe Z-2193-50/-200 50/200 µl 116 ZytoLight SPEC MEF2D/BCL9 TriCheck™ Probe Z-2277-50 50 µl 31 ZytoLight SPEC MYB Dual Color Break Apart Probe Z-2143-50/-200 50/200 µl 70 ZytoLight SPEC NUP98 Dual Color Break Apart Probe Z-2266-50 50 µl 102 ZytoLight SPEC NUP214 Dual Color Break Apart Probe Z-2265-50 50 µl 95 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck™ Probe Z-2209-50 50 µl 55 ZytoLight SPEC SPI1 Dual Color Break Apart Probe Z-2291-50 50 µl 104 ZytoLight SPEC ZNF384 Dual Color Break Apart Probe Z-2275-50 50 µl 117

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

22 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 22 ILE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Acute Myelogenous Leukemia (AML) ZytoLight CEN 8 Probe Z-2004-50/-200 50/200 µl 170 f. ZytoLight SPEC ABL2 Dual Color Break Apart Probe Z-2200-50 50 µl 33 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe Z-2111-50/-200 50/200 µl 94 ZytoLight SPEC CBFB Dual Color Break Apart Probe Z-2207-50 50 µl 136 ZytoLight SPEC CREBBP Dual Color Break Apart Probe Z-2267-50 50 µl 134 ZytoLight SPEC CSF1R/D5S23,D5S721 Dual Color Probe Z-2268-50 50µl 62 ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe Z-2214-50 50 µl 75 ZytoLight SPEC EGR1/5p15 Dual Color Probe Z-2107-50/-200 50/200 µl 59 ZytoLight SPEC EGR1/D5S23,D5S721 Dual Color Probe Z-2211-50 50 µl 60 ZytoLight SPEC FGFR1 Dual Color Break Apart Probe Z-2168-50/-200 50/200 µl 81 ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe Z-2287-50 50 µl 46 ZytoLight SPEC KMT2A Dual Color Break Apart Probe Z-2193-50/-200 50/200 µl 116 ZytoLight SPEC NUP98 Dual Color Break Apart Probe Z-2266-50 50 µl 102 ZytoLight SPEC NUP214 Dual Color Break Apart Probe Z-2265-50 50 µl 95 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck™ Probe Z-2209-50 50 µl 55 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe Z-2197-50 50 µl 63 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe Z-2113-50/-200 50/200 µl 132 ZytoLight SPEC PTPRT/20q11 Dual Color Probe Z-2213-50 50 µl 158 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe Z-2112-50/-200 50/200 µl 83

Chronic Lymphocytic Leukemia (CLL) ZytoLight SPEC ATM/CEN 11 Dual Color Probe Z-2297-50 50 µl 110 ZytoLight SPEC ATM/CEN 12 Dual Color Probe Z-2296-50 50 µl 111 ZytoLight SPEC BCL2 Dual Color Break Apart Probe Z-2192-50/-200 50/200 µl 151 ZytoLight SPEC CCND1 Dual Color Break Apart Probe Z-2108-50/-200 50/200 µl 105 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC D13S319/13q34/CEN 12 Triple Color Probe Z-2160-50/-200 50/200 µl 113 ZytoLight SPEC D13S319/13q34 Dual Color Probe Z-2280-50 50 µl 114 ZytoLight SPEC MYB/CEN 6 Dual Color Probe Z-2281-50 50 µl 71 ZytoLight SPEC MYC/CEN 8 Dual Color Probe Z-2092-50/-200 50/200 µl 85 ZytoLight SPEC RB1/13q12 Dual Color Probe Z-2165-50/-200 50/200 µl 129 ZytoLight SPEC TP53/ATM Dual Color Probe Z-2159-50/-200 50/200 µl 112 ZytoLight SPEC TP53/CEN 17 Dual Color Probe Z-2153-50/-200 50/200 µl 140

Chronic Myelogenous Leukemia (CML) ZytoLight CEN 8 Probe Z-2004-50/-200 50/200 µl 170 f. ZytoLight SPEC ABL1 Dual Color Break Apart Probe Z-2199-50 50 µl 93 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe Z-2111-50/-200 50/200 µl 94 ZytoLight SPEC JAK2 Dual Color Break Apart Probe Z-2294-50 50 µl 88 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe Z-2197-50 50 µl 63 ZytoLight SPEC TP53/17q22 Dual Color Probe Z-2198-50 50 µl 139

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

23 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 23 ILE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Multiple Myeloma ZytoLight SPEC CCND1 Dual Color Break Apart Probe Z-2108-50/-200 50/200 µl 105 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe Z-2125-50/-200 50/200 µl 107 ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe Z-2276-50 50 µl 29 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe Z-2170-50/-200 50/200 µl 52 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe Z-2282-50 50µl 54 ZytoLight SPEC IGH Dual Color Break Apart Probe Z-2110-50/-200 50/200 µl 130 ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe Z-2270-50 50 µl 137 ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe Z-2271-50 50 µl 138 ZytoLight SPEC RB1/13q12 Dual Color Probe Z-2165-50/-200 50/200 µl 129 ZytoLight SPEC TP53/CEN 17 Dual Color Probe Z-2153-50/-200 50/200 µl 140

Myelodysplastic Syndrome (MDS) ZytoLight CEN 8 Probe Z-2004-50/-200 50/200 µl 170 f. ZytoLight SPEC CREBBP Dual Color Break Apart Probe Z-2267-50 50 µl 134 ZytoLight SPEC CSF1R/D5S23,D5S721 Dual Color Probe Z-2268-50 50 µl 62 ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe Z-2214-50 50 µl 75 ZytoLight SPEC EGR1/5p15 Dual Color Probe Z-2107-50/-200 50/200 µl 59 ZytoLight SPEC EGR1/D5S23,D5S721 Dual Color Probe Z-2211-50 50 µl 60 ZytoLight SPEC ETV6 Dual Color Break Apart Probe Z-2176-50/-200 50/200 µl 118 ZytoLight SPEC NUP98 Dual Color Break Apart Probe Z-2266-50 50 µl 102 ZytoLight SPEC NUP214 Dual Color Break Apart Probe Z-2265-50 50 µl 95 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe Z-2197-50 50 µl 63 ZytoLight SPEC PTPRT/20q11 Dual Color Probe Z-2213-50 50 µl 158 ZytoLight SPEC TERT/5q31 Dual Color Probe Z-2091-50/-200 50/200 µl 57

Non-Hodgkin Lymphoma, other ZytoLight SPEC 11q gain/loss Triple Color Probe Z-2216-50 50 µl 115 ZytoLight SPEC BCL2 Dual Color Break Apart Probe Z-2192-50/-200 50/200 µl 151 ZytoLight SPEC BCL2/CEN 18 Dual Color Probe Z-2174-50 50 µl 152 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe Z-2114-50/-200 50/200 µl 153 ZytoLight SPEC BCL6 Dual Color Break Apart Probe Z-2177-50/-200 50/200 µl 51 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe Z-2146-50/-200 50/200 µl 109 ZytoLight SPEC CCND1 Dual Color Break Apart Probe Z-2108-50/-200 50/200 µl 105 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe Z-2071-50/-200 50/200 µl 106 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe Z-2125-50/-200 50/200 µl 107 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe Z-2170-50/-200 50/200 µl 52 ZytoLight SPEC IGH Dual Color Break Apart Probe Z-2110-50/-200 50/200 µl 130 ZytoLight SPEC IGK Dual Color Break Apart Probe Z-2288-50 50 µl 40 ZytoLight SPEC IGL Dual Color Break Apart Probe Z-2286-50 50 µl 163 ZytoLight SPEC IRF4,DUSP22 Dual Color Break Apart Probe Z-2210-50 50 µl 64 ZytoLight SPEC MALT1 Dual Color Break Apart Probe Z-2196-50/-200 50/200 µl 154 ZytoLight SPEC MYC Dual Color Break Apart Probe Z-2090-50/-200 50/200 µl 84 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe Z-2105-50/-200 50/200 µl 86

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

24 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 24 ILE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Genetics Sex Mismatched Bone-Marrow Transplantant Management ZytoLight CEN X Probe Z-2008-200 200 µl 170 f. ZytoLight CEN X/Y Dual Color Probe Z-2120-200 200 µl 170 f. ZytoLight CEN X/Yq12 Dual Color Probe Z-2016-50/-200 50/200 µl 170 f. ZytoLight CEN Y (DYZ3) Probe Z-2123-200 200 µl 170 f. ZytoLight CEN Yq12 Probe Z-2010-200 200 µl 170 f.

Prenatal, Postnatal, and Preimplantation Genetics ZytoLight Aneuploidy Panel 18/X/Y and 13/21 Z-2279-20 20 tests 172 ZytoLight Aneuploidy Panel X/Y and 13/18/21 Z-2104-5/-20 5/20 tests 173 ZytoLight SPEC 13q12 Probe Z-2085-200 200 µl 170 f. ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe Z-2095-50/-200 50/200 µl 170 f. ZytoLight SPEC 13/21 Dual Color Probe Z-2164-200 200 µl 170 f. ZytoLight CEN 18 Probe Z-2007-200 200 µl 170 f. ZytoLight SPEC 18/CEN X/Y Triple Color Probe Z-2163-200 200 µl 170 f. ZytoLight SPEC 21q22 Probe Z-2086-200 200 µl 170 f. ZytoLight SPEC 21/CEN X/Yq12 Triple Color Probe Z-2180-200 200 µl 170 f. ZytoLight CEN X Probe Z-2008-200 200 µl 170 f. ZytoLight CEN X/Y Dual Color Probe Z-2120-200 200 µl 170 f. ZytoLight CEN X/Yq12 Dual Color Probe Z-2016-50/-200 50/200 µl 170 f. ZytoLight CEN Y (DYZ3) Probe Z-2123-200 200 µl 170 f. ZytoLight CEN Yq12 Probe Z-2010-200 200 µl 170 f. ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe Z-2299-50 50 µl 161 ZytoLight SPEC DiGeorge Triple Color Probe Z-2289-50 50 µl 162

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

25 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 25 ILE001-1-20 ZytoLight ® Glioma 1p/19q Probe Set

Background Probe Description Results Deletions affecting the short arm of chro- The ZytoLight ® Glioma 1p/19q Probe Set Using the SPEC 1p36/1q25 Dual Color mosome 1 (1p36) and the long arm of includes the ZytoLight ® SPEC 1p36/1q25 Probe or the SPEC 19q13/19p13 Dual chromosome 19 (19q13) are frequently Dual Color Probe and the ZytoLight ® Color Probe in a normal interphase nucle- found in human gliomas. According to the SPEC 19q13/19p13 Dual Color Probe us, two orange and two green signals are 2016 WHO criteria for classification of for the detection of both 1p36 and 19q13 expected. In a cell with deletions affecting tumors of the central nervous system, the loci, and the innovative ZyBlack™ Quench- the 1p36 or 19q13 locus, one or no copy detection of 1p/19q loss is required for ing Solution to reduce autofluorescence of the orange signal will be observed. the diagnosis of WHO grade II or III “oli- on both formalin-fixed paraffin-embedded godendroglioma, IDH-mutant and 1p/19q and frozen sections. codeleted”. Since both, astrocytomas and oligodendrogliomas, can exhibit IDH mu- ZytoLight ® SPEC 1p36/1q25 Dual Color Probe tations, evaluation of 1p/19q status plays D1S1384 RH50089 a critical role in differentiating astrocyto- 3’ 5’ ma from oligodendroglioma. CHD5 Oligodendroglioma morphology, IDH-mu- ~365 kb tant genotype, and 1p/19q codeletion Tel 1p36.31 Cen are associated with better response to SPEC 1p36 Probe map (not to scale). chemotherapy and improved survival. Hence, determination of 1p and 19q sta- RH77882 STSG34156 tus may aid in therapeutic decisions and predict outcome in patients with diffuse

gliomas. ~715 kb Cen 1q25.3 Tel Brain tissue section hybridized with the Several types of tissue tend to emit intense ZytoLight ® SPEC 1p36/1q25 Dual Color Probe autofluorescence including brain, liver, SPEC 1q25 Probe map (not to scale). without ZyBlack™ Quenching Solution. kidney and myocardium, making it difficult to evaluate FISH results. The ZyBlack™ ZytoLight ® SPEC 19q13/19p13 Dual Color Probe Quenching Solution reduces autofluores- RH55727 RH120143 cence without adversely affecting tissue integrity or specific fluorescence signals.

References Barbashina V, et al. (2005) Clin Cancer Res 11: 1119-28. ~485 kb Cairncross JG, et al. (1998) J Natl Cancer Inst 90: 1473-9. Tel 19p13.3 Cen Cairncross G, et al. (2013) J Clin Oncol 31: 337-43. Griffin CA, et al. (2006) J Neuropathol Exp Neurol 65: 988-94. Louis DN, et al. (ed.) (2016) WHO Classification of Tumours of the SPEC 19p13 Probe map (not to scale). Central Nervous System (Revised 4th Edition). Reifenberger G, et al. (2017) Nat Rev Clin Oncol 14: 434-52. Rosenberg JE, et al. (1996) Oncogene 13: 2483-5. RH68396 RH14160 Smith JS, et al. (1999) Oncogene 18: 4144-52. 5’ 3’ 5’3’ 5’ 3’ Smith JS, et al. (2000) Genes Chromosomes Cancer 29: 16-25.

GLTSCR1 GLTSCR2 CRX

~515 kb Cen 19q13.32-q13.33 Tel Brain tissue section hybridized with the ZytoLight ® SPEC 1p36/1q25 Dual Color Probe SPEC 19q13 Probe map (not to scale). with ZyBlack™ Quenching Solution.

Prod. No. Product Label Tests* (Volume) Z-2272-20 ZytoLight Glioma 1p/19q Probe Set 20 Incl. ZytoLight SPEC 1p36/1q25 Dual Color Probe, 0.2 ml; ZytoLight SPEC 19q13/19p13 Dual Color Probe, 0.2 ml; ZyBlack Quenching Solution, 8 ml Related Products Z-2075-200 ZytoLight SPEC 1p36/1q25 Dual Color Probe / 20 (200 μl) Z-2076-200 ZytoLight SPEC 19q13/19p13 Dual Color Probe / 20 (200 μl) Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

26 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 26 FE149-1-20 ZytoLight ® SPEC 1p36/1q25 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC 1p36/1q25 Dual The SPEC 1p36/1q25 Dual Color Probe Using the SPEC 1p36/1q25 Dual Color Color Probe is designed for the detection is a mixture of an orange fluorochrome Probe in a normal interphase nucleus, two of 1p deletions. direct labeled 1p36 probe specific for the orange and two green signals are expect- Deletions affecting the short arm of chro- smallest region of consistent deletion (SRD) ed. In a cell with deletions affecting the mosome 1 (1p) are frequently found in of chromosome 1 defined in neuroblasto- 1p36 locus, one or no copy of the orange human gliomas and neuroblastomas, but ma at 1p36.31 and a green fluorochrome signal will be observed. also in breast, lung, endometrial, ovarian, direct labeled 1q25 probe specific for and colorectal carcinomas. 1q25.3. Deletions affecting the long arm of chromosome 19 (19q) are frequently found in human malignant gliomas as well as in neuroblastomas and epithelial ovarian cancers. Combined loss of the complete 1p/19q chromosome arms, caused by an unbalanced t(1;19)(q10;p10) translocation, is characteristic of oligodendrogliomas. According to the 2016 WHO criteria

for classification of tumors of the central SPEC 1p36/1q25 Dual Color Probe hybridized nervous system, the detection of 1p/19q to a glioma tissue section with 1p36 deletion as indicated by one orange signal in each nucleus. loss is required for the diagnosis of WHO grade II or III “oligodendroglioma, Ideogram of chromosome 1 IDH-mutant and 1p/19q codeleted”. Since indicating the hybridization locations. both, astrocytomas and oligodendroglio- D1S1384 RH50089 mas, can exhibit IDH mutations, evaluation 3’ 5’ of 1p/19q status plays a critical role in CHD5 differentiating astrocytoma from oligoden- ~365 kb droglioma. Tel 1p36.31 Cen Oligodendroglioma morphology, IDH-mu- SPEC 1p36 Probe map (not to scale). tant genotype, and 1p/19q codeletion References RH77882 STSG34156 Barbashina V, et al. (2005) Clin Cancer Res 11: 1119-28. are associated with better response to Cairncross JG, et al. (1998) J Natl Cancer Inst 90: 1473-9. Cairncross G, et al. (2013) J Clin Oncol 31: 337-43. chemotherapy and improved survival. Caron H, et al. (1996) N Engl J Med 334: 225-30. Griffin CA, et al. (2006) J Neuropathol Exp Neurol 65: 988-94. Hence, determination of 1p and 19q sta- Louis DN, et al. (ed.) (2016) WHO Classification of Tumours of the ~715 kb Central Nervous System (Revised 4th Edition). tus may aid in therapeutic decisions and Ragnarsson G, et al. (1999) Br J Cancer 79: 1468-74. Cen 1q25.3 Tel Reifenberger G, et al. (2017) Nat Rev Clin Oncol 14: 434-52. predict outcome in patients with diffuse Rosenberg JE, et al. (1996) Oncogene 13: 2483-5. SPEC 1q25 Probe map (not to scale). Smith JS, et al. (1999) Oncogene 18: 4144-52. gliomas. Smith JS, et al. (2000) Genes Chromosomes Cancer 29: 16-25.

Prod. No. Product Label Tests* (Volume) Z-2075-50 ZytoLight SPEC 1p36/1q25 Dual Color Probe / 5 (50 μl) Z-2075-200 ZytoLight SPEC 1p36/1q25 Dual Color Probe / 20 (200 μl) Related Products Z-2272-20 ZytoLight Glioma 1p/19q Probe Set 20 Incl. ZytoLight SPEC 1p36/1q25 Dual Color Probe, 0.2 ml; ZytoLight SPEC 19q13/19p13 Dual Color Probe, 0.2 ml; ZyBlack Quenching Solution, 8 ml Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

27 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 27 FE021-1-20 ZytoLight ® SPEC 19q13/19p13 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC 19q13/19p13 Dual The SPEC 19q13/19p13 Dual Color Using the SPEC 19q13/19p13 Dual Color Probe is designed for the detection Probe is a mixture of an orange fluo- Color Probe in a normal interphase nucle- of 19q deletions. rochrome direct labeled 19q13 probe us, two orange and two green signals are Deletions affecting the long arm of chro- specific for the region of common deletion expected. In a cell with deletions affecting mosome 19 (19q) are frequently found in gliomas at 19q13.32-q13.33 and a the 19q13 locus, one or no copy of the in human malignant gliomas as well as green fluorochrome direct labeled 19p13 orange signal will be observed. in neuroblastomas and epithelial ovarian probe specific for 19p13.3. cancers. Deletions affecting the short arm of chromosome 1 (1p) are frequently found in human gliomas and neuroblastomas, but also in breast, lung, endometrial, ovarian, Ideogram of chromosome 19 and colorectal carcinomas. indicating the hybridization locations. Combined loss of the complete 1p/19q RH55727 RH120143 chromosome arms, caused by an unbalanced t(1;19)(q10;p10) translocation, is characteristic of oligodendrogliomas. ~485 kb According to the 2016 WHO criteria Tel 19p13.3 Cen for classification of tumors of the central SPEC 19p13 Probe map (not to scale). SPEC 19q13/19p13 Dual Color Probe hybridized to nervous system, the detection of 1p/19q a glioma tissue section with 19q13 deletion as indicated by one orange signal in each nucleus. RH68396 RH14160 loss is required for the diagnosis of 5’ 3’ 5’3’ 5’ 3’ WHO grade II or III “oligodendroglioma, IDH-mutant and 1p/19q codeleted”. Since GLTSCR1 GLTSCR2 CRX both, astrocytomas and oligodendroglio- ~515 kb Cen 19q13.32-q13.33 Tel mas, can exhibit IDH mutations, evaluation of 1p/19q status plays a critical role in SPEC 19q13 Probe map (not to scale). differentiating astrocytoma from oligodendroglioma. Oligodendroglioma morphology, IDH-mu-

tant genotype, and 1p/19q codeletion References Barbashina V, et al. (2005) Clin Cancer Res 11: 1119-28. are associated with better response to Cairncross JG, et al. (1998) J Natl Cancer Inst 90: 1473-9. Cairncross G, et al. (2013) J Clin Oncol 31: 337-43. chemotherapy and improved survival. Caron H, et al. (1996) N Engl J Med 334: 225-30. Griffin CA, et al. (2006) J Neuropathol Exp Neurol 65: 988-94. Hence, determination of 1p and 19q sta- Louis DN, et al. (ed.) (2016) WHO Classification of Tumours of the Central Nervous System (Revised 4th Edition). tus may aid in therapeutic decisions and Ragnarsson G, et al. (1999) Br J Cancer 79: 1468-74. Reifenberger G, et al. (2017) Nat Rev Clin Oncol 14: 434-52. predict outcome in patients with diffuse Rosenberg JE, et al. (1996) Oncogene 13: 2483-5. Smith JS, et al. (1999) Oncogene 18: 4144-52. gliomas. Smith JS, et al. (2000) Genes Chromosomes Cancer 29: 16-25.

Prod. No. Product Label Tests* (Volume) Z-2076-50 ZytoLight SPEC 19q13/19p13 Dual Color Probe / 5 (50 μl) Z-2076-200 ZytoLight SPEC 19q13/19p13 Dual Color Probe / 20 (200 μl) Related Products Z-2272-20 ZytoLight Glioma 1p/19q Probe Set 20 Incl. ZytoLight SPEC 1p36/1q25 Dual Color Probe, 0.2 ml; ZytoLight SPEC 19q13/19p13 Dual Color Probe, 0.2 ml; ZyBlack Quenching Solution, 8 ml Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

28 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 28 FE125-1-20 ZytoLight ® SPEC CKS1B/CDKN2C Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC CKS1B/CDKN2C The SPEC CKS1B/CDKN2C Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- ange and two green signals are expected. detection of gains/amplifications affecting chrome direct labeled SPEC CKS1B probe In a cell with a gain/amplification of the the chromosomal region 1q21.3-q22 hybridizing to the chromosomal region CKS1B gene locus, multiple copies of the (CKS1B) and/or deletions of the chromo- 1q21.3-q22 and a green labeled SPEC orange signal or orange signal clusters somal region 1p32.2 (CDKN2C). CDKN2C probe hybridizing to the chro- will be observed. Chromosome 1 abnormalities are among mosomal region 1p32.2. In a cell with deletion of the CDKN2C the most common cytogenetic findings in gene locus one or no copy of the green multiple myeloma (MM). This B-cell malig- signal will be observed. Deletions affect- nancy is characterized by slow prolifera- ing only parts of the CDKN2C locus might tion of malignant plasma cells localized CDKN2C result in a normal signal pattern with primarily in the bone marrow. Copy green signals of reduced size. number alterations (CNA) occur frequently in this entity. The short arm of chromosome 1 is often affected by deletions whereas the long arm is mainly affected by gains/ CKS1B amplifications. The CKS1B gene is located on the long arm of chromosome 1 at 1q21. Tandem duplications and jumping translocations of the 1q21 band are acquired during Ideogram of chromosome 1 myeloma disease progression, and 1q indicating the hybridization locations. amplifications have been linked to a poor D1S427 STSG32509FS SPEC CKS1B/CDKN2C Dual Color Probe prognosis in MM patients. 5’ 3’ hybridized to normal interphase cells as indicated The CDKN2C gene maps to the chromo- by two orange and two green signals and to CDKN2C metaphase chromosomes of a normal cell. somal region 1p32.2 and belongs to the ~380 kb INK4 gene family that consists of tumor Tel 1p32.2 Cen suppressor genes which play a role in the SPEC CDKN2C Probe map (not to scale). regulation of cell proliferation. SHGC-145644 RH49721 MM patients harboring a deletion of the 5’ 3’ CDKN2C gene region have a worse overall survival compared to those lacking CKS1B this alteration. ~425 kb Cen 1q21.3-q22 Tel Hence, Fluorescence in situ Hybridization may be a helpful tool for diagnosis and SPEC CKS1B Probe map (not to scale). therapy decisions. References Bone marrow smear of a pediatric ALL case with Chang H, et al. (2010) Bone Marrow Transplant 45: 117-21. amplification affecting the CKS1B locus as indicated Kulkarni MS, et al. (2002) Leukemia 16: 127-34. Shaughnessy J, et al. (2005) Hematology 10: 117-26. by three or more orange signals. Walker BA, et al. (2010) Blood 116: 56-65. Zhan F, et al. (2007) Blood 109: 4995-5001. Material kindly provided by Paediatric Oncology/ Haematology, Charieté – Universitätsmedizin Berlin.

Prod. No. Product Label Tests* (Volume) Z-2276-50 ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

Incl. Cytology Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 50 ml; 10x MgCl2, 50 ml; 10x PBS, 50 ml; Cytology Stringency Wash Buffer SSC, 500 ml; Cytology Wash Buffer SSC, 500 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

29 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 29 FE146-1-20 ZytoLight ® SPEC MCL1/1p12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MCL1/1p12 Dual The SPEC MCL1/1p12 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of a green fluorochrome direct ange and two green signals are expected. of MCL1 gene amplifications. labeled SPEC MCL1 probe hybridizing to In a cell with amplification of the MCL1 The MCL1 (MCL1, BCL2 family apoptosis the MCL1 gene in the chromosomal region gene locus, multiple copies of the green regulator, a.k.a. BCL2L3) gene is located 1q21.3 and an orange fluorochrome signal or green signal clusters will be in the chromosomal region 1q21.3 and direct labeled SPEC 1p12 probe specific observed. encodes for an anti-apoptotic protein that for the chromosomal region 1p12. Due to belongs to the BCL2 family. These genes cross-hybridizations of chromosome 1 al- are involved in a wide variety of cellular pha satellites to other centromeric regions, activities including lymphocyte develop- probes specific for 1p12 are frequently ment and hematopoiesis. used for chromosome 1 copy number MCL1 amplifications have been reported detection. in several human cancers including bladder, gastric, ovarian, lung, breast, melanoma, and hematologic malignancies. Overexpression of MCL1 reduces MYC-induced apoptosis in immortalized bronchial epithelial cells. Furthermore, MCL1 amplifications are 1p12 H2110 cell line with interphase cells showing amplifi- found in many tumor cell lines with resist- cation of the MCL1 gene locus as indicated by multiple green signals in each nucleus. ance to chemotherapeutic agents. How- MCL1 ever, many MCL1 amplified cell lines are sensitive to treatment with the cyclin-dependent kinase (CDK) inhibitor dinaciclib. Targeting the BCL2 family proteins with small non-peptidic compounds, so called Ideogram of chromosome 1 BH3-mimetics, is currently investigated in indicating the hybridization locations. clinical trials. D1S2342E LASS2 Hence, the identification of MCL1 amplifi- 3’5’ cations by Fluorescence in situ Hybridiza- MCL1 tion and the inhibition of MCL1 signaling ~575 kb may be of therapeutic significance in Cen 1q21.3 Tel various types of tumors. SPEC MCL1 Probe map (not to scale). References Beroukhim R, et al. (2010) Nature 463: 899-905. Booher RN, et al. (2014) PloS One 9: e108371. SHGC-141884 RH105871 Sochalska M, et al. (2015) FEBS J 282: 834-49. 3’ 5’ 5’ 3’ Yasui K, et al. (2004) Cancer Res 64: 1403-10. WARS2 HAO2

~285 kb Tel 1p12 Cen

SPEC 1p12 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2173-200 ZytoLight SPEC MCL1/1p12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

30 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 30 FE108-1-20 ZytoLight ® SPEC MEF2D/BCL9 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC MEF2D/BCL9 The SPEC MEF2D/BCL9 TriCheck™ Probe In an interphase nucleus without re- TriCheck™ Probe is designed to detect in- is a mixture of three direct labeled probes arrangement of the MEF2D/BCL9 locus, versions involving the chromosomal region hybridizing to the long arm of chromo- two green/orange fusion signals and two 1q21.2 harboring the BCL9 gene and the some 1. The green fluorochrome direct blue signals are expected. chromosomal region 1q22 harboring the labeled probe hybridizes proximal to the A MEF2D-BCL9 inversion is indicated by one MEF2D gene. Moreover, using this probe MEF2D gene breakpoint region at 1q22, separate green signal, one separate orange it is possible to discriminate between the orange fluorochrome direct labeled signal, and an additional blue signal. MEF2D-BCL9 inversions and MEF2D trans- probe hybridizes distal to the MEF2D A MEF2D translocation, without BCL9 locations not affecting BCL9. gene breakpoint region at 1q22-q23.1, involvement, is indicated by one separate Rearrangements of the myocyte enhancer and the blue fluorochrome direct labeled green signal and one separate orange sig- factor 2D (MEF2D) have been frequently probe hybridizes to the BCL9 gene region nal, without an additional blue signal. found in acute lymphoblastic leukemia at 1q21.1-q21.2. Gain of the aberrant region may be ob- (ALL). Recurring rearrangements have been served and is indicated by multiple copies of found in 3-4% of pediatric and up to 7% of the respective signal pattern. adult ALL patients, respectively. In B-pro- Signal patterns other than those described genitor ALL cases the most common trans- above may indicate deviant rearrangements. location partner of MEF2D is BCL9. Other known translocation partners are CSF1R (5q32), DAZAP1 (19p13.3), HNRNPUL1 (19q13.2), SS18 (18q11.2), and FOXJ2 BCL9 (12p13.31). ALL cases harboring MEF2D MEF2D rearrangements are often associated with copy number alterations of the aberrant locus and display an increased sensitivity to histone deacetylase inhibitor (HDAC) treatment. MEF2D gene translocation de- Ideogram of chromosome 1 rived ALL cases show a markedly high ex- indicating the hybridization locations. pression of HDAC9 inducing resistance to RH121816 SHGC-142538 conventional chemotherapy and in case of SHGC-148333 SHGC-146655 SPEC MEF2D/BCL9 TriCheck™ Probe on 3’ 5’ MEF2D-BCL9 gene fusion also a resistance normal interphase cells with non-rearranged MEF2D loci (two green/orange fusion signals), to dexamethasone treatment. MEF2D-re- MEF2D and non-rearranged BCL9 loci (two blue signals). arranged ALL represents a distinct form of ~625 kb ~510 kb high-risk leukemia; consequently MEF2D- Cen 1q22-q23.1 Tel BCL9 fusion detection by Fluorescence SPEC MEF2D Probe map (not to scale). in situ Hybridization (FISH) might be of RH112365 RH94160 diagnostic and therapeutic relevance. 5’ 3’

References Gu Z, et al. (2016) Nat Commun 7: 13331. BCL9 Liu YF, et al. (2016) EBioMedicine 8: 173-83. Suzuki K, et al. (2016) J Clin Oncol 34: 3451-9. ~800 kb Cen 1q21.1-q21.2 Tel

SPEC BCL9 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2277-50 ZytoLight SPEC MEF2D/BCL9 TriCheck Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

31 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 31 FE148-1-20 ZytoLight ® SPEC NTRK1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NTRK1 Dual Color The SPEC NTRK1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 1q22-q23.1 band, translocations involving the chromosomal probes hybridizing to the 1q22-q23.1 two orange/green fusion signals are ex- region 1q23.1 harboring the NTRK1 band. The green fluorochrome direct la- pected representing two normal (non-rear- (neurotrophic receptor tyrosine kinase 1, beled probe hybridizes proximal and the ranged) 1q22-q23.1 loci. A signal pattern a.k.a. TRKA or TRK) gene. orange fluorochrome direct labeled probe consisting of one orange/green fusion NTRK1 encodes a tyrosine kinase (TK) re- hybridizes distal to the NTRK1 gene. signal, one orange signal, and a sepa- ceptor for the nerve growth factor (NGF). rate green signal indicates one normal The NTRK1 gene was found to be rear- 1q22-q23.1 locus and one 1q22-q23.1 ranged in about 12% of papillary thyroid locus affected by a translocation. carcinoma (PTC) cases. PTC accounts for about 80% of all thyroid cancers. NTRK1 rearrangements result in the fusion of the 3‘ end of the NTRK1 gene with the 5‘ end of different activating genes (TPM3, TPR, or TFG). All these fusion genes encode NTRK1 hybrid proteins comprising the TK domain of NTRK1 and the N-terminus of the partner proteins carrying coiled-coil domains. NTRK1 rearrangements were shown to be involved in thyroid carcinogenesis. Ideogram of chromosome 1 Several studies showed that NTRK1 indicating the hybridization locations. rearrangements may be associated with a SPEC NTRK1 Dual Color Break Apart Probe hybridi- RH47613 STSG28995 worse clinical course when compared with zed to normal interphase cells as indicated by RH38210 RH45945 two orange/green fusion signals per nucleus. NTRK1 rearrangement-negative PTCs. 5’ 3’

Recently, NTRK1 rearrangements were NTRK1

also found in lung adenocarcinomas. ~535 kb ~440 kb Various inhibitors targeting the NTRK1-de- Cen 1q22-q23.1 Tel

rived fusion proteins were shown in vitro SPEC NTRK1 Probe map (not to scale). to inhibit proliferation of cells expressing the fusion genes. This indicates that these fusion genes are potential therapeutic targets. Hence, detection of NTRK1 rearrangements by Fluorescence in situ Hybridiza- References Alberti L, et al. (2003) J Cell Physiol 195: 168-86. tion represents a useful tool for studying Bongarzone I, et al. (1998) Clin Cancer Res 4: 223-8. Doebele RC, et al. (2013) J Clin Oncol 31 Suppl: Abstr. 8023. thyroid carcinogenesis and may be of Greco A, et al. (2010) Mol Cell Endocrinol 321: 44-9. Musholt TJ (2000) Surgery 128: 984-93. Lung cancer tissue section with translocation prognostic and therapeutic significance. Russell JP, et al. (2000) Oncogene 19: 5729-35. of the NTRK1 gene as indicated by one non-rearranged orange/green fusion signal, one orange and one separate green signal.

Image kindly provided by Prof. Büttner, Cologne, Germany.

Prod. No. Product Label Tests* (Volume) Z-2167-50 ZytoLight SPEC NTRK1 Dual Color Break Apart Probe / 5 (50 μl) Z-2167-200 ZytoLight SPEC NTRK1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

32 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 32 FE077-1-20 ZytoLight ® SPEC ABL2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ABL2 Dual Color The SPEC ABL2 Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving the rearrangements involving the chromosom- probes hybridizing to the 1q25.2 band. 1q25.2 band, two orange/green fusion al region 1q25.2 harboring the ABL2 (ABL The orange fluorochrome direct labeled signals are expected representing two proto-oncogene 2, non-receptor tyrosine probe hybridizes proximal to the ABL2 normal (non-rearranged) 1q25.2 loci. A kinase, a.k.a. ARG) gene. gene at 1q25.2, the green fluorochrome signal pattern consisting of one orange/ The ABL2 gene encodes for a non-receptor direct labeled probe hybridizes distal to green fusion signal, one orange signal, tyrosine kinase (TK) with high homology the ABL2 gene at 1q25.2 and a separate green signal indicates one to ABL1. ABL1 and ABL2 proteins belong normal 1q25.2 locus and one 1q25.2 to the Abelson family and link diverse locus affected by a translocation. extracellular stimuli to signaling pathways controlling cell growth, survival, invasion, and migration. The translocation t(1;12)(q25.2;p13.2) involving ABL2 was shown to result in a chimeric protein consisting of the helix-loop-helix (HLH) domain of ETV6 and the TK domain of ABL2. The HLH domain ABL2 of ETV6 is known to confer oncogenic activity to chimeric tyrosine kinase proteins by forming ligand-independent oligomers. The ETV6-ABL2 fusion gene has been Ideogram of chromosome 1 detected in a patient with AML-M3 and in indicating the hybridization locations. SPEC ABL2 Dual Color Break Apart Probe a T-cell ALL cell line. hybridized to normal interphase cells as indicated RH42831 RH50182 by two orange/green fusion signals per nucleus Further ABL2 fusion partners have been SHGC-141303 RH13275 and to metaphase chromosomes of a normal cell. identified in patients with Philadelphia 3’5’

chromosome-like ALL, including PAG1, ABL2

RCSD1, and ZC3HAV1. Cell lines express- ~520 kb ~520 kb ing ABL2 fusions were shown to respond Cen 1q25.2 Tel

to tyrosine kinase inhibitors. Moreover, a SPEC ABL2 Probe map (not to scale). patient with B-ALL positive for RCSD1-ABL2 fusion was reported to respond to treatment with the ABL1 inhibitor imatinib. Hence, detection of ABL2 rearrangements by FISH may help in selecting patients eligible for therapy with TK inhibitors.

References Cazzaniga G, et al. (1999) Blood 94: 4370-3. De Braekeleer E, et al. (2012) Leuk Res 36: 945-61. Blood smear with translocation of the ABL2 Greuber EK, et al. (2013) Nat Rev Cancer 13: 559-71. Roberts KG, et al. (2014) N Engl J Med 371: 1005-15. gene as indicated by one non-rearranged orange/green fusion signal, one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2200-50 ZytoLight SPEC ABL2 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

33 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 33 FE101-1-20 ZytoLight ® SPEC MDM4/1p12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MDM4/1p12 Dual The SPEC MDM4/1p12 Dual Color Probe In a normal interphase nucleus two orange Color Probe is designed for the detection is a mixture of a green fluorochrome direct and two green signals are expected. Nu- of MDM4 gene amplifications found in labeled SPEC MDM4 probe hybridizing clei with amplification of the MDM4 gene 10-20% of various tumors such as lung, distal and proximal to the human MDM4 locus or aneuploidy of chromosome 1 will colon, stomach, and breast cancers, as gene in the chromosomal region 1q32.1 show multiple copies of the green signal well as in 65% of retinoblastomas. and an orange fluorochrome direct or large green signal clusters. The MDM4 (MDM4, p53 regulator) gene labeled SPEC 1p12 probe hybridizing (a.k.a. HDMX or MDMX) is located in in close proximity to the centromere of the chromosomal region 1q32.1 and chromosome 1 at the chromosomal region encodes a 490-amino acid protein which 1p12. Due to cross-hybridizations of shows significant structural similarity to the chromosome 1 alpha satellites to other p53-binding protein MDM2. centromeric regions, probes specific for Like MDM2, the oncogene MDM4 can 1p12 are frequently used for chromosome bind to p53 thereby inactivating the func- 1 copy number detection. tion of p53 as a transcriptional activator. In addition, MDM4 has been shown to bind to MDM2 resulting in inhibition of MDM2 degradation. Antitumor strategies employing combined SPEC MDM4/1p12 Dual Color Probe hybridized inhibitors of the two oncogenic proteins to normal interphase cells as indicated by two orange and two green signals in each nucleus. MDM2 and MDM4 may lead to an effective activation of the tumor suppressor p53.

References Duhamel LA, et al. (2012) Histopathology 60: 357-9. Laurie NA, et al. (2006) Nature 444: 61-6. Shvarts A, et al. (1996) EMBO J 15: 5349-57. Shvarts A, et al. (1997) Genomics 43: 34-42. Tanimura S et al. (1999) FEBS Lett 447: 5-9. Toledo F & Wahl GM (2006) Nat Rev Cancer 6: 909-23. Toledo F & Wahl GM (2007) Int J Biochem Cell Biol 39: 1476-82. Ideogram of chromosome 1 indicating the hybridization locations.

SHGC-141884 RH105871 3’ 5’ 5’ 3’

WARS2 HAO2

~285 kb Tel 1p12 Cen

SPEC 1p12 Probe map (not to scale).

RH11064 D1S2326E 5’ 3’

MDM4

~755 kb Cen 1q32.1 Tel

SPEC MDM4 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2080-200 ZytoLight SPEC MDM4/1p12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

34 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 34 FE027-1-20 ZytoLight ® SPEC MYCN/2q11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MYCN/2q11 Dual The SPEC MYCN/2q11 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of a green fluorochrome direct ange and two green signals are expected. of MYCN amplification which represents labeled SPEC MYCN probe hybridizing to In a cell with amplification of the MYCN the most powerful unfavorable prognostic the human MYCN gene in the chromosom- gene locus, multiple copies of the green factor for neuroblastoma. Less frequently al region 2p24.3 and an orange fluoro- signal or green signal clusters will be amplifications are found in retinoblastoma, chrome direct labeled SPEC 2q11 probe observed. small cell lung cancer, astrocytoma and specific for the AFF3 (AF4/FMR2 family, other tumors derived from the neuroecto- member 3) gene region in 2q11.2. Due to derm. cross-hybridizations of chromosome 2 al- The MYCN (MYCN proto-oncogene, pha satellites to other centromeric regions, bHLH transcription factor, a.k.a. NMYC) probes specific for 2q11 are frequently gene is located in the chromosomal region used for chromosome 2 copy number 2p24.3 and encodes a 62-64 kDa tran- detection. scription factor mainly expressed in the developing nervous system. MYCN Amplification of the MYCN gene is found in about 25% of primary neuroblastomas and is strongly associated with rapid tumor progression, advanced stages of the SPEC MYCN/2q11 Dual Color Probe hybridized to normal interphase cells as indicated by two orange disease, and poor prognosis. Hence, am- and two green signals in each nucleus. plification status is increasingly being used for stratification of patients to different treatment protocols.

References Gessi M, et al. (2014) Neuro Oncol 16: 924-32. Kaneko M, et al. (1998) Med Pediatr Oncol 31: 1-7. Lee WH, et al. (1984) Nature 309: 458-60. Slamon DJ, et al. (1986) Science 232: 768-72. Suita S, et al. (2007) J Pediatr Surg 42: 489-93. Ideogram of chromosome 2 indicating the hybridization locations.

SHGC-140713 D2S2295 5’3’

MYCN

~670 kb Tel 2p24.3 Cen

SPEC MYCN Probe map (not to scale).

D2S2776 RH121214 3’ 5’

AFF3

~490 kb Cen 2q11.2 Tel

SPEC 2q11 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2074-50 ZytoLight SPEC MYCN/2q11 Dual Color Probe / 5 (50 μl) Z-2074-200 ZytoLight SPEC MYCN/2q11 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

35 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 35 FE019-1-20 ZytoLight ® SPEC ALK/EML4 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC ALK/EML4 TriCheck™ The SPEC ALK/EML4 TriCheck™ Probe is In an interphase nucleus without rear- Probe is designed to detect inversions a mixture of three direct labeled probes rangement of the EML4-ALK locus, two involving the chromosomal region hybridizing to the short arm of chromo- orange/green fusion signals and two blue 2p23.1-p23.2 harboring the ALK gene some 2. The orange fluorochrome direct signals are expected. and the chromosomal region 2p21 har- labeled probe hybridizes distal to the ALK An EML4-ALK inversion is indicated by one boring the EML4 gene. Moreover, using gene breakpoint region at 2p23.2, the separate green signal, one separate orange this probe it is possible to discriminate green fluorochrome direct labeled probe signal, and an additional blue signal. between EML4-ALK inversions and translo- hybridizes proximal to the ALK gene An ALK translocation is indicated by sepa- cations affecting ALK, but not EML4, such breakpoint region at 2p23.1-p23.2, and rated orange and green signals without an as ALK-TFG or ALK-KIF5B translocations. the blue fluorochrome direct labeled probe additional blue signal. Inversions in the short arm of chromosome hybridizes to the EML4 gene region at EML4-ALK inversion with deletion of 5’- 2 [inv(2)(p21p23)] have been frequently 2p21. ALK sequences is indicated by loss of one detected in non-small cell lung cancer green signal and co-localization of the (NSCLC) and lead to the formation of isolated orange signal with a blue signal. EML4-ALK fusion transcripts. A few reports also identified EML4-ALK fusion transcripts in breast, gastric, and colorectal cancers. Many different breakpoints affecting ALK and EML4 were identified in these respective inversions. Thus, multiple EML4-ALK transcript variants have been identified, all of which involve the intracellular kinase domain of ALK. ALK kinase targeted therapies may represent a very effective therapeutic strategy Ideogram of chromosome 2 SPEC ALK/EML4 TriCheck™ Probe on normal in NSCLC patients carrying EML4-ALK indicating the hybridization locations. interphase cells with non-rearranged ALK loci rearrangements. For the detection of this (two orange/green fusion signals), and non- rearranged EML4 loci (two blue signals). subset of NSCLC patients, the specific de- D2S2934 D2S405 RH12489 SHGC-142979 tection of EML4-ALK rearrangements using 3’ 5’ Fluorescence in situ Hybridization is a ALK helpful tool for diagnosis and for selecting ~210 kb ~635 kb treatment. Tel 2p23.1-p23.2 Cen References Inamura K, et al. (2009) Mod Pathol 22: 508-15. SPEC ALK Probe map (not to scale). Koivunen JP, et al. (2008) Clin Cancer Res 14: 4275-83. Lawce HJ & Olson S (2013) J Assoc Genet Technol 39: 66-71. SHGC-105957 D2S1583E Martelli MP, et al. (2009) Am J Pathol 174: 661-70. 5’ 3’ Perner S, et al. (2008) Neoplasia 10: 298-302. Preusser M, et al. (2013) Lung Cancer 80: 278-83. Rodig SJ, et al. (2009) Clin Cancer Res 15: 5216-23. EML4 Sasaki T, et al. (2010) Eur J Cancer 46: 1773-80. Schildgen V, et al. (2012) Per Med 9: 801-3. ~1.8 Mb Schildhaus HU, et al. (2013) Mod Pathol 26: 1468-77. Schoppmann SF, et al. (2013) Eur J Cancer 49: 1876-81. Tel 2p21 Cen Thunnissen E, et al. (2012) Virchows Arch 461: 245-57. NSCLC tissue section with an EML4-ALK inversion Von Laffert M, et al. (2013) Lung Cancer 81: 200-6. SPEC EML4 Probe map (not to scale). as indicated by one green, one separated orange, and one additional blue signal.

Prod. No. Product Label Tests* (Volume) Z-2117-50 ZytoLight SPEC ALK/EML4 TriCheck Probe •/•/• 5 (50 μl) Z-2117-200 ZytoLight SPEC ALK/EML4 TriCheck Probe •/•/• 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

36 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 36 FE039-1-20 ZytoLight ® SPEC ALK Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ALK Dual Color The SPEC ALK Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving rearrangements involving the chromosom- probes hybridizing to the 2p23.1-p23.2 the 2p23.1-p23.2 band, two orange/ al region 2p23.1-p23.2 harboring the band. The orange fluorochrome direct green fusion signals are expected rep- ALK (ALK receptor tyrosine kinase, a.k.a. labeled probe hybridizes distal to the ALK resenting two normal (non-rearranged) CD246) gene. gene breakpoint region at 2p23.2, the 2p23.1-p23.2 loci. A signal pattern con- ALK encodes a transmembrane receptor green fluorochrome direct labeled probe sisting of one orange/green fusion signal, tyrosine kinase. This gene exerts charac- hybridizes proximal to the ALK gene one orange signal, and a separate green teristic oncogenic activities through fusion breakpoint region at 2p23.1-p23.2. signal indicates one normal 2p23.1-p23.2 to several gene partners or mutations both locus and one 2p23.1-p23.2 locus affect- in hematopoietic and non-hematopoietic ed by a translocation or inversion. solid tumors. EML4-ALK inversion with deletion of Translocations affecting the ALK gene 5’-ALK sequences is indicated by one or locus are frequently found in anaplastic multiple isolated orange signals. large cell lymphoma (ALCL), an aggressive non-Hodgkin lymphoma arising from T-cells. The most frequent translocation t(2;5) results in a fusion with the NPM1 (nucleophosmin a.k.a. nucleolar phos- phoprotein B23, numatrin) gene located on chromosome 5q35. This rearrangement results in a NPM1/ALK fusion protein, Ideogram of chromosome 2 which is constitutively activated through indicating the hybridization locations. autophosphorylation, and that in turn D2S2934 D2S405 mediates malignant cell transformation by RH12489 SHGC-142979 SPEC ALK Dual Color Break Apart Probe 3’ 5’ activating downstream effectors like e.g. hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. STAT3. ALK Additionally, inversions affecting the ALK ~210 kb ~635 kb gene located on the short arm of chro- Tel 2p23.1-p23.2 Cen

mosome 2 [inv(2)(p21p23)] have been SPEC ALK Probe map (not to scale). frequently detected in non-small cell lung cancer (NSCLC) and lead to the formation of EML4-ALK fusion transcripts. ALK kinase targeted therapies may represent a very effective therapeutic strategy References in NSCLC patients carrying EML4-ALK Inamura K, et al. (2009) Mod Pathol 22: 508-15. Koivunen JP, et al. (2008) Clin Cancer Res 14: 4275-83. rearrangements. Martelli MP, et al. (2009) Am J Pathol 174: 661-70. Palmer RH, et al. (2009) Biochem J 420: 345-61. Perner S, et al. (2008) Neoplasia 10: 298-302. Lung carcinoma tissue section with translocation Rodig SJ, et al. (2009) Clin Cancer Res 15: 5216-23. affecting the 2p23 locus as indicated by Sasaki T, et al. (2010) Eur J Cancer 46: 1773-80. Von Laffert M, et al. (2013) Lung Cancer 81: 200-6. one orange/green fusion (non-rearranged) signal, Zhang Q, et al. (2007) Nat Med 11: 1341-8. one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2124-50 ZytoLight SPEC ALK Dual Color Break Apart Probe / 5 (50 μl) Z-2124-200 ZytoLight SPEC ALK Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

37 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 37 FE045-1-20 ZytoLight ® SPEC ALK/2q11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ALK/2q11 Dual The SPEC ALK/2q11 Dual Color Probe is In a normal interphase nucleus, two or- Color Probe is designed for the detection a mixture of a green fluorochrome direct ange and two green signals are expected. of amplifications of the chromosomal labeled SPEC ALK probe hybridizing to In a cell with amplification of the ALK region harboring the ALK gene. the human ALK gene in the chromosomal gene locus, multiple copies of the green The ALK (ALK receptor tyrosine kinase, region 2p23.1-p23.2 and an orange signal or green signal clusters will be a.k.a. CD246) gene is located on chro- fluorochrome direct labeled SPEC 2q11 observed. mosome 2p23.1-p23.2 and encodes a probe. The SPEC 2q11 probe is specific transmembrane receptor tyrosine kinase. for the AFF3 (AF4/FMR2 family, member ALK was originally identified as a fusion 3) gene region in 2q11.2. Due to cross-hy- partner of NPM1. This gene fusion is fre- bridizations of chromosome 2 alpha satel- quently found in anaplastic large cell lym- lites to other centromeric regions, probes phoma (ALCL), an aggressive non-Hodgkin specific for 2q11 are frequently used for lymphoma. Rearrangements affecting the chromosome 2 copy number detection. ALK gene locus have also been found to play a role in carcinogenesis of a variety ALK of hematopoietic and non-hematopoietic solid tumors, including non-small cell lung cancer (NSCLC). Moreover, ALK amplifications and copy 2q11 SPEC ALK/2q11 Dual Color Probe hybridized to normal interphase cells as indicated by two orange number gains have been reported to occur and two green signals in each nucleus. in a variety of tumors including NSCLC and alveolar rhabdomyosarcoma (ARMS). In colorectal cancer, ALK amplification was correlated with nodal status suggesting that ALK amplified tumors have a more aggressive phenotype. ALK copy number Ideogram of chromosome 2 gains and amplifications are also a fre- indicating the hybridization locations. quent genetic event in the tumorigenesis of D2S405 SHGC-142979 neuroblastomas and were found to result 3’ 5’ in high ALK expression correlating with an unfavorable neuroblastoma phenotype. ALK Hence, the identification of ALK gene copy ~635 kb Tel 2p23.1-p23.2 Cen number changes by in situ Hybridization Neuroblastoma tissue section with tetrasomy of SPEC ALK Probe map (not to scale). might be of prognostic and therapeutic chromosome 2 as indicated by four orange (2q11) and four green (ALK) signals in each nucleus. relevance. D2S2776 RH121214 3’ 5’ References Carén H, et al. (2008) Biochem J 416: 153-9. Corao DA, et al. (2009) Pediatr Dev Pathol 12: 275-83. AFF3 Koivunen JP, et al. (2008) Clin Cancer Res 14: 4275-83. Montagut C, et al. (2010) J Clin Oncol 28: Suppl 10537. ~490 kb Pelosi G, et al. (2012) Lung Cancer 77: 507-14. Salido M, et al. (2011) J Thorac Oncol 6: 21-7. Cen 2q11.2 Tel Subramaniam MM, et al. (2009) Hum Pathol 40: 1638-42. SPEC 2q11 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2161-200 ZytoLight SPEC ALK/2q11 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

38 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 38 FE073-1-20 ZytoLight ® SPEC EML4 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC EML4 Dual Color The SPEC EML4 Dual Color Break Apart In an interphase nucleus of a normal cell Break Apart Probe is designed to detect Probe is a mixture of two direct labeled lacking an inversion involving the 2p21 rearrangements involving the chromosom- probes hybridizing to the 2p21 band. The band, two orange/green fusion signals al region 2p21 harboring the EML4 (echi- orange fluorochrome direct labeled probe are expected representing two normal noderm microtubule-associated protein-like hybridizes proximal, the green fluoro- (non-rearranged) 2p21 loci. A signal pat- 4, a.k.a. ROPP120) gene. chrome direct labeled probe hybridizes tern consisting of one orange/green fusion Inversions in the short arm of chromosome distal to the EML4 gene breakpoint region signal, one orange signal, and a separate 2 [inv(2)(p21p23)] have been frequently at 2p21. green signal indicates one normal 2p21 detected in non-small cell lung cancer locus and one 2p21 locus affected by an (NSCLC) and lead to the formation of inversion or translocation. EML4-ALK fusion transcripts. A few reports EML4 also identified these fusion transcripts in breast, gastric, and colorectal cancers. The fusion genes comprise variably truncated N-terminal portions of the EML4 gene and the intracellular signaling domain of the ALK receptor tyrosine kinase (a.k.a. CD246). It was found that EML4 mediates ligand-independent dimerization of ALK, resulting in constitutive kinase activity. EML4-ALK was shown to possess transforming activity in vitro and in vivo. Ideogram of chromosome 2 indicating the hybridization locations. The EML4-ALK fusion transcript is found in SPEC EML4 Dual Color Break Apart Probe about 5% of NSCLC, predominantly ad- D2S2932 RH81694 hybridized to normal interphase cells as indicated D2S2305 D2S1482 by two orange/green fusion signals per nucleus. enocarcinomas, and is considered to be 5’ 3’ mutually exclusive to EGFR or KRAS mu- EML4 tations. The detection of the inversion by ~610 kb ~585 kb Fluorescence in situ Hybridization might Tel 2p21 Cen represent a valuable tool to identify a SPEC EML4 Probe map (not to scale). subpopulation of NSCLC likely to respond to ALK kinase targeting therapies. The SPEC EML4 Dual Color Break Apart Probe can be used to subsequently confirm EML4-ALK inversion if an ALK Break Apart Probe has been used for initial diagnosis. References Choi YL, et al. (2008) Cancer Res 69: 4971-6. Inamura K, et al. (2009) Mod Pathol 22: 508-15. Lin E, et al. (2009) Mol Cancer Res 7: 1466-76. NSCLC tissue section with inversion Perner S, et al. (2008) Neoplasia 10: 298-302. affecting the EML4 locus at 2p21 as indicated by Rodig SJ, et al. (2009) Clin Cancer Res 15: 5216-23. Soda M, et al. (2007) Nature 448: 561-6. one orange/green fusion (non-rearranged) signal, Shaw AT, et al. (2009) J Clin Oncol 27: 4247-53. one green signal, and one separate orange signal.

Prod. No. Product Label Tests* (Volume) Z-2136-50 ZytoLight SPEC EML4 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

39 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 39 FE058-1-20 ® ZytoLight SPEC IGK Dual Color Break Apart ProbeHome saltwater

Background Probe Description Results The ZytoLight ® SPEC IGK Dual Color The SPEC IGK Dual Color Break Apart In an interphase nucleus lacking a Break Apart Probe is designed to detect Probe is a mixture of a green fluorochrome translocation involving the IGK locus at rearrangements affecting the chromosomal direct labeled probe hybridizing distal to 2p11.2, two orange/green fusion signals region 2p11.2 harboring the IGK the IGK breakpoint region at 2p11.2 and are expected. A signal pattern consisting (immunoglobulin kappa locus, a.k.a. IGK@, an orange fluorochrome direct labeled of one orange/green fusion signal, one IGκ) gene cluster region. probe hybridizing proximal to the IGK orange signal, and a separate green Translocations involving the immunoglobulin breakpoint region. Due to homologous signal indicates one normal IGK locus and (IG) genes are recurring events of B-cell sequence segments proximal to the IGK one IGK locus affected by a translocation. oncogenesis. In all of these translocations, breakpoint region, the orange probe Due to the two hybridization regions of an oncogene is activated and overexpressed has two hybridization regions in close the orange probe, orange signals may by juxtaposing this oncogene to IG proximity. appear as paired signal dots. regulatory sequences. Burkitt lymphoma (BL) is characterized by reciprocal translocations involving the MYC gene and one of the IG loci. The majority of translocations involve the immunoglobulin heavy chain (IGH) locus while a minor part IGK involves the immunoglobulin light chain loci, either the kappa light chain (IGK) or the lambda light chain (IGL). IGK and IGL rearrangements resulting from the variant translocations t(2;8)(p11.2;q24.21) and SPEC IGK Dual Color Break Apart Probe hybridized t(8;22)(q24.21;q11.2), respectively, have to normal interphase cells as indicated by two orange/green fusion signals in each nucleus and to meta- been detected in up to 25% of BL cases. phase chromosomes of a normal cell. Orange signals In non-Hodgkin lymphoma (NHL) harboring Ideogram of chromosome 2 may appear as paired signal dots. indicating the hybridization locations. IG-MYC rearrangements, the MYC BV209599 SHGC-145092 SHGC-17726 translocation partner is IGK and IGL in SHGC-143963 RH109567 SHGC-7625 8 and 22% of the cases, respectively. 5’ 3’ IG translocations have been reported in several B-cell lineage malignancies other IGK than BL including atypical Burkitt/Burkitt-like ~770 kb ~365 kb ~235 kb Tel 2p11.2 Cen lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, IGK Probe map (not to scale). and multiple myeloma. Other rearrangement events involve the IGK and IGL gene with the BCL2 and BCL6 oncogenes as translocation Burkitt lymphoma with an IGK translocation affecting the 2p11.2 locus as indicated by partners. The detection of IGK and IGL one non-rearranged orange/green fusion signal, References one orange signal (may appear as paired involvement in lymphomas by Fluorescence Cario G, et al. (2000) Br J Haematol 110: 537-46. Einerson RR, et al. (2006) Leukemia 20: 1790-9. signal dots), and one separate green signal. in situ Hybridization may prove a valuable Henglein B, et al. (1989) Mol Cell Biol 9: 2105-13. Martín-Subero JI, et al. (2002) Int J Cancer 98: 470-4. Specimen kindly provided by Dr. Brändle, Vienna, Austria. diagnostic and prognostic tool. Poulsen TS, et al. (2002) Leukemia 16: 2148-55.

Prod. No. Product Label Tests* (Volume) Z-2288-50 ZytoLight SPEC IGK Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

40 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 40 FE156-1-20 ZytoLight ® SPEC ERBB4/2q11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ERBB4/2q11 Dual The SPEC ERBB4/2q11 Dual Color Probe Using the SPEC ERBB4/2q11 Dual Color Color Probe is designed for the detection is a mixture of a green fluorochrome direct Probe in a normal interphase nucleus, of amplifications of the chromosomal labeled SPEC ERBB4 probe hybridizing two green and two orange signals are regions harboring the ERBB4 gene. to intronic sequences of the human ERBB4 expected. In a cell with amplification of The ERBB4 (a.k.a. HER4) gene encodes a gene in the chromosomal region 2q34 the ERBB4 gene locus, multiple copies of transmembrane glycoprotein acting as a and an orange fluorochrome direct labe- the green signal or green signal clusters cellular growth factor receptor. It belongs led SPEC 2q11 probe. The SPEC 2q11 will be observed. to the epidermal growth factor receptor probe is specific for the AFF3 (AF4/FMR2 subgroup of the receptor tyrosine kinase family, member 3) gene region in 2q11.2. superfamily also including ERBB1 (EGFR), Due to cross-hybridizations of chromo- ERBB2, which is known to be affected by some 2 alpha satellites to other centromer- gene amplifications in a number of malig- ic regions, probes specific for 2q11 are nant tumors, and ERBB3. frequently used for chromosome 2 copy Although EGFR and ERBB2 have been number detection. shown to represent good predictive markers and appropriate targets for therapeutic approaches, relatively less is known of comparable significance for ERBB3 and ERBB4. However, there is growing evidence that cooperation of all four members of the ERBB gene family contributes to SPEC ERBB4/2q11 Dual Color Probe hybridized to normal interphase cells as indicated by two green and a more aggressive tumor phenotype and two orange signals in each nucleus. influences therapeutic response. Accordingly, it is assumed that the assessment of the combined amplification ERBB4 status of ERBB1 to ERBB4 may improve the diagnostic value significantly. Recently it was shown in a retrospective Ideogram of chromosome 2 indicating the hybridization locations. study that responsiveness to Herceptin™ turned out to be more efficient if tumor RH4706 SHGC-144718 3’ 5’ cells show ERBB4 gene amplification.

References ERBB4 Alimandi M, et al. (1995) Oncogene 10: 1813-21. Begnami MD, et al. (2011) J Clin Oncol 29: 3030-6. ~1040 kb Brockhoff G, et al. (2011) Acta Derm Venereol 91: 488-90. Cen 2q34 Tel Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Plowman GD, et al. (1993) Proc Natl Acad Sci U S A 90: 1746-50. SPEC ERBB4 Probe map (not to scale). Sassen A, et al. (2008) Breast Cancer Res 10: R2. Sassen A, et al. (2009) Breast Cancer Res 11: R50. Breast cancer tissue section with amplification of Zaczek A, et al. (2005) Histol Histopathol 20: 1005-15. D2S2776 RH121214 the ERBB4 gene (green), SPEC 2q11 (orange). Zimonjic DB, et al. (1995) Oncogene 10: 1235-7. 3’ 5’ Image kindly provided by Prof. Brockhoff, Regensburg, Germany. AFF3

~490 kb Cen 2q11.2 Tel

SPEC 2q11 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2057-200 ZytoLight SPEC ERBB4/2q11 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

41 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 41 FE132-1-20 ZytoLight ® SPEC VHL/CEN 3 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC VHL/CEN 3 Dual The SPEC VHL/CEN 3 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of deletions affecting the VHL gene. direct labeled CEN 3 probe specific for In a cell with deletions affecting the VHL The tumor suppressor gene VHL (von the alpha satellite centromeric region of gene, one or no copy of the green signal Hippel-Lindau tumor suppressor) is located chromosome 3 (D3Z1) and a green fluo- will be observed. on 3p25.3 and encodes a 30 kDa protein rochrome direct labeled SPEC VHL probe with ubiquitin ligase E3 activity. The pro- spanning the VHL gene at 3p25.3. tein is involved in the ubiquitination and degradation of hypoxia-inducible-factor (HIF), which is a transcription factor that plays a critical role in the regulation of gene expression by oxygen. Loss of heterozygosity (LOH) at chromosome 3p and inactivation of the VHL gene by deletion or mutation is the most frequent genetic change in sporadic as well as VHL disease-associated conventional renal cell carcinomas (RCC) whereas alterations of this region are rarely seen in Ideogram of chromosome 3 SPEC VHL/CEN 3 Dual Color Probe hybridized to indicating the hybridization locations. normal interphase cells as indicated by two orange papillary and chromophobe RCC. Recent and two green signals in each nucleus. studies suggest that the determination of RH58266 D3S3107 5’3’ the VHL status by FISH can significantly improve the accuracy of kidney tumor VHL biopsy evaluation, providing prognostic ~550 kb information that can guide management Tel 3p25.3 Cen decisions. SPEC VHL Probe map (not to scale).

References Barocas DA, et al. (2006) BJU Int 99: 290-5. Broom RJ, et al. (2012) Clin Genitourin Cancer 10: 202-6. Dagher J, et al. (2013) Hum Pathol 44: 2106-15. Hosoe S, et al. (1990) Genomics 8: 634-40. Latif F, et al. (1993) Science 260: 1317-20. Sükösd F, et al. (2003) Cancer Res 63: 455-7.

Trisomy of chromosome 3 as indicated by three orange (CEN 3) and three green (VHL) signals in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2084-200 ZytoLight SPEC VHL/CEN 3 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

42 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 42 FE022-1-20 ZytoLight ® SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe

Background Probe Description Results The ZytoLight ® SPEC VHL/1p12/CEN 7/17 The SPEC VHL/1p12/CEN 7/17 Quad- In a normal interphase nucleus, two green, Quadruple Color Probe is designed for an ruple Color Probe is a mixture of a green two red, two gold, and two blue signals are accurate identification of renal cell carcinoma fluorochrome direct labeled SPEC VHL probe expected. In a cell with deletion affecting the (RCC) subtypes by the simultaneous detection spanning the VHL gene at 3p25.3, a gold fluo- VHL gene, a reduced number of green signals of VHL gene status and enumeration of chromo- rochrome direct labeled CEN 7 probe specific will be observed. In cells with aneusomy of somes 1, 7, and 17 in tumor cells. for the alpha satellite centromeric region of chromosome 1, 7, or 17, more or less signals Clear cell RCC (ccRCC), papillary RCC (pRCC), chromosome 7 (D7Z1), a blue fluorochrome of the respective color will be visible. chromophobe RCC (chRCC) and renal onco- direct labeled CEN 17 probe specific for the cytomas (ROs) are the most frequent renal cell alpha satellite centromeric region of chromo- tumor subtypes. Patients with ccRCC have a some 17 (D17Z1), and a red fluorochrome poorer prognosis than patients with pRCC and direct labeled SPEC 1p12 hybridizing in close chRCC. RO is considered to be a benign neo- proximity to the centromere of chromosome plasm. The differentiation between RCC types 1 at the chromosomal region 1p12. Due to may sometimes be difficult on histopathological cross-hybridizations of chromosome 1 alpha features alone. However, the different subtypes satellites to other centromeric regions, probes of kidney tumors are characterized by distinct specific for 1p12 are frequently used for chro- genetic patterns. Chromosome 3p deletion, mosome 1 copy number detection. Renal cell carcinoma tissue section with including deletion of the tumor suppressor gene deletion of the VHL gene as indicated by RH58266 D3S3107 VHL (von Hippel-Lindau tumor suppressor) in 5’3’ one green signal in each nucleus. 3p25.3, is the most typical genetic abnor- VHL mality in ccRCC. pRCC is characterized by ~550 kb trisomy/polysomy of chromosomes 7 and 17. Tel 3p25.3 Cen Combined losses of chromosomes 1, 2, 6, 10, SPEC VHL Probe map (not to scale). 13, 17, and 21 (with 1, 2, 6, and 17 being SHGC-141884 RH105871 affected most frequently) are the most common 3’ 5’ 5’ 3’ changes in chRCC, whereas ROs often show rearrangements involving 11q13.3 harboring WARS2 HAO2 the CCND1 gene or losses of chromosomes 1, ~285 kb Tel 1p12 Cen 14, and sex chromosomes. Renal cell carcinoma tissue section with polysomy SPEC 1p12 Probe map (not to scale). Consequently, the ZytoLight ® SPEC VHL/1p12/ of the chromosome 7 and 17 as indicated by multiple gold and/or blue signals in each nucleus. CEN 7/17 Quadruple Color Probe is designed to differentiate between ccRCC, pRCC, and some chRCC tumors and should be used in combination with the ZytoLight ® SPEC CCND1 VHL Break Apart/2q11/CEN 6 Quadruple Color Probe which helps to especially differentiate 1p12 between chRCC and ROs. CEN 7 (D7Z1) CEN 17 (D17Z1) References Brunelli M, et al. (2005) Modern Pathology 18: 161-9. Iqbal MA, et al. (2000) Diagn Cytopathol 22: 3-6. Jhang JS, et al. (2004) Cancer Genet Cytogenet 149: 114-9. Mertz KD, et al. (2006) Urologe 45: 316-22. Moch H (2013) Semin Cancer Biol 23: 3-9. Sanjmyatav J, et al. (2013) Eur Urol 64: 689-91. Sukov WR, et al. (2009) Hum Pathol 40: 1296-303.

Ideograms of chromosomes 3, 1, 7, and 17 indicating the hybridization locations.

Prod. No. Product Label Tests* (Volume) Z-2102-200 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe •/ / /• 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

43 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 43 FE078-1-20 ZytoLight ® SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe

Background Probe Description Results The ZytoLight ® SPEC CCND1 Break The SPEC CCND1 Break Apart/2q11/ In a normal interphase nucleus, two red/ Apart/2q11/CEN 6 Quadruple Color CEN 6 Quadruple Color Probe is a green fusion signals, two blue, and two Probe is designed for an accurate iden- mixture of a green and a red fluorochrome gold signals are expected. In a cell with tification of renal cell carcinoma (RCC) direct labeled probe hybridizing proximal translocation of the CCND1 gene locus, subtypes by the simultaneous detection and distal to the breakpoint on 11q13.3, a signal pattern consisting of one red/ of rearrangements affecting the CCND1 respectively, a gold fluorochrome direct la- green fusion signal, one red, and a sep- (cyclin D1, a.k.a. BCL2 or PRAD1) gene beled CEN 6 probe specific for the alpha arate green signal indicates one normal in 11q13.3 and enumeration of chromo- satellite centromeric region of chromosome CCND1 locus and one CCND1 locus somes 2 and 6 in tumor cells. 6 (D6Z1), and a blue fluorochrome direct affected by an 11q13.3 translocation. In Clear cell RCC (ccRCC), papillary RCC labeled SPEC 2q11 probe. The SPEC cells with aneusomy of chromosome 2 or (pRCC), chromophobe RCC (chRCC), and 2q11 probe is specific for the AFF3 (AF4/ 6, more or less signals of the respective renal oncocytomas (ROs) are the most FMR2 family, member 3) gene region in color will be visible. frequent renal cell tumor subtypes. Patients 2q11.2. Due to cross-hybridizations of with ccRCC have a poorer prognosis than chromosome 2 alpha satellites to other patients with pRCC and chRCC. RO is centromeric regions, probes specific for considered to be a benign neoplasm. The 2q11 are frequently used for chromosome differentiation between RCC types may 2 copy number detection. sometimes be difficult on histopathological features alone. However, the different sub- RH47914 PMC133966P1 RH40065 RH92087 types of kidney tumors are characterized 5’3’ 5’ 3’ by distinct genetic patterns. Chromosome MYEOV CCND1 3p deletion, including deletion of the ~455 kb ~580 kb tumor suppressor gene VHL (von Hip- Cen 11q13.2-q13.3 Tel Renal cell carcinoma tissue section with translocation affecting the 11q13.3 locus as indicated by one pel-Lindau tumor suppressor) in 3p25.3, SPEC CCND1 Probe map (not to scale). non-rearranged red/green fusion signal, one red is the most typical genetic abnormality signal, and one separate green signal. D2S2776 RH121214 in ccRCC. pRCC is characterized by 3’ 5’ trisomy/polysomy of chromosomes 7 and 17. Combined losses of chromosomes 1, AFF3 2, 6, 10, 13, 17, and 21 (with 1, 2, 6, ~490 kb Cen 2q11.2 Tel and 17 being affected most frequently) SPEC 2q11 Probe map (not to scale). are the most common changes in chRCC, whereas ROs often show rearrangements involving 11q13.3 or losses of chromosomes 1, 14, and sex chromosomes. CEN 6 (D6Z1) Consequently, the ZytoLight ® SPEC 2q11 CCND1 CCND1 Break Apart/2q11/CEN 6 Renal cell carcinoma tissue section with monosomy of chromosome 2 and 6 as indicated by one blue Quadruple Color Probe is designed to and one gold signal in each nucleus. especially differentiate between chRCC and ROs and should be used in combina- References ® tion with the ZytoLight SPEC VHL/1p12/ Ideograms of chromosomes 11, 2, and 6 Brunelli M, et al. (2005) Modern Pathology 18: 161-9. Iqbal MA, et al. (2000) Diagn Cytopathol 22: 3-6. CEN 7/17 Quadruple Color Probe which indicating the hybridization locations. Jhang JS, et al. (2004) Cancer Genet Cytogenet 149: 114-9. Mertz KD, et al. (2006) Urologe 45: 316-22. is designed for the differentiation between Moch H (2013) Semin Cancer Biol 23: 3-9. Sanjmyatav J, et al. (2013) Eur Urol 64: 689-91. ccRCC, pRCC, and some chRCC tumors. Sukov WR, et al. (2009) Hum Pathol 40: 1296-303.

Prod. No. Product Label Tests* (Volume) Z-2118-200 ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe •/ /•/ 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

44 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 44 FE079-1-20 ZytoLight ® SPEC FHIT/CEN 3 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC FHIT/CEN 3 Dual The SPEC FHIT/CEN 3 Dual Color probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of FHIT gene deletions frequently observed direct labeled CEN 3 probe specific for In a cell with deletion of the FHIT gene in most of the common epithelial neo- the alpha satellite centromeric region of locus, a reduced number of green signals plasms. chromosome 3 (D3Z1) and a green fluo- will be observed. Deletions affecting only The FHIT (fragile histidine triad) gene is lo- rochrome direct labeled SPEC FHIT probe parts of introns 4 and/or 5 of the FHIT cated in the chromosomal region 3p14.2 hybridizing to sequences of introns 4 and gene might result in a normal signal pat- and encodes a 16.8 kDa member of the 5 of the human FHIT gene at 3p14.2. tern with green signals of reduced size. HIT superfamily of nucleoside monophos- phate hydrolases and transferases. The 1.6 Mb FHIT gene encompasses the most carcinogen-sensitive common fragile region FRA3B and the t(3;8) translocation breakpoint associated with hereditary renal carcinoma. The tumor suppressor gene FHIT is inactivated by deletions in a variety of human tumors e.g. lung, kidney, gastric, breast, pancreatic, and cervical tumors. Since loss of the FHIT locus occurs in a number of Ideogram of chromosome 3 indicating the hybridization locations. preneoplastic lesions, FHIT may represent SPEC FHIT/CEN 3 Dual Color Probe hybridized a potential marker for the detection of SHGC-145137 HUMUT2337 to interphase cells each showing three orange 3’ 5’ and two green signals. tumor precursor cells.

References FHIT Broom RJ, et al. (2012) Clin Genitourin Cancer 10: 202-6. ~695 kb Cirombella R, et al. (2010) Cancer Lett 291: 230-6. Huebner K, et al. (1998) Annu Rev Genet 32: 7-31. Tel 3p14.2 Cen Ishii H, et al. (2003) J Exp Ther Oncol 3: 291-6. Ohta M, et al. (1996) Cell 84: 587-97. SPEC FHIT Probe map (not to scale). Pekarsky Y, et al. (2002) Lancet Oncol 3: 748-54. Schwarz S, et al. (2008) Cytometry A 73: 305-11. Vieira J, et al. (2010) Genes Chromosomes Cancer 49: 935-47.

Prod. No. Product Label Tests* (Volume) Z-2062-200 ZytoLight SPEC FHIT/CEN 3 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

45 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 45 FE011-1-20 ZytoLight ® SPEC GATA2/MECOM Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC GATA2/MECOM The SPEC GATA2/MECOM Dual Color In a normal interphase nucleus, two Dual Color Dual Fusion Probe is designed to Dual Fusion Probe is a mixture of an green and two orange signals are detect the inversion inv(3)(q21q26.2) and orange fluorochrome direct labeled probe expected. An aberration involving the the translocation t(3;3)(q21;q26.2) both spanning the GATA2 gene region at chromosomal regions of GATA2 and affecting the GATA2 (a.k.a. NFE1B) gene 3q21.3 and a green fluorochrome direct MECOM generates a fusion signal on in the chromosomal region 3q21.3 and the labeled probe spanning the MECOM each of the chromosomes involved in case MECOM (MDS1 and EVI1 complex locus, gene region at 3q26.2. This probe is of a t(3;3) or two fusion signals on the a.k.a. MDS1, EVI1) gene in 3q26.2. approved to be used with a hybridization involved chromosome in case of an inv(3). MECOM and GATA2 are transcription time of 2 hours on cytological specimens. The chromosomal regions that are not factors that play an essential role in the translocated are indicated by the single proliferation of hematopoietic stem cells. green and orange signal, respectively. Inv(3)/t(3;3), and less commonly ins(3;3) Other relevant signal patterns may also be (q26.2;q21q26.2), occur in 1-2.5% of observed as a result of ins(3;3) or 3q26.2 acute myeloid leukemia (AML) and are also rearrangements without the involvement of observed in myelodysplastic syndromes the GATA2 locus. and in the blastic phase of chronic myeloid GATA2 leukemia. A variety of other MECOM translocations involving other fusion partner MECOM genes have also been reported in various types of myeloid malignancies. 3q26.2 Ideogram of chromosome 3 rearrangements are associated with minimal indicating the hybridization locations. to no response to chemotherapy and poor RH112766 RH66627 clinical outcome. 3’ 5’ The inv(3) and t(3;3) result in overexpression GATA2 of the MECOM gene due to its ~660 kb juxtaposition to enhancer sequences of the Cen 3q21.3 Tel GATA2 gene and simultaneously confer SPEC GATA2/MECOM Dual Color Dual Fusion SPEC GATA2 Probe map (not to scale). Probe hybridized to normal interphase cells as GATA2 haploinsufficiency, leading to indicated by two orange and two green signals leukemogenesis. D3S3084 RH10606 and to metaphase chromosomes of a normal cell. In the revised 2016 WHO classification of 3’ 5’

myeloid neoplasms and acute leukemia, MECOM

“AML with inv(3)(q21.3q26.2) or t(3;3) ~1.5 Mb (q21.3;q26.2); GATA2, MECOM” is Cen 3q26.2 Tel classified as its own entity, emphasizing the SPEC MECOM Probe map (not to scale). unique clinicopathologic features and poorer prognosis of this subgroup of AML patients. Chromosome 3q26.2 rearrangements may References Arber DA, et al. (2016) Blood 127: 2391-405. be cryptic on standard karyotype analysis. Bobadilla D, et al. (2007) Br J Haematol 136: 806-13. De Braekeleer E, et al. (2011) Anticancer Res 31: 3441-8. Bone marrow smear with rearrangement affecting Hence, FISH may be a helpful tool to confirm Pintado T, et al. (1985) Cancer 55: 535-41. the GATA2/MECOM loci as indicated by one Tang Z, et al. (2019) Cancer Genet 233-234: 21-31. the diagnosis of this distinct AML subgroup. Yamazaki H, et al. (2014) Cancer Cell 25: 415-27. separate orange signal, one separate green signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2287-50 ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

46 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 46 FE159-1-20 ZytoLight ® SPEC WWTR1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC WWTR1 Dual Color The SPEC WWTR1 Dual Color Break In an interphase nucleus lacking a translo- Break Apart Probe is designed for the Apart Probe is a mixture of two direct cation involving the 3q24-3q25.1 band, detection of translocations involving the labeled probes hybridizing to the 3q24- two orange/green fusion signals are chromosomal region 3q25.1 harboring the 3q25.1 band. The green fluorochrome expected representing two normal (non-re- WWTR1 (WW domain containing tran- direct labeled probe hybridizes in 3q24- arranged) 3q24-3q25.1 loci. A signal pat- scription regulator 1, a.k.a. TAZ) gene. 3q25.1 proximal and the orange fluoro- tern consisting of one orange/green fusion Epithelioid vascular tumors encompass chrome direct labeled probe hybridizes in signal, one orange signal, and a separate a spectrum of diseases that includes 3q25.1 distal to the WWTR1 gene. green signal indicates one normal 3q24- epithelioid hemangioma (EH), a benign 3q25.1 locus and one 3q24-3q25.1 locus neoplasm, epithelioid hemangioendothe- affected by a translocation. lioma (EHE), a low to intermediate grade malignancy, and epithelioid angiosarcoma (EAS), a high grade malignancy. Although certain morphologic features allow to distinguish EHE from EH and EAS, the diagnosis can be challenging due to considerable morphologic overlap, particularly on small WWTR1 biopsies or when EAS lacks vasoformative properties. Clinical behavior and, consequently, treatment and prognosis vary signif- Ideogram of chromosome 3 indicating the hybridization locations. icantly among vascular tumors. Therefore, it is paramount to effectively distinguish them RH17784 RH58195 D3S3082 SHGC-154409 from each other. 3’ 5’ SPEC WWTR1 Dual Color Break Apart Probe The recurrent translocation t(1;3) hybridized to normal interphase cells as indicated WWTR1 by two orange/green fusion signals per nucleus (p36.3;q25.1) was identified in approxi- and to metaphase chromosomes of a normal cell. mately 90% of EHE cases, but not in other ~700 kb ~505 kb Cen 3q24-q25.1 Tel vascular tumors. t(1;3) results in the SPEC WWTR1 Probe map (not to scale). WWTR1-CAMTA1 fusion gene which encodes a putative chimeric transcription factor which is under the transcriptional control of the WWTR1 promoter. A recurrent YAP1-TFE3 gene fusion has been identified in WWTR1-CAMTA1 negative EHEs. Thus, FISH analysis for the presence of WWTR1 translocation may serve as a useful molecular tool in the differential diagnosis of challenging cases.

References Anderson T, et al. (2015) Am J Surg Pathol 39: 132-9. Epitheloid hemangioendothelioma interphase Errani C, et al. (2011) Genes Chromosomes Cancer 50: 644-53. cells showing translocation of the WWTR1 Mendlick MR, et al. (2001) Am J Surg Pathol 25: 684-7. gene as indicated by one non-rearranged Puls F, et al. (2015) Virchows Arch 466: 473-8. orange/green fusion signal, one orange Tanas MR, et al. (2011) Sci Transl Med 3: 98ra82. signal and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2212-50 ZytoLight SPEC WWTR1 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

47 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 47 FE130-1-20 ZytoLight ® SPEC TERC/CEN 3 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC TERC/CEN 3 Dual The SPEC TERC/CEN 3 Dual Color Probe Using the SPEC TERC/CEN 3 Dual Color Color Probe is designed to detect amplifi- is a mixture of an orange fluorochrome Probe in a normal interphase nucleus, two cations affecting the chromosomal region direct labeled probe spanning the TERC orange and two green signals are expected. 3q26.2 harboring the TERC (telomerase gene region at 3q26.2 and a green fluo- In a cell with gain of the TERC gene locus, RNA component, a.k.a. hTERC, TRC3) rochrome direct labeled probe hybridizing multiple copies of the orange signal or gene. to the alpha satellite centromeric region of orange signal clusters will be observed. 3q copy number gains including the TERC chromosome 3 (D3Z1). gene locus, have been found in several epithelial carcinomas such as cervix carcinoma, prostate cancer, non-small cell lung cancer, and lung squamous cell carcinoma. TERC amplifications are not only found in cancers but also in pre-cancerous CEN 3 (D3Z1) lesions such as atypical squamous cell of undetermined significance (ASCUS). For cervical carcinoma, which is the second most common malignancy among women TERC worldwide, TERC amplifications have become a molecular marker to distinguish Ideogram of chromosome 3 indicating the hybridization locations. between low-grade dysplasia and high- SPEC TERC/CEN 3 Dual Color Probe grade cervical neoplasia and invasive SHGC-68502 RH10606 hybridized to normal interphase cells as indicated 3’ 5’ by two orange and two green signals per nucleus. carcinoma. Only a minority of cases which are cytologically diagnosed as low-grade TERC squamous intraepithelial lesion (LSIL) show ~495 kb Cen 3q26.2 Tel a development to high-grade squamous intraepithelial lesions (HSIL). Since an SPEC TERC Probe map (not to scale). increase in TERC gene copy number has been shown to be strongly associated with the progression of cervical intraepithelial neoplasia (CIN) to invasive carcinoma, TERC amplification has been proposed as prognostic marker to identify low-grade lesions with high risk to progress to high- grade disease and cancer. Fluorescence in situ Hybridization (FISH) may be a reliable SPEC TERC/CEN 3 Dual Color Probe hybridized to diagnostic tool to complement Pap-testings CaSki cells with TERC amplification as indicated by and may be of prognostic relevance. three or four orange signals in each nucleus.

References Andersson S, et al. (2009) Am J Pathol 175: 1831-47. Heselmeyer-Haddad K, et al. (2005) Am J Pathol 166: 1229-38. Heselmeyer K, et al. (1996) Proc Natl Acad Sci U S A 93: 479-84. Pelosi G, et al. (2007) Clin Cancer Res 13: 1995-2004. Yokoi S, et al. (2003) Clin Cancer Res 9: 4705-13.

Prod. No. Product Label Tests* (Volume) Z-2284-200 ZytoLight SPEC TERC/CEN 3 Dual Color Probe •/• 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

48 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 48 FE153-1-20 ZytoLight ® SPEC PIK3CA/CEN 3 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC PIK3CA/CEN 3 The SPEC PIK3CA/CEN 3 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of an orange fluo- ange and two green signals are expected. detection of PIK3CA gene amplifications rochrome direct labeled CEN 3 probe Nuclei with amplification of the PIK3CA frequently found in a variety of human specific for the alpha satellite centromeric gene locus 3q26.32-q26.33 or aneu- cancers. region of chromosome 3 (D3Z1) and a ploidy of chromosome 3 will show multiple The PIK3CA (a.k.a. PI3K-alpha) gene is green fluorochrome direct labeled SPEC copies of the green signal or large green located on chromosome 3q26.32 and PIK3CA probe specific for the chromo- signal clusters. encodes the 110 kDa catalytic subunit of somal region 3q26.32-q26.33 harboring the phosphatidylinositol 3-kinase (PI3K). the PIK3CA gene. Amplifications of PIK3CA were found e.g. in cervical, ovarian, endometrial, breast, gastric, and lung cancer. In ovarian cancer as well as cervical cancer cells increased copy numbers were shown to be associated with increased CEN 3 (D3Z1) expression of the gene product and PI3K activity. Furthermore, treatment with a PI3K inhibitor leads to decreased proliferation and increased apoptosis. It was concluded PIK3CA

that PIK3CA is an important oncogene in SPEC PIK3CA/CEN 3 Dual Color Probe hybridized these tumors. Ideogram of chromosome 3 to normal interphase cells as indicated by two indicating the hybridization locations. orange and two green signals in each nucleus. Likewise in endometrial carcinomas detec- D3S3730 SHGC-141305 tion of PIK3CA amplification is associated 5’ 3’ with tumor grade and stage. A significant correlation between PIK3CA PIK3CA amplification and poor survival was found ~760 kb Cen 3q26.32-q26.33 Tel for gastric cancer patients. SPEC PIK3CA Probe map (not to scale). PIK3CA amplification was also frequently found in non-small cell lung cancer (NSCLC) and was shown to be associated with certain clinicopathologic features. PIK3CA amplification seems to promote tumorigenesis through aberrant activation of the PI3K/Akt signaling pathway. Hence, Human breast cancer cell line with amplification this pathway might represent an effective of the PIK3CA gene as indicated by therapeutic target in several cancer types. multiple green signals in each nucleus. References Ji M, et al. (2011) BMC Cancer 11: 147. Ma YY, et al. (2000) Oncogene 19: 2739-44. Shayesteh L, et al. (1999) Nat Genet 21: 99-102. Shi J, et al. (2012) BMC Cancer 12: 50. Volinia S, et al. (1994) Genomics 24: 472-7.

Prod. No. Product Label Tests* (Volume) Z-2140-200 ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

49 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 49 FE060-1-20 ZytoLight ® SPEC SOX2/CEN 3 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC SOX2/CEN 3 Dual The SPEC SOX2/CEN 3 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of SOX2 gene amplifications frequently direct labeled CEN 3 probe specific for Nuclei with amplification of the SOX2 observed in squamous cell carcinoma the alpha satellite centromeric region of gene locus 3q26.33 or aneuploidy of (SCC) of the lung, the esophagus, the oral chromosome 3 (D3Z1) and a green fluoro- chromosome 3 will show multiple copies cavity, and further organ sites. In addition, chrome direct labeled SPEC SOX2 probe of the green signal or large green signal amplifications and/or overexpression specific for the SOX2 gene at 3q26.33. clusters. were found in glioma, breast cancer, and other tumor types. The SOX2 (SRY-box 2, a.k.a. ANOP3) gene is located on chromosome 3q26.33 and encodes a High Mobility Group domain transcription factor that is a regulator CEN 3 (D3Z1) of normal stem cell function in embryonic and neural stem cells. Amplification of the SOX2 gene was found in about 20% of lung SCC and SOX2 15% of esophageal SCC and results in oncogenic SOX2 overexpression. In a Ideogram of chromosome 3 indicating the hybridization locations. large series of lung SCC it was shown that SPEC SOX2/CEN 3 Dual Color Probe hybridized to amplification of SOX2 was associated SHGC-111316 B317YC5 normal interphase cells as indicated by two orange 5’3’ and two green signals in each nucleus. with lower tumor grade and hence with favorable prognosis. SOX2 However, in glioma and glioma cell lines ~825 kb Cen 3q26.33 Tel SOX2 expression seems to show a positive correlation with malignancy grade. SPEC SOX2 Probe map (not to scale).

References Alonso MM, et al. (2011) PLoS One 6: e26740. Annovazzi L, et al. (2011) Cancer Genomics Proteomics 8: 139-47. Bass AJ, et al. (2009) Nat Genet 41: 1238-42. Hussenet T, et al. (2010) PLoS One 5: e8969. Kokalj Vokac N, et al. (2014) Mol Cytogenet 7: 5. Maier S, et al. (2011) Hum Pathol 42: 1078-88. Wilbertz T, et al. (2011) Mod Pathol 24: 944-53.

Lung cancer tissue section with amplification of the SOX2 gene (green) and trisomy of chromosome 3 (orange).

Prod. No. Product Label Tests* (Volume) Z-2127-200 ZytoLight SPEC SOX2/CEN 3 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

50 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 50 FE051-1-20 ZytoLight ® SPEC BCL6 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC BCL6 Dual Color The SPEC BCL6 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed for the Probe is a mixture of two direct labeled location involving the 3q27.3-q28 band, detection of translocations involving the probes hybridizing to the 3q27.3-q28 two orange/green fusion signals are ex- chromosomal region 3q27.3 harboring band. The green fluorochrome direct labe- pected representing two normal (non-rear- the BCL6 (BCL6 transcription repressor, led probe hybridizes at 3q27.3 proximal ranged) 3q27.3-q28 loci. A signal pattern a.k.a. ZNF51, LAZ3) gene. to the BCL6 gene, and the orange fluoro- consisting of one orange/green fusion The BCL6 protein acts as a transcriptional chrome direct labeled probe hybridizes at signal, one orange signal, and a sepa- repressor that is involved in the regulation 3q27.3-q28 distal to the BCL6 gene. rate green signal indicates one normal of lymphoid development and function. 3q27.3-q28 locus and one 3q27.3-q28 Chromosomal rearrangements of the locus affected by a translocation. BCL6 gene region were found to occur in different types of non-Hodgkin lymphoma (NHL), including diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL). The most common BCL6 translocation t(3;14)(q27;q32.3) results in the IGH-BCL6 gene fusion. In addition, more than 20 partner loci have been identified BCL6 including immunoglobulin (Ig) genes but also a number of non-Ig genes. As a result Ideogram of chromosome 3 indicating the hybridization locations. of these translocations, the rearranged BCL6 gene comes under the control of the SST_PRO RH123158 RH58741 D3S4143 promoter of the partner gene leading to 3’5’ SPEC BCL6 Dual Color Break Apart Probe deregulated expression of BCL6. hybridized to normal interphase cells as indicated BCL6 by two orange/green fusion signals per nucleus. In DLBCL, the most common histologic ~665 kb ~350 kb subtype of NHL, BCL6 translocations rep- Cen 3q27.3-q28 Tel resent one of the most frequent cytogenetic SPEC BCL6 Probe map (not to scale). abnormality, occurring in 20% to 40% of the cases. Several studies reported a correlation of BCL6 translocation with an inferior overall survival. Moreover, DLBCL which are positive for both BCL6 and MYC rearrangements have been shown to have an extremely poor prognosis. Hence, the detection of BCL6 rearrangements by Fluorescence in situ Hybridiza- References Akyurek N, et al. (2012) Cancer 118: 4173-83. tion may help in predicting the clinical Cady FM, et al. (2008) J Clin Oncol 26: 4814-9. Ohno H (2004) Histol Histopathol 19: 637-50. DLBCL tissue section with translocation outcome in patients with NHL. Ohno H (2006) J Clin Exp Hematop 46: 43-53. of the BCL6 gene as indicated by one non-rearranged orange/green fusion signal, one orange, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2177-50 ZytoLight SPEC BCL6 Dual Color Break Apart Probe / 5 (50 μl) Z-2177-200 ZytoLight SPEC BCL6 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

51 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 51 FE086-1-20 ZytoLight ® SPEC FGFR3 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR3 Dual Color The SPEC FGFR3 Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving the rearrangements involving the chromosom- probes hybridizing to the 4p16.3 band. 4p16.3 band, two orange/green fusion al region 4p16.3 harboring the FGFR3 The orange fluorochrome direct labeled signals are expected representing two (fibroblast growth factor receptor 3, a.k.a. probe hybridizes proximal, the green fluo- normal (non-rearranged) 4p16.3 loci. A JTK4) gene. rochrome direct labeled probe hybridizes signal pattern consisting of one orange/ Rearrangements affecting the FGFR3 gene distal to the FGFR3 gene at 4p16.3. green fusion signal, one orange signal, are frequently found in carcinomas of and a separate green signal indicates one various types including multiple myeloma FGFR3 normal 4p16.3 locus and one 4p16.3 (MM), bladder cancer, glioblastoma, locus affected by a translocation. peripheral T-cell lymphoma, and lung squamous cell carcinoma. FGFR3 encodes for a transmembrane receptor tyrosine kinase which dimerizes after ligand binding leading to activation of downstream signaling cascades. This gene develops characteristic oncogenic activities after fusion to several gene part- Ideogram of chromosome 4 ners which often leads to ligand-independ- indicating the hybridization locations. ent activation of the tyrosine kinase of the RH11100 ECD24349 FGFR3 fusion protein. SHGC-110784 SHGC-51532 5’3’ Several in vivo and in vitro studies have SPEC FGFR3 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated by two demonstrated the therapeutic potential of FGFR3 orange/green fusion signals per nucleus. FGFR inhibitors in cell lines and animal ~635 kb ~525 kb models harboring FGFR3 fusion genes. Tel 4p16.3 Cen Hence, the detection of FGFR3 transloca- SPEC FGFR3 Probe map (not to scale). tions by Fluorescence in situ Hybridization may be a useful predictive biomarker in the selection of patients for FGFR-targeted therapy.

References Cheng T, et al. (2013) PLoS One 8: e57284. Fonseca R, et al. (2009) Leukemia 23: 2210-21. Kang S, et al. (2009) Mol Cell Biol 29: 2105-17. Knowles MA (2007) World J Urol 25: 581-93. Parker BC, et al. (2014) J Pathol 232: 4-15. Williams SV, et al. (2012) Hum Mol Genet 22: 795-803.

Breast cancer tissue section with translocation affecting the FGFR3 gene as indicated by one non-rearranged orange/green fusion signal, one orange, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2170-50 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe / 5 (50 μl) Z-2170-200 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

52 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 52 FE080-1-20 ZytoLight ® SPEC FGFR3/4p11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR3/4p11 Dual The SPEC FGFR3/4p11 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of a green fluorochrome direct ange and two green signals are expected. of FGFR3 gene amplifications. labeled SPEC FGFR3 probe hybridizing In a cell with amplification of the FGFR3 The FGFR3 (fibroblast growth factor recep- to the FGFR3 gene in the chromosomal gene locus, multiple copies of the green tor 3) gene is located in the chromosomal region 4p16.3 and an orange fluoro- signal or large green signal clusters will region 4p16.3 and encodes a receptor chrome direct labeled SPEC 4p11 probe be observed. tyrosine kinase. specific for the ZAR1 (zygote arrest 1) FGFR family members differ from one gene region in 4p11. For an unambigu- another in their ligand affinities and tissue ous enumeration of chromosome 4 the distribution. The extracellular portion of SPEC 4p11 is found to be more suitable. the protein interacts with fibroblast growth factors, setting in motion a cascade of FGFR3 downstream signals, ultimately influencing mitogenesis and differentiation. The 4p11 FGFR3 protein binds acidic and basic fibroblast growth hormone and plays a role in bone development and maintenance. Activating mutations are associated with multiple myeloma, cervical SPEC FGFR3/4p11 Dual Color Probe hybridized to carcinoma, and carcinoma of the bladder. normal interphase cells as indicated by two orange and two green signals in each nucleus. Additionally, it was found that copy num- Ideogram of chromosome 4 ber gains at 4p16.3 occurred significantly indicating the hybridization locations.

more frequently in recurred/metastasized D4S865 D4S2774E salivary gland adenoid cystic carcinoma 5’ 3’

(ACC) compared with indolent ACC. FGFR3 References Keegan K, et al. (1991) Proc Natl Acad Sci U S A 88: 1095-9. ~545 kb L’Hôte CG & Knowles MA (2005) Exp Cell Res 304: 417-31. Tel 4p16.3 Cen Thompson LM, et al. (1991) Genomics 11: 1133-42. Vékony H, et al. (2007) Clin Cancer Res 13: 3133-9. SPEC FGFR3 Probe map (not to scale).

RH67343 RH82989 5’3’

ZAR1

~435 kb Tel 4p11 Cen

SPEC 4p11 Probe map (not to scale). Bladder cancer tissue section with interphase cells showing polysomy of chromosome 4 as indicated by multiple green and orange signals in the nuclei.

Prod. No. Product Label Tests* (Volume) Z-2082-200 ZytoLight SPEC FGFR3/4p11 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

53 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 53 FE023-1-20 ZytoLight ® SPEC FGFR3/IGH Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR3/IGH Dual The SPEC FGFR3/IGH Dual Color Dual In a normal interphase nucleus, two Color Dual Fusion Probe is designed to de- Fusion Probe is a mixture of an orange orange and two green signals are tect the translocation t(4;14)(p16.3;q32.3) direct labeled probe spanning the FGFR3 expected. A reciprocal translocation affecting the FGFR3 (fibroblast growth gene region at 4p16.3 and a green direct involving two breakpoints splits the two factor receptor 3, a.k.a. JTK4) gene in labeled probe hybridizing to the IGH signals and generates a fusion signal the chromosomal region 4p16.3 and the gene locus a 14q32.33. on each of the chromosomes involved. IGH (immunoglobulin heavy locus, a.k.a. The chromosomal regions which are not IGH@) locus in 14q32.33. FGFR3 translocated are indicated by the single FGFR3 encodes for a receptor tyrosine ki- orange and green signal, respectively. nase, which regulates downstream signaling cascades after ligand binding. Fusion to several partner genes (including the IGH locus) can lead to a ligand-independent activation of the tyrosine kinase of the resulting FGFR3 fusion protein, frequently found in multiple myeloma (MM). FGFR3/IGH translocations are observed in approximately 15–20% of patients with MM. The breaking points for the 4p16.3 locus are found between the FGFR3 gene and the 5’ end of the NSD2 gene. The t(4;14)(p16.3;q32.3) translocation is associated with upregulation of the FGFR3 IGH SPEC FGFR3/IGH Dual Color Dual Fusion Probe hybridized to normal interphase cells as indicated by and the myeloma NSD2 (a.k.a. MMSET) Ideograms of chromosomes 4 (above) and two orange and two green signals in each nucleus. domain protein. Patients with FGFR3/IGH 14 (below) indicating the hybridization locations. translocation demonstrate an overall poor D4S865 D4S43 prognosis that is only partially mitigated 5’3’ 5’ 3’ by the use of the novel agents bortezomib FGFR3 NSD2 and lenalidomide. ~855 kb With conventional cytogenetics, the Tel 4p16.3 Cen

t(4;14)(p16.3;q32.3) translocation is SPEC FGFR3 Probe map (not to scale). difficult to identify. Thus, the detection BV448316 RH119884 of FGFR3/IGH translocations by Fluo- 3’ 5’ 3’ 5’ rescence in situ Hybridization may be of JAG2 IGH diagnostic and prognostic relevance. ~1.5 Mb References Bergsagel PL & Kuehl WM (2001) Oncogene 20: 5611-22. Cen 14q32.33 Tel Chesi M, et al. (1998) Blood 92: 3025-34. Fabris S, et al. (2005) Genes Chromosomes Cancer 42: 117-27. SPEC IGH Probe map (not to scale). Fenton JA, et al. (2003) Oncogene 22: 1103-13. Kalff A & Spencer A (2012) Blood Cancer J 7: e89. Sonneveld P, et al. (2016) Blood 127: 2955-62. Walker BA, et al. (2013) Blood 121: 3413-19.

Prod. No. Product Label Tests* (Volume) Z-2282-50 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

54 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 54 FE150-1-20 ZytoLight ® SPEC PDGFRA/FIP1L1 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC PDGFRA/FIP1L1 The SPEC PDGFRA/FIP1L1 TriCheck™ In an interphase nucleus lacking a deletion TriCheck™ Probe is designed to detect rear- Probe is a mixture of three direct labeled or translocation involving the 4q12 band, rangements involving the chromosomal region probes hybridizing to the 4q12 band. The two tricolor orange/green/blue fusion 4q12 harboring the PDGFRA gene. The green fluorochrome direct labeled probe signals are expected representing two PDGFRA (platelet-derived growth factor hybridizes proximal to the FIP1L1 gene, normal 4q12 loci. receptor alpha) gene encodes a transmem- the orange fluorochrome direct labeled A PDGFRA-FIP1L1 fusion resulting from an brane glycoprotein that belongs to the type III probe hybridizes distal to the FIP1L1 gene interstitial DNA deletion is indicated by receptor tyrosine kinase family and has a key and proximal to the PDGFRA gene, and the loss of the orange signal leading to a role in a variety of cellular processes. the blue fluorochrome direct labeled probe separate green signal co-localizing with a PDGFRA gene rearrangements are rarely hybridizes distal to the PDGFRA gene. blue signal. genetic events detected in myeloid and A PDGFRA translocation without involve- lymphoid neoplasms. These rearrangements ment of FIP1L1 is indicated by one orange/ most frequently occur in chronic eosinophilic green fusion signal and one separate blue leukemia (CEL), but can be also detected in PDGFRA, FIP1L1 signal. acute myeloid leukemia (AML), and T-lymphoblastic leukemia/lymphoma (T-ALL). The most common gene fusion partner for PDGFRA is the FIP1-like 1 (FIP1L1) gene caused by an 800 kb interstitial deletion on chromosome 4q12. The result of this deletion

is the loss of the CHIC2 gene and the fusion Ideogram of chromosome 4 of the 5’ end of the FIP1L1 gene with the 3’ indicating the hybridization locations. end of the PDGFRA gene. SHGC-112148 RH110746 Although FIP1L1 is the most common fusion RH26448 SHGC-153776 RH77462 D4S769 partner of PDGFRA, five other partner genes 5’ 3’ 3’ 5’ 5’ 3’

have been identified, including BCR, ETV6, FIP1L1 CHIC2 PDGFRA

KIF5B, STRN, and CDK5RAP2. ~685 kb ~400 kb ~730 kb SPEC PDGFRA/FIP1L1 TriCheck™ Probe hybridized to Identification of patients harboring a Cen 4q12 Tel normal interphase cells as indicated by two tricolor orange/green/blue fusion signals per nucleus. PDGFRA rearrangement is important as SPEC PDGFRA/FIP1L1 Probe map (not to scale). these patients respond very well to a targeted therapy with imatinib. In CEL patients harboring a PDGFRA-FIP1L1 fusion a good response to other tyrosine kinase inhibitors like dastinib, nilotinib, sorafenib, and midostaurin could be demonstrated. Hence, detection of PDGFRA rearrange- References Bain BJ (2010) Haematologica 95: 696-8. ments by Fluorescence in situ Hybridization Cools J, et al. (2003) N Engl J Med 348: 1201-14. Curtis CE, et al. (2007) Br J Haematol 138: 77-81. (FISH) may be of diagnostic and predictive Gotlib J, et al. (2004) Blood 103: 2879-91. Savage N, et al. (2013) Int J Lab Hematol 35: 491-500. relevance. Vega F, et al. (2015) Am J Clin Pathol 144: 377-92.

Prod. No. Product Label Tests* (Volume) Z-2209-50 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

55 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 55 FE122-1-20 ZytoLight ® SPEC TERT Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC TERT Dual Color The SPEC TERT Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cation involving the 5p15.33 band, two translocations involving the chromosomal probes hybridizing to the 5p15.33 band. orange/green fusion signals are expected, region 5p15.33 harboring the TERT gene. The orange fluorochrome labeled TERT representing two normal (non-rearranged) The TERT (telomerase reverse transcriptase, probe hybridizes proximal to the TERT 5p15.33 loci. A signal pattern consisting a.k.a. EST2, TCS1) gene encodes the gene, the green fluorochrome labeled of one orange/green fusion signal, one reverse transcriptase component, a cata- TERT probe hybridizes distal to that gene. orange signal, and a separate green lytic domain of the enzyme telomerase. signal indicates one normal 5p15.33 Telomerases are necessary to maintain the TERT locus and one 5p15.33 locus affected by ends of chromosomes and are inactive in a TERT translocation. the majority of somatic cells but active in cancer cells. Remodeling of the genomic context, due to 5p15.33 rearrangements, abrogates transcriptional silencing of the TERT gene. TERT translocations are the second common aberration found in neuroblastomas. These translocations are observed either in a 50 kb region Ideogram of chromosome 5 indicating the hybridization locations. proximal or more rarely in a 40 kb region SHGC-147290 D5S678 distal to the gene. Molecular genetic RH60332 RH41110 3’ 5’ studies showed that rearrangements of the chromosomal region 5p15.33 occur TERT ~540 kb ~235 kb exclusively in high-risk neuroblastomas. SPEC TERT Dual Color Break Apart Probe hybridized Therefore, TERT rearrangements are con- Tel 5p15.33 Cen to normal interphase cells as indicated by two orange/green fusion signals in each nucleus and SPEC TERT Probe map (not to scale). sidered to define a sub-group of high-risk to metaphase chromosomes of a normal cell. neuroblastomas with a poor prognosis. Rearrangements of TERT are found in chromophobe renal cell carcinomas, non-Hodgkin lymphomas, and mantle cell lymphomas. FISH is a suitable tool for the detection of these TERT rearrangements and thus may be of prognostic relevance.

References Davis CF, et al. (2014) Cancer Cell 26: 319-30. Nagel I, et al. (2010) Blood 116: 1317-20. Pfeifer M, et al. (2015) Nature 526: 700-4. Schilling G, et al. (2013) Leukemia Res 37: 280-6.

Prod. No. Product Label Tests* (Volume) Z-2273-50 ZytoLight SPEC TERT Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

56 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 56 FE142-1-20 ZytoLight ® SPEC TERT/5q31 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC TERT/5q31 Dual The SPEC TERT/5q31 Dual Color Probe is In a normal interphase nucleus two orange Color Probe is designed for the detection a mixture of a green fluorochrome direct and two green signals are expected. of TERT gene amplifications and chromo- labeled SPEC TERT probe hybridizing to In a cell with amplification of the TERT somal gains found in a variety of human the TERT gene in the chromosomal region gene locus or aneuploidy of chromosome tumors. 5p15.33 and an orange fluorochrome di- 5, multiple copies of the green signal or The TERT (telomerase reverse transcriptase) rect labeled SPEC 5q31 probe specific for green signal clusters will be observed. gene is located in the chromosomal region the chromosomal region 5q31.2 harbor- 5p15.33 and encodes the reverse tran- ing the EGR1 gene. Since chromosomes scriptase component of the human telomer- 1, 5, and 19 share the same repetitive ase. Telomerase, the ribonucleoprotein sequences, probes specific for 5q31.2 are enzyme complex necessary to maintain commonly used for chromosome 5 copy the ends of chromosomes, is absent from number detection. the majority of somatic cells but is present and active in the majority of immortal cell lines and human cancers. Chromosomal gain or amplification of the TERT gene was found in various human tumors such as lung, cervical, bladder,

breast, hepatocellular and colorectal SPEC TERT/5q31 Dual Color Probe hybridized carcinomas as well as in neuroblastoma to melanoma tissue section showing normal cells as indicated by two green and two orange signals in and melanoma. It was shown that TERT each nucleus and cells with TERT gene amplification amplification is a poor prognostic factor as indicated by multiple green signals per nucleus. in non-small cell lung cancer (NSCLC) and Ideogram of chromosome 5 indicating the hybridization locations. is associated with poorly differentiated D10S18 RH41110 histopathology of hepatocellular carcino- 3’ 5’ mas. Thus, detection of TERT amplification may have useful applications in cancer TERT diagnosis and prognosis. ~615 kb Tel 5p15.33 Cen References Bryce LA, et al. (2000) Neoplasia 2: 197-201. SPEC 5p15 Probe map (not to scale). Cao Y, et al. (2008) Cancer Sci 99: 1092-9. Morin GB (1989) Nature 353: 454-6. Takuma Y, et al. (2004) J Gastroenterol Hepatol 19: 1300-4. RH48032 RH111439 Zhu C-Q, et al. (2006) Br J Cancer 94: 1452-9. 5’3’

EGR1

~605 kb Cen 5q31.2 Tel

SPEC 5q31 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2091-50 ZytoLight SPEC TERT/5q31 Dual Color Probe / 5 (50 μl) Z-2091-200 ZytoLight SPEC TERT/5q31 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

57 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 57 FE026-1-20 ZytoLight ® SPEC RICTOR/5q31.1 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC RICTOR/5q31.1 The SPEC RICTOR/5q31.1 Dual Color Using the SPEC RICTOR/5q31.1 Dual Dual Color Probe is designed to detect Probe is a mixture of a green fluorochrome Color Probe in a normal interphase amplifications affecting the chromosomal direct labeled probe spanning the RICTOR nucleus, two orange and two green region 5p13.1 harboring the RICTOR gene region at 5p13.1 and an orange signals are expected. In a cell with gain (RPTOR independent companion of MTOR fluorochrome direct labeled SPEC 5q31.1 of the RICTOR gene locus, multiple copies complex 2, a.k.a. AVO3, KIAA1999) probe specific for the chromosomal region of the green signal or green signal clusters gene. 5q31.1 harboring the ZCCHC10 gene. will be observed. RICTOR is part of the RICTOR-mTOR protein complex which is involved in the phosphorylation and regulation of the RICTOR AKT/Protein Kinase B (PKB) which in turn triggers cell proliferation and cell survival. Amplification of the RICTOR gene region leads to a dysregulation of PKB, 5q31.1 known to be a responsible factor in tumor pathogenesis. In 8-13% of non-small cell lung carcinomas (NSCLC) and small cell Ideogram of chromosome 5 indicating the hybridization locations. lung cancer (SCLC) an amplification

of the RICTOR gene region has been RH113556 RH76719 SPEC RICTOR/5q31.1 Dual Color Probe hybridized 3’ 5’ demonstrated. Amplifications of the to normal interphase cells as indicated by two orange and two green signals in each nucleus and RICTOR gene region represent one of the RICTOR to metaphase chromosomes of a normal cell. most abundant genetic events in SCLC ~610 kb patients. Hence, RICTOR amplifications Tel 5p13.1 Cen have been proposed as biomarker for a SPEC RICTOR Probe map (not to scale). targeted therapy with mTOR inhibitory REN71705 D5S2456 agents in patients diagnosed with lung 3’ 5’ cancer. Fluorescence in situ Hybridization ZCCHC10 may be a suitable diagnostic tool for the ~660 kb detection of RICTOR amplifications. Cen 5q31.1 Tel

References SPEC 5q31.1 Probe map (not to scale). Cheng H, et al. (2015) Cancer Discov 5: 1262-70. Ross JS, et al. (2014) J Clin Pathol 67: 772-6. Sarbassov DD, et al. (2005) Science 307: 1098-101.

Squamous cell carcinoma section with RICTOR amplification as indicated by multiple green signals in each nucleus.

Kindly provided by Prof. Dr. Schildhaus, Essen, Germany.

Prod. No. Product Label Tests* (Volume) Z-2278-200 ZytoLight SPEC RICTOR/5q31.1 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

58 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 58 FE157-1-20 ZytoLight ® SPEC EGR1/5p15 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC EGR1/5p15 Dual The SPEC EGR1/5p15 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of a green fluorochrome direct ange and two green signals are expected. of EGR1 gene deletions. labeled SPEC EGR1 probe hybridizing to In a cell with deletions affecting the EGR1 The EGR1 (early growth response 1) gene the EGR1 gene in the chromosomal region gene locus, one or no copy of the green is located in the chromosomal region 5q31.2 and an orange fluorochrome signal will be observed. 5q31.2. Deletions spanning the region direct labeled SPEC 5p15 probe specific 5q31.2 are among the most common for the chromosomal region 5p15.33. reoccurring abnormalities detectable in myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). The EGR1 protein belongs to the EGR family of C2H2-type zinc-finger proteins. It is a nuclear protein and functions as a transcriptional regulator. Deletion of EGR1 in estrogen receptor negative (ER-) breast carcinomas is correlated with a higher tumor grade, sug- Ideogram of chromosome 5 gesting that loss of the EGR1 gene (and indicating the hybridization locations. SPEC EGR1/5p15 Dual Color Probe hybridized thereby loss of functioning EGR1 protein) to normal interphase cells as indicated by two RH47863 RH67940 orange and two green signals in each nucleus. may contribute to the pathogenesis of ER- 5’3’ breast carcinomas. In patients with therapy-related MDS and EGR1 AML, dicentric chromosomes have often ~700 kb Cen 5q31.2 Tel been observed. In such conditions, many SPEC EGR1 Probe map (not to scale). patients show a complex karyotype with several marker chromosomes unidentifia- D10S18 RH41110 3’ 5’ ble by conventional cytogenetics. Fluo- rescence in situ Hybridization (FISH) has TERT now made the characterization of these ~615 kb rearrangements much easier. Tel 5p15.33 Cen

References SPEC 5p15 Probe map (not to scale). Graubert TA, et al. (2009) PLoS One 4: e4583. Herry A, et al. (2007) Cancer Genet Cytogenet 175: 125-31. Ronski K, et al. (2005) Cancer 104: 925-30. Sun Y & Cook JR (2010) Leuk Res 34: 340-3. SPEC EGR1/5p15 Dual Color Probe hybridized to bone marrow biopsy section with deletion of the EGR1 gene as indicated by one green signal and two orange signals in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2107-50 ZytoLight SPEC EGR1/5p15 Dual Color Probe / 5 (50 μl) Z-2107-200 ZytoLight SPEC EGR1/5p15 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

59 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 59 FE035-1-20 ZytoLight ® SPEC EGR1/D5S23,D5S721 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC EGR1/D5S23, The SPEC EGR1/D5S23,D5S721 Dual In a normal interphase nucleus, two or- D5S721 Dual Color Probe is designed for Color Probe is a mixture of a green ange and two green signals are expected. the detection of EGR1 gene deletions. fluorochrome direct labeled SPEC EGR1 In a cell with deletion of the EGR1 gene The EGR1 (early growth response 1) gene probe hybridizing to the EGR1 gene in locus, one or no copy of the green signal is located in the chromosomal region the chromosomal region 5q31.2 and an will be observed. 5q31.2 and encodes a zinc finger tran- orange fluorochrome direct labeled SPEC scription factor which is associated with cell D5S23,D5S721 probe specific for the proliferation, differentiation, and transfor- chromosomal region 5p15.2-p15.31. mation. Since in diverse solid tumors the chro- Deletions spanning the region 5q31.2 are mosomal region 5p15.33 is affected among the most common reoccurring abnor- by amplifications, probes targeting the malities detectable in myelodysplastic syn- D5S23,D5S721 region are more suitable dromes (MDS) and acute myeloid leukemia for the enumeration of chromosome 5. (AML). In therapy-related MDS or AML, 40% of the patients exhibit a 5q deletion. D5S23, D5S721 Deletion of EGR1 in estrogen receptor negative (ER-) breast carcinomas is correlated with a higher tumor grade, suggesting that SPEC EGR1/D5S23,D5S721 Dual Color Probe loss of the EGR1 gene may contribute to the hybridized to normal interphase calls as indicated by two orange and two green signals in each nucleus. pathogenesis of ER- breast carcinomas. Transfusion-dependent, lower-risk MDS EGR1 patients with 5q deletion are treated with the thalidomide analog lenalidomide which Ideogram of chromosome 5 is approved by the FDA. indicating the hybridization locations. In patients with de novo or therapy-related RH47863 RH67940 MDS and AML, dicentric chromosomes, 5’3’ involving chromosome 5, have often been observed. These patients frequently show a EGR1 complex karyotype. In such conditions the ~700 kb Cen 5q31.2 Tel characterization of rearrangement is nearly SPEC EGR1 Probe map (not to scale). not feasible by conventional cytogenetics. Hence, Fluorescence in situ Hybridization D5S1880 D5S721 D5S23 D5S1030 SPEC EGR1/D5S23,D5S721 Dual Color Probe (FISH) may be a helpful tool for diagnosis hybridized to an ALL specimen with deletion of and therapy decisions. the EGR1 gene as indicated by one green and two orange signals in each nucleus. References ~735 kb Boultwood J, et al. (2010) Blood 116: 5803-11. Specimens kindly provided by Paediatric Oncology/Haematology, Coleman JF, et al. (2011) Am J Clin Pathol 135: 915-20. Tel 5p15.2-p15.31 Cen Charité - Universitätsmedizin Berlin, Germany. Herry A, et al. (2007) Cancer Genet Cytogenet 175: 125-31. Horrigan SK, et al. (2000) Blood 95: 2372-7. SPEC D5S23,D5S721 Probe map (not to scale). Sukhatme V, et al. (1988) Cell 53: 37-43. Sun Y & Cook JR (2010) Leuk Res 34: 340-3. Tian J, et al. (2016) Intractable Rare Dis Res 5: 76-82. Wei S, et al. (2009) Proc Natl Acad Sci U S A 106: 12974-9. Zhao N, et al. (1997) Proc Natl Acad Sci U S A 94: 6948-53.

Prod. No. Product Label Tests* (Volume) Z-2211-50 ZytoLight SPEC EGR1/D5S23,D5S721 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

60 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 60 FE127-1-20 ZytoLight ® SPEC CSF1R Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CSF1R Dual Color The SPEC CSF1R Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving the rearrangements involving the chromo- probes hybridizing to the 5q32-q33.1 5q32-q33.1 band, two orange/green somal region 5q32 harboring the CSF1R band. The orange fluorochrome direct fusion signals are expected representing (colony stimulating factor 1 receptor, labeled probe hybridizes proximal to the two normal (non-rearranged) 5q32-q33.1 a.k.a. FMS) gene. CSF1R gene at 5q32, the green fluoro- loci. A signal pattern consisting of one The CSF1 receptor is activated by dimer- chrome direct labeled probe hybridizes orange/green fusion signal, one orange ization upon binding of its ligand CSF1 distal to the CSF1R gene at 5q32-q33.1. signal, and a separate green signal and is involved in macrophage develop- indicates one normal 5q32-q33.1 locus ment. and one 5q32-q33.1 locus affected by a Rearrangement of the CSF1R gene was translocation. first detected in an acute megakaryoblas- Duplication of the 5q32 locus will result in tic leukemia (AMKL) cell line generating additional orange signals. the RBM6-CSF1R fusion gene. A MEF2D- CSF1R fusion gene was described in a patient with primary pre-B cell acute CSF1R lymphoblastic leukemia (pre-B ALL). Both fusion proteins contain the intact kinase Ideogram of chromosome 5 domain of CSF1R. indicating the hybridization locations. Philadelphia chromosome-like ALL (Ph-like STSG34261 BV208428 ALL) is a subgroup of B-cell precursor SHGC-149706 RH25007 ALL and is associated with a high risk of 3’ 5’

treatment failure. SSBP2-CSF1R fusions CSF1R PDGFRB were detected in some patients with Ph-like ~365 kb ~570 kb ALL. They result from either the balanced Cen 5q32-q33.1 Tel SPEC CSF1R Dual Color Break Apart Probe translocation t(5;5)(q14;q32) or the dupli- hybridized to normal interphase cells as indicated SPEC CSF1R Probe map (not to scale). by two orange/green fusion signals per nucleus. cation dup(5)(q14q32). Expression of this fusion gene results in cytokine-independent growth and enhanced STAT5 activation which are inhibited by dasatinib in vitro. CSF1R signaling was also shown to be suppressed by the ABL1 kinase inhibitor imatinib. Hence, the detection of CSF1R rearrangements by FISH may help in selecting ALL patients eligible for treatment with CSF1R inhibitors. References Dewar AL, et al. (2005) Blood 105: 3127-32. Blood smear with translocation of the CSF1R gene as Gu TL, et al. (2007) Blood 110: 323-33. indicated by one non-rearranged orange/green fusion Lilljebjörn H, et al. (2014) Leukemia 28: 977-9. Roberts KG, et al. (2014) N Engl J Med 371: 1005-15. signal, one orange and one separate green signal. Schwab C, et al. (2014) Blood 124: 3773.

Prod. No. Product Label Tests* (Volume) Z-2202-50 ZytoLight SPEC CSF1R Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

61 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 61 FE109-1-20 ZytoLight ® SPEC CSF1R/D5S23,D5S721 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC CSF1R/ The SPEC CSF1R/D5S23,D5S721 Dual In a normal interphase nucleus, two or- D5S23,D5S721 Dual Color Probe is de- Color Probe is a mixture of a green fluoro- ange and two green signals are expected. signed for the detection of 5q deletions. chrome direct labeled SPEC CSF1R probe In a cell with deletion of the CSF1R gene The CSF1R (colony stimulating factor 1 hybridizing to the CSF1R gene in the locus, one or no copy of the green signal receptor, a.k.a. C-FMS) gene is located in chromosomal region 5q32-q33.1 and an will be observed. the chromosomal region 5q32. orange fluorochrome direct labeled SPEC The interstitial deletion of chromosome 5q D5S23,D5S721 probe specific for the is a characteristic hallmark of the myelo- chromosomal region 5p15.2-p15.31. dysplastic syndrome (MDS) with isolated del(5q). The size of the deletion as well D5S23,D5S721 as the breakpoints are variable but a commonly deleted region (CDR) has been narrowed to the approximately 1.5 Mb in- terval at 5q32-q33.1 flanked by the DNA marker D5S413 and the GLRA1 gene. One candidate gene for the development CSF1R of MDS in patients with 5q- syndrome is

RPS14 (ribosomal protein 14), a tumor SPEC CSF1R/D5S23,D5S721 Dual Color Probe suppressor gene located in the chromo- Ideogram of chromosome 5 hybridized to normal interphase cells as indicated by indicating the hybridization locations. two orange and two green signals in each nucleus somal region 5q33.1. Haploinsufficiency and to metaphase chromosomes of a normal cell. BV166744 D5S636 (caused by hemizygous deletion) of RPS14 3’ 5’ 3’ 5’ is the probable cause of the erythroid defect that characterizes the 5q- syn- CSF1R RPS14 drome. Lenalidomide has been reported ~ 640 kb Cen 5q32-q33.1 Tel to overcome the pathogenic effect of 5q deletion in MDS. SPEC CSF1R Probe map (not to scale).

Despite the severe phenotype of the 5q- D5S1880 D5S721 D5S23 D5S1030 syndrome, it has a relatively low (10%) transformation risk to acute myeloid

leukemia (AML). Therefore, FISH may be ~735 kb a helpful tool for diagnosis and therapy Tel 5p15.2-p15.31 Cen decision. SPEC D5S23,D5S721 Probe map (not to scale). References Bone marrow biopsy tissue section of an Boultwood J, et al. (1991) Proc Natl Acad Sci U S A 88: 6176-80. Boultwood J, et al. (2010) Blood 116: 5803-11. ALL case showing hemizygous deletion of Giagounidis AA, et al. (2004) Clin Cancer Res 12: 5-10. the CSF1R gene as indicated by the loss Van den Berghe H & Michaux JL (1974) Nature 251: 437-8. of one green signal in each nucleus. Swerdlow SH, et al. (ed.) (2008) WHO classification of tumours of haematopoietic and lymphoid tissues.

Prod. No. Product Label Tests* (Volume) Z-2268-50 ZytoLight SPEC CSF1R/D5S23,D5S721 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

62 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 62 FE141-1-20 ZytoLight ® SPEC PDGFRB Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC PDGFRB Dual Color The SPEC PDGFRB Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 5q32-q33.1 band, translocations involving the chromosomal probes hybridizing to the 5q32-q33.1 two orange/green fusion signals are ex- region 5q32 harboring the PDGFRB gene. band. The green fluorochrome direct labe- pected representing two normal (non-rear- The PDGFRB (platelet derived growth led probe hybridizes distal to the PDGFRB ranged) 5q32-q33.1 loci. A signal pattern factor receptor beta) gene encodes a gene, and the orange fluorochrome direct consisting of one orange/green fusion transmembrane glycoprotein that belongs labeled probe hybridizes proximal to the signal, one orange signal, and a sepa- to the type III receptor tyrosine kinase PDGFRB locus. rate green signal indicates one normal family and has a key role in a variety of 5q32-q33.1 locus and one 5q32-q33.1 cellular processes. locus affected by a translocation. Translocations involving the PDGFRB gene are rare genetic disorders and are identified in myelodysplastic/myeloproliferative neoplasms (MDS/MPNs), chronic myeloproliferative disorders (CMPD), acute myeloid leukemia (AML), and also PDGFRB in atypical (BCR-ABL1-negative) chronic myeloid leukemia/chronic myelomonocytic leukemia (CML/CMML)-like diseases, Ideogram of chromosome 5 indicating the hybridization locations. often with eosinophilia and splenomegaly. The most common translocation involving GDB:181638 BV208428 SHGC-149706 RH76992 PDGFRB is the t(5;12)(q32;p13.2). Result 3’ 5’ of this translocation is the fusion protein CSF1R PDGFRB SPEC PDGFRB Dual Color Break Apart Probe ETV6-PDGFRB, in which the pointed hybridized to normal interphase cells as indicated ~415 kb ~735 kb by two orange/green fusion signals per nucleus. domain of ETV6 is juxtaposed next to Cen 5q32-q33.1 Tel the transmembrane and entire tyrosine SPEC PDGFRB Probe map (not to scale). kinase domain of PDGFRB. As a result, the tyrosine kinase is constitutively activated leading to hematopoietic cell proliferation. Patients with myeloid malignancies bear- ing PDGFRB fusion genes were shown to achieve durable long-term remissions under imatinib treatment. Recent studies revealed that sorafenib is a further potential inhibitor of patients with ETV6-PDGFRB translocation. References Bain BJ (2010) Haematologica 95: 696-8. Cross NC & Reiter A (2008) Acta Haematol 119: 199-206. Jones AV & Cross NC (2004) Cell Mol Life Sci 61: 2912-23. Keene P, et al. (1987) Br J Haematol 67: 25-31. Lierman E, et al. (2007) Haematologica 92: 27-34. Savage N, et al. (2013) Int J Lab Hematol 35: 491-500. Steer EJ & Cross NC (2002) Acta Haematol 107: 113-22. Vega F, et al. (2015) Am J Clin Pathol 144: 377-92.

Prod. No. Product Label Tests* (Volume) Z-2197-50 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

63 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 63 FE103-1-20 ZytoLight ® SPEC IRF4,DUSP22 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC IRF4,DUSP22 Dual The SPEC IRF4,DUSP22 Dual Color Break In an interphase nucleus lacking a trans- Color Break Apart Probe is designed Apart Probe is a mixture of two direct location involving the 6p25.3 band, two for the detection of translocations involv- labeled probes hybridizing to the 6p25.3 orange/green fusion signals are expected ing the chromosomal region 6p25.3 band. The green fluorochrome direct la- representing two normal (non-rearranged) harboring the DUSP22 (dual specificity beled probe hybridizes proximal and the 6p25.3 loci. A signal pattern consisting phosphatase 22, a.k.a. JKAP) and IRF4 orange fluorochrome direct labeled probe of one orange/green fusion signal, one (interferon regulatory factor 4, a.k.a. hybridizes distal to the IRF4 and DUSP22 orange signal, and a separate green MUM1) genes. genes. signal indicates one normal 6p25.3 locus IRF4 is normally expressed in plasma cells, and one 6p25.3 locus affected by a IRF4, DUSP22 melanocytes, some B-cells, and in activat- translocation. ed T-cells. The IRF4 protein is required at several stages of B-cell development, and is also critical for T-cell differentiation. Rearrangements of the IRF4/DUSP22 chromosomal region have been detected in various B-cell and T-cell lymphomas. Large B-cell lymphoma (LBCL) with IRF4 rearrangement, which occurs most com- Ideogram of chromosome 6 monly in children and young adults, is indicating the hybridization locations. considered a distinct new provisional enti- RH40877 RH45976 ty. These lymphomas most typically occur RH12503 RH120056 5’ 3’5’3’ in Waldeyer ring and/or cervical lymph

nodes. Most cases have IG/IRF4 fusions DUSP22 IRF4 SPEC IRF4, DUSP22 Dual Color Break Apart Probe and have a favorable prognosis. hybridized to normal interphase cells as indicated by ~160 kb ~385 kb two orange/green fusion signals per nucleus. Rearrangements of IRF4 and/or DUSP22 Tel 6p25.3 Cen have also been described in peripher- SPEC IRF4, DUSP22 Probe map (not to scale). al T-cell lymphomas and in cutaneous anaplastic large cell lymphoma (ALCL). ALCL is difficult to distinguish from other CD30-positive T-cell lymphoproliferative disorders. IRF4 translocation has a high specificity for cutaneous ALCL supporting the clinical utility of FISH for IRF4 in the differential diagnosis of T-cell lymphoproliferative disorders.

Moreover, DUSP22 rearrangement in References Iida S, et al. (1997) Nat Genet 17: 226-30. ALK-negative ALCL is associated with a fa- Parrilla Castellar ER, et al. (2014) Blood 124: 1473-80. Pham-Ledard A, et al. (2010) J Invest Dermatol 130: 816-25. T-cell lymphoma tissue section with translocation vorable outcome indicating the usefulness Salaverria I, et al. (2011) Blood 118: 139-47. affecting the 6p25.3 locus as indicated by one of DUSP22 as a predictive biomarker. Swerdlow SH, et al. (2016) Blood 127: 2375-90. non-rearranged orange/green fusion signal, one Wada DA, et al. (2011) Mod Pathol 24: 596-605. orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2210-50 ZytoLight SPEC IRF4,DUSP22 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

64 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 64 FE124-1-20 ZytoLight ® SPEC RREB1/MYB/CEN 6 Triple Color Probe

Background Probe Description Results The ZytoLight ® SPEC RREB1/MYB/CEN 6 The SPEC RREB1/MYB/CEN 6 Triple In a normal interphase nucleus, two green, Triple Color Probe is designed for the Color Probe is a mixture of a green two orange, and two blue signals are detection of copy number changes of the fluorochrome direct labeled SPEC RREB1 expected. In a cell with amplification of chromosomal regions harboring the RREB1 probe hybridizing to the RREB1 locus at the RREB1 or the MYB gene locus, multiple and the MYB gene, respectively. 6p24.3-p25.1, an orange fluorochrome copies of the green or orange signal will The RREB1 (ras responsive element bind- direct labeled SPEC MYB probe hybrid- be observed, respectively. In a cell with ing protein 1, a.k.a. HNT) gene is located izing to the MYB locus at 6q23.2-q23.3, deletion of the RREB1 or the MYB gene lo- in 6p24.3 and encodes a zinc finger tran- and a blue fluorochrome direct labeled cus, a reduced number of green or orange scription factor. The MYB (MYB proto-on- CEN 6 probe specific for the alpha satel- signals will be observed, respectively. cogene, transcription factor, a.k.a. c-myb) lite centromeric region of chromosome 6 gene is located in 6q23.3 and encodes (D6Z1). a transcription factor that is implicated in RREB1 proliferation, survival, and differentiation of hematopoietic progenitor cells. Overexpression of the RREB1 protein CEN 6 (D6Z1) was detected in prostate cancer and in a medullary thyroid cancer cell line. RREB1 is suggested to play a role in Ras and Raf signal transduction in medullary thyroid MYB cancer. MYB has been found to be amplified in a Ideogram of chromosome 6 variety of human cancers. In pancreatic indicating the hybridization locations. SPEC RREB1/MYB/CEN 6 Triple Color cancer, MYB amplification was mainly Probe hybridized to normal interphase cells RH110101 D8S1159 as indicated by two green, two orange, found in advanced and metastatic tumors. 5’ 3’ and two blue signals in each nucleus. In breast tumors from BRCA1 germline RREB1 mutation carriers, MYB amplification was ~495 kb observed in 29% of the cases and resulted Tel 6p24.3-25.1 Cen in overexpression of the MYB protein. SPEC RREB1 Probe map (not to scale). Moreover, duplication of the MYB gene D6S1976 RH33702 occurs in 8.4% of individuals with T-cell 5’ 3’ acute lymphoblastic leukemia (T-ALL). MYB References Kauraniemi P, et al. (2000) Cancer Res 60: 5323-8. ~575 kb Lahortiga I, et al. (2007) Nat Genet 39: 593-5. Thiagalingam A, et al. (1996) Mol Cell Biol 16: 5335-45. Cen 6q23.2-q23.3 Tel Wallrapp C, et al. (1997) Cancer Res 57: 3135-9. Zou J, et al. (2011) Prostate 71: 1518-24. SPEC MYB Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2152-50 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe / / 5 (50 μl) Z-2152-200 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe / / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

65 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 65 FE113-1-20 ZytoLight ® SPEC PHF1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC PHF1 Dual Color The SPEC PHF1 Dual Color Break In an interphase nucleus lacking a trans- Break Apart Probe is designed for the Apart Probe is a mixture of two di- location involving the 6p21.31-p21.32 detection of translocations involving the rect labeled probes hybridizing to the band, two orange/green fusion signals chromosomal region 6p21.32 harboring 6p21.31-p21.32 band. The green fluo- are expected representing two normal the PHF1 (PHD finger protein 1, a.k.a. rochrome direct labeled probe hybridizes (non-rearranged) 6p21.31-p21.32 loci. A MTF2L2, PCL1) gene. The PHF1 protein in 6p21.31-p21.32 proximal and the signal pattern consisting of one orange/ is known to affect processes, such as de- orange fluorochrome direct labeled probe green fusion signal, one orange signal, velopment and cell proliferation, through hybridizes in 6p21.32 distal to the PHF1 and a separate green signal indicates modulation of histone H3 methylation. gene. one normal 6p21.31-p21.32 locus and Endometrial stromal tumors (ESTs) are the one 6p21.31-p21.32 locus affected by a second most common pure mesenchy- translocation. mal tumors of the uterus. ESTs may pose PHF1 Deletion of 5‘-PHF1 sequences is indicated diagnostic challenges particularly when by one or multiple isolated green signals. they exhibit variant histologic appearanc- es, involve extrauterine sites, or present as metastatic disease. Several rearrangements involving the genes JAZF1, PHF1, or YWHAE have been identified in ESTs, detection of which may be helpful in the Ideogram of chromosome 6 differential diagnosis of these tumors. indicating the hybridization locations. PHF1 rearrangements were found to occur in endometrial stromal sarcomas but not in BV448045 BV447978 BV208812 SHGC-152483 endometrial stromal nodules or undifferen- 5’ 3’ tiated endometrial sarcomas. Moreover, recurrent rearrangements of the PHF1 ~195 kb ~455 kb PHF1 gene have also been detected in up SPEC PHF1 Dual Color Break Apart Probe hybridized Tel 6p21.31-p21.32 Cen to 85% of ossifying fibromyxoid tumors to normal interphase cells as indicated by two SPEC PHF1 Probe map (not to scale). orange/green fusion signals per nucleus and (OFMTs) including benign and malignant to metaphase chromosomes of a normal cell. cases. Thus, FISH analysis for the detection of PHF1 translocation may also serve as a diagnostic tool to identify OFMT cases. References Antonescu CR, et al. (2014) Genes Chromosomes Cancer 53: 183-93. D‘Angelo E, et al. (2013) Am J Surg Pathol 37: 514-21. Gebre-Medhin S, et al. (2012) Am J Pathol 181: 1069-77. Hodge JC, et al. (2016) J Mol Diagn 18: 516-26. Micci F, et al. (2006) Cancer Res 66: 107-12.

Sarcoma tissue section with translocation of the PHF1 gene as indicated by one non-rearranged orange/green fusion signal, one orange, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2215-50 ZytoLight SPEC PHF1 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

66 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 66 FE136-1-20 ZytoLight ® SPEC VEGFA/CEN 6 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC VEGFA/CEN 6 Dual The SPEC VEGFA/CEN 6 Dual Color In a normal interphase nucleus, two or- Color Probe is designed for the detection Probe is a mixture of a green fluoro- ange and two green signals are expected. of amplifications involving the chromosom- chrome direct labeled SPEC VEGFA probe In a cell with amplification of the VEGFA al region 6p21.1 harboring the VEGFA specific for the VEGFA gene at 6p21.1 gene locus, multiple copies of the green gene (vascular endothelial growth factor and an orange fluorochrome direct labeled signal or large green signal clusters will A, a.k.a. VEGF, VPF). CEN 6 probe specific for the alpha satel- be observed. The VEGFA protein is involved in vascular lite centromeric region of chromosome 6 permeability, angiogenesis, cell migration, (D6Z1). and inhibition of apoptosis. In addition, binding of VEGFA to its receptors activates the RAS/MEK/MAPK pathway, thus, lead- VEGFA ing to mitotic activation. CEN 6 (D6Z1) Amplification of the VEGFA gene locus was found in several types of malignancy, such as osteosarcoma, hepatocellular carcinoma (HCC), and colorectal cancers. In patients with osteosarcoma, VEGFA gene amplification results in elevated ex- Ideogram of chromosome 6 pression of VEGFA and is associated with indicating the hybridization locations. SPEC VEGFA/CEN 6 Dual Color Probe hybridized to adverse tumor-free survival. normal interphase calls as indicated by two orange D6S1173E SHGC-155289 and two green signals in each nucleus. VEGFA amplifications occur in 3-6% of 5’ 3’ colorectal cancers and result in a highly aggressive disease. VEGFA HCC patients with VEGFA gain responded ~352 kb Tel 6p21.1 Cen better to sorafenib, a multi-kinase inhibitor SPEC VEGFA Probe map (not to scale). that blocks, i.a., receptors of the VEGFA protein, resulting in improved survival of the patients. This suggests that VEGFA is a potential biomarker for response to sorafenib therapy in HCC. Hence, detection of VEGFA amplifications by Fluorescence in situ Hybridization may help in selecting patients eligible for an HCC tissue section with interphase cells anti-VEGFA therapy. showing a polysomy of chromosome 6 as References indicated by multiple green (VEGFA) and Horwitz E, et al. (2014) Cancer Discov 4: 730-43. orange (CEN 6) signals in each nucleus. Vlajnic T, et al. (2011) Mod Pathol 24: 1404-12. Yang J, et al. (2011) Cancer 117: 4925-38.

Prod. No. Product Label Tests* (Volume) Z-2195-200 ZytoLight SPEC VEGFA/CEN 6 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

67 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 67 FE098-1-20 ZytoLight ® SPEC ROS1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ROS1 Dual Color The SPEC ROS1 Dual Color Break Apart In an interphase nucleus lacking an aber- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled ration involving the 6q22.1 band, two translocations involving the chromosomal probes hybridizing to the 6q22.1 band. orange/green fusion signals are expected region 6q22.1 harboring the ROS pro- The orange fluorochrome direct labeled representing two normal (non-rearranged) to-oncogene 1, receptor tyrosine kinase probe hybridizes distal, the green fluoro- 6q22.1 loci. A signal pattern consisting (ROS1, a.k.a. MCF3) gene. chrome direct labeled probe hybridizes of one orange/green fusion signal, one The ROS1 gene is located on 6q22.1 proximal to the ROS1 breakpoint region orange signal, and a separate green signal and encodes a receptor tyrosine kinase. at 6q22.1. indicates one normal 6q22.1 locus and one Translocations affecting ROS1 have been 6q22.1 locus affected by a translocation. detected in glioblastoma, cholangiocar- Isolated green signals are the result of dele- cinoma, and non-small cell lung cancer tions distal to the ROS1 breakpoint region (NSCLC). or are due to unbalanced translocations In NSCLC several ROS1 translocation part- affecting this chromosomal region. ners have been detected all of which result in the fusion of variably truncated forms of ROS1 e.g. TPM3, SDC4, SLC34A2, CD74, EZR, or LRIG3 to the kinase domain of ROS1. GOPC has also been found to be fused Ideogram of chromosome 6 to ROS1 in NSCLC. GOPC-ROS1 fusions indicating the hybridization locations. result from interstitial deletion of approx. RH69070 RH104060 240 kb on 6q22.1. D6S1391E RH78348 ROS1 rearrangements are thought to 3’ 5’ 3’5’

define a molecular subset of NSCLC with ROS1 GOPC

distinct clinical characteristics that are sim- ~715 kb ~215 kb ilar to those observed in patients with ALK Cen 6q22.1 Tel SPEC ROS1 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated rearranged NSCLC. SPEC ROS1 Probe map (not to scale). by two orange/green fusion signals per nucleus. First evidence suggests that administration of ROS1 kinase inhibitors may represent a very effective therapeutic strategy in NSCLC patients harboring activating ROS1 rearrangements. Accordingly, detection of ROS1 rearrangements using Fluorescence in situ Hybridization might be a helpful tool for the identification of patients likely to respond to ROS1 kinase targeting therapies.

References Bergethon K, et al. (2012) J Clin Oncol 30: 863-70. Birchmaier C, et al. (1987) Proc Natl Acad Sci U S A 84: 9270-4. Bos M, et al. (2013) Lung Cancer 81: 142-3. Section of paraffin embedded NSCLC cell line Lee SE, et al. (2015) Mod Pathol 28: 468-79. with translocation affecting the 6q22.1 locus Rikova K, et al. (2007) Cell 131: 1190-203. Rimkunas VM, et al. (2012) Clin Cancer Res 18: 4449-57. harboring ROS1 as indicated by one orange/ Suehara Y, et al. (2012) Clin Cancer Res 18: 6599-608. green fusion signal (non-rearranged), one orange Takeuchi K, et al. (2012) Nat Med 18: 378-81. signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2144-50 ZytoLight SPEC ROS1 Dual Color Break Apart Probe / 5 (50 μl) Z-2144-200 ZytoLight SPEC ROS1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

68 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 68 FE061-1-20 ZytoLight ® SPEC ROS1/CEN 6 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ROS1/CEN 6 Dual The SPEC ROS1/CEN 6 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of amplifications of the chromosomal direct labeled CEN 6 probe specific for In a cell with amplification of the ROS1 region harboring the ROS1 gene. the alpha satellite centromeric region of gene locus, multiple copies of the green The ROS1 (ROS proto-oncogene 1, recep- chromosome 6 (D6Z1) and a green fluoro- signal or green signal clusters will be tor tyrosine kinase, a.k.a. MCF3) gene is chrome direct labeled SPEC ROS1 probe observed. located on 6q22.1 and encodes a recep- specific for the ROS1 gene at 6q22.1. tor tyrosine kinase of the insulin receptor family. ROS1 has been found to undergo genetic rearrangements in a variety of human cancers including glioblastoma, CEN 6 (D6Z1) cholangiocarcinoma, and non-small cell lung cancer (NSCLC). ROS1 rearrangements, detected in ade- ROS1 nocarcinoma of the lung, are thought to define a molecular subset of NSCLC with distinct clinical characteristics that are Ideogram of chromosome 6 similar to those observed in patients with indicating the hybridization locations. ALK rearranged NSCLC. Targeting ROS1 SPEC ROS1/CEN 6 Dual Color Probe hybridized BV208912 RH47666 fusion proteins with the kinase inhibitor 3’ 5’ to normal interphase cells as indicated by two orange and two green signals in each nucleus. crizotinib was shown to be a promising ROS1 and effective therapy in NSCLC patients ~465 kb whose tumors are positive for this genetic Cen 6q22.1 Tel aberration. SPEC ROS1 Probe map (not to scale). Recently, copy number gain of the ROS1 gene was reported to occur in NSCLC patients and to be associated with poor prognosis. Hence, detection of ROS1 amplification by FISH could help to identify patients who might be selected for further clinical examinations with regard to potential ROS1 targeting treatments. Lung cancer tissue section with interphase References cells showing a polysomy of chromosome 6 Bergethon K, et al. (2012) J Clin Oncol 30: 863-70. Bos M, et al. (2013) Lung Cancer 81: 142-3. as indicated by multiple orange (CEN 6) Jin Y, et al. (2015) Virchows Arch 466: 45-52. and green (ROS1) signals in each nucleus. Mazières J, et al. (2015) J Clin Oncol 33: 992-9. Takeuchi K, et al. (2012) Nat Med 18: 378-81.

Prod. No. Product Label Tests* (Volume) Z-2162-200 ZytoLight SPEC ROS1/CEN 6 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

69 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 69 FE074-1-20 ZytoLight ® SPEC MYB Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC MYB Dual Color The SPEC MYB Dual Color Break Apart In an interphase nucleus lacking a transloca- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled tion involving the 6q23.2-q23.3 band, two translocations involving the chromosomal probes hybridizing to the 6q23.2-q23.3 orange/green fusion signals are expected region 6q23.3 harboring the MYB (MYB band. The orange fluorochrome direct representing two normal (non-rearranged) proto-oncogene, transcription factor, labeled probe hybridizes distal and the 6q23.2-q23.3 loci. A signal pattern consist- a.k.a. c-myb) gene. green fluorochrome direct labeled probe ing of one orange/green fusion signal, one The MYB gene is expressed predominantly hybridizes proximal to the MYB break- orange signal, and a separate green signal in immature progenitor cells of all hemato- point cluster region. indicates one normal 6q23.2-q23.3 locus poietic lineages and is highly expressed in and one 6q23.2-q23.3 locus affected by a most leukemias and in some solid tumors. translocation. Translocations affecting MYB have been detected in T-cell acute lymphoblastic leukemia (T-ALL) and adenoid cystic carcinoma (ACC). Recent studies have identified a subgroup of T-ALL with reciprocal translocation t(6;7) MYB (q23.3;q34) that juxtaposes MYB and TCRB (T-cell receptor beta locus) leading Ideogram of chromosome 6 to the activation of MYB expression. Since indicating the hybridization locations. the translocation breakpoints in 6q23 map SHGC-141085 RH69238 to two clusters located 5 kb and more RH112660 RH106508 5’ 3’ 3’ 5’ than 50 kb telomeric of MYB, no true MYB SPEC MYB Dual Color Break Apart Probe hybridized to normal interphase cells as indicated by two fusion gene is generated. It is assumed MYB AHI1 orange/green fusion signals per nucleus. that the abnormal MYB expression could ~645 kb ~660 kb confer oncogenic properties and that Cen 6q23.2-q23.3 Tel MYB might represent a potential target for SPEC MYB Probe map (not to scale). therapeutic intervention in T-ALL. In ACC a recurrent translocation t(6;9) (q22-23;p23-24) is found in about one third of karyotypically abnormal cases. The translocation results in the fusion of the two transcription factor genes MYB and NFIB (nuclear factor I/B) which leads to enhanced expression of the MYB-NFIB fusion protein. The detection of MYB rear- Adenoid cystic carcinoma tissue section rangements using FISH might represent a with translocation affecting the 6q23.3 locus References as indicated by one orange/green fusion powerful adjunctive diagnostic tool useful Clappier E, et al. (2007) Blood 110: 1251-61. Persson M, et al. (2009) Proc Natl Acad Sci U S A 106: 18740-4. (non-rearranged) signal, one orange signal, in the differential diagnosis of ACC. Stenman G, et al. (2010) Cell Cycle 9: 2986-95. and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2143-50 ZytoLight SPEC MYB Dual Color Break Apart Probe / 5 (50 μl) Z-2143-200 ZytoLight SPEC MYB Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

70 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 70 FE067-1-20 ZytoLight ® SPEC MYB/CEN 6 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MYB/CEN 6 Dual The SPEC MYB/CEN 6 Dual Color Probe In a normal interphase nucleus, two green Color Probe is designed to detect dele- is a mixture of a green fluorochrome and two orange signals are expected. tions affecting the chromosomal region direct labeled CEN 6 probe specific for In a cell with deletion affecting the 6q23.3 6q23.3 harboring the MYB gene. the alpha satellite centromeric region of locus, one or no copy of the orange signal The MYB (MYB proto-oncogene, transcrip- chromosome 6 (D6Z1) and an orange will be observed. tion factor, a.k.a c-myb) gene encodes for fluorochrome direct labeled SPEC MYB a transcription factor which is primarily probe specific for the chromosomal region expressed in premature lymphoid and 6q23.2-23.3 harboring the MYB gene. myeloid T-cells. Aberrations of 6q are the most commonly found chromosomal changes for different types of lymphoid neoplasms. Several major deletion regions CEN 6 (D6Z1) have been detected on the long arm of chromosome 6, one of them is 6q23. 3-10% of chronic lymphocytic leukemia (CLL) cases have been shown to harbor MYB structural aberrations in the chromosomal region 6q. Deletions of MYB often occur Ideogram of chromosome 6 SPEC MYB/CEN 6 Dual Color Probe hybridized to as secondary changes indicating disease indicating the hybridization locations. normal interphase cells as indicated by two orange progression. CLL patients presenting a and two green signals in each nucleus and to metaphase chromosomes of a normal cell. 6q23 deletion seem to exhibit a more D6S1976 RH33702 5’ 3’ favorable prognosis than patients with 11q23.3 and 17p13 deletions. However, MYB the prognostic relevance of 6q deletions in ~575 kb CLL is still controversially discussed. Cen 6q23.2-q23.3 Tel Since conventional cytogenetic methods SPEC MYB Probe map (not to scale). often miss alterations in CLL, investigation by molecular cytogenetic methods such as Fluorescence in situ Hybridization (FISH) may be of diagnostic and prognostic relevance.

References Döhner H, et al. (1999) J Mol Med 77: 266-81. Johansson B, et al. (1993) Genes Chromosomes Cancer 8: 205-18. Stilgenbauer S, et al. (1999) Leukemia 13: 1331-4. Urbankova H, et al. (2014) Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub 158: 56-64. Blood smear with deletion of the MYB gene as Wang DM, et al. (2011) Leuk Lymphoma 52: 230-7. indicated by one orange signal in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2281-50 ZytoLight SPEC MYB/CEN 6 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

71 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 71 FE151-1-20 ZytoLight ® SPEC ESR1/CEN 6 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ESR1/CEN 6 Dual The SPEC ESR1/CEN 6 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of ESR1 gene amplification frequently direct labeled CEN 6 probe specific for In a cell with amplification of the ESR1 observed in breast cancer. the alpha satellite centromeric region of gene locus, multiple copies of the green The ESR1 (estrogen receptor 1) gene is lo- chromosome 6 (D6Z1) and a green fluo- signal or green signal clusters will be cated in the chromosomal region 6q25.1 rochrome direct labeled SPEC ESR1 probe observed. and encodes estrogen receptor alpha (ER). hybridizing to the ESR1 locus at 6q25.1. ER expression is one of the most important known factors in the development of breast cancer, and assessing its status by immunohistochemistry is important for de- termining the use of anti-estrogen receptor therapies. ESR1 gene amplification has been found frequently in ER-positive breast tumors. Additionally, it has been shown very recently for breast cancer patients receiving Ideogram of chromosome 6 adjuvant tamoxifen monotherapy that indicating the hybridization locations. survival is significantly longer in cases of SPEC ESR1/CEN 6 Dual Color Probe hybridized to RH40922 RH72992 ESR1 gene amplification as determined by 5’ 3’ normal interphase cells as indicated by two orange and two green signals in each nucleus. FISH compared to immunohistochemically ESR1 ER-positive cases without gene amplifi- ~395 kb cation. Additionally, it has been shown Cen 6q25.1 Tel that response to tamoxifen is dependent SPEC ESR1 Probe map (not to scale). on the absolute ESR1 copy number. Thus, determination of ESR1 amplification may identify a subgroup of breast cancer patients particularly likely to respond to anti-estrogen therapy.

References Burkhardt L, et al. (2010) Breast Cancer Res Treat 123: 757-65. Holst F, et al. (2007) Nature Genet 39: 655-60. Lacroix M (2006) Endocr Relat Cancer 13: 1033-67. Moelans CB, et al. (2010) Anal Cell Pathol 33: 13-8. Nembrot M, et al. (1990) Biochem Biophys Res Comm 166: 601-7. Nessling M, et al. (2005) Cancer Res 65: 439-47. Pentheroudakis G, et al. (2013) PLoS One 8: e70634. ESR1 gene amplification as indicated by mutiple green Rahman MT, et al. (2013) Anticancer Res 33: 3775-81. ESR1 specific signals in each nucleus. Sassen A, et al. (2009) Breast Cancer Res 11: R50. Tomita S, et al. (2009) Cancer Sci 100: 1012-7.

Prod. No. Product Label Tests* (Volume) Z-2069-50 ZytoLight SPEC ESR1/CEN 6 Dual Color Probe / 5 (50 μl) Z-2069-200 ZytoLight SPEC ESR1/CEN 6 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

72 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 72 FE014-1-20 ZytoLight ® SPEC JAZF1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC JAZF1 Dual Color The SPEC JAZF1 Dual Color Break Apart In an interphase nucleus lacking a Break Apart Probe is designed to detect Probe is a mixture of two direct labeled translocation involving the 7p15.1-p15.2 translocations involving the chromosom- probes hybridizing to the 7p15.1-p15.2 band, two orange/green fusion signals al region 7p15.1-p15.2 harboring the band. The orange fluorochrome direct are expected representing two normal JAZF1 (JAZF zinc finger 1) gene. labeled probe hybridizes proximal, the (non-rearranged) 7p15.1-p15.2 loci. A Translocations involving the region green fluorochrome direct labeled probe signal pattern consisting of one orange/ 7p15.1-p15.2 are frequently found in hybridizes distal to the JAZF1 breakpoint green fusion signal, one orange signal, endometrial stromal sarcoma (ESS). The region. and a separate green signal indicates most common cytogenetic abnormality one normal 7p15.1-p15.2 locus and one 7p15.1-p15.2 locus affected by a detected in 33-80% of ESS is t(7;17) JAZF1 (p15.1-p15.2;q11.2) which results in the translocation. fusion of the JAZF1 gene at 7p15.1-p15.2 to the JJAZ1 (Joined to JAZF1; a.k.a. SUZ12) gene at 17q11.2. Both genes involved contain zinc finger domains characteristic for DNA binding proteins. It was shown that the fusion protein JAZF1-JJAZ1 Ideogram of chromosome 7 can promote cell proliferation when the indicating the hybridization locations. wild-type JJAZ1 is silenced as it is in ESS harboring the t(7;17). RH65631 D7S547 ECD04964 W1204 In 25-30% of ESS the JAZF1 gene is 3’ 5’ disrupted by another translocation t(6;7) JAZF1 where the first zinc finger domain of ~700 kb ~600 kb JAZF1 is fused to both zinc finger domains Tel 7p15.1-p15.2 Cen SPEC JAZF1 Dual Color Break Apart Probe of the PHF1 (PHD finger protein 1) gene hybridized to normal interphase cells as indicated SPEC JAZF1 Probe map (not to scale). by two orange/green fusion signals per nucleus. at 6p21.32. As a result the entire coding region of PHF1 is regulated by the JAZF1 promoter. Since the diagnosis of ESS is often difficult in cases showing diverse histological differentiation or in undifferentiated endometrial sarcoma (UES), the detection of the JAZF1 translocations can serve as a diagnostic tool to confirm the diagnosis of ESS.

References Hrzenjak A, et al. (2005) J Mol Diagn 7: 388-95. Koontz JI, et al. (2001) Proc Natl Acad Sci U S A 98: 6348-53. Li H, et al. (2007) Proc Natl Acad Sci U S A 104: 20001-6. Micci F, et al. (2003) Cancer Genet Cytogenet 144: 119-24. Endometrial stromal sarcoma with translocation affecting JAZF1 at 7p15.1-p15.2 as well as monosomy of chromosome 7 as indicated by one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2132-50 ZytoLight SPEC JAZF1 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

73 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 73 FE054-1-20 ZytoLight ® SPEC EGFR/CEN 7 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC EGFR/CEN 7 Dual The SPEC EGFR/CEN 7 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of EGFR gene amplification frequently direct labeled CEN 7 probe specific for In a cell with amplification of the EGFR observed in solid neoplasms including the alpha satellite centromeric region of gene locus, multiple copies of the green non-small cell lung cancer (NSCLC) and chromosome 7 (D7Z1) and a green fluoro- signal or green signal clusters will be glioblastoma. chrome direct labeled SPEC EGFR probe observed. The EGFR gene (a.k.a. ERBB1 and HER1) specific for the EGFR gene at 7p11.2 is located in the chromosomal region 7p11.2 and encodes a transmembrane glycoprotein acting as a cellular growth factor receptor. The protein belongs to the EGFR (epidermal growth factor receptor) subgroup of the RTK (receptor tyrosine kinase) superfamily also including ERBB2 (HER2), ERBB3 (HER3), and ERBB4 (HER4). Ideogram of chromosome 7 Overexpression of EGFR has been shown indicating the hybridization locations. in a number of tumor entities and is asso- SWSS2426 RH74846 ciated with poor prognosis. EGFR copy 5’ 3’ SPEC EGFR/CEN 7 Dual Color Probe hybridized to number identified by FISH is thought to be normal interphase cells as indicated by two orange a molecular predictor in neoplasms. EGFR and two green signals in each nucleus. ~635 kb References Ach T, et al. (2013) Virchows Arch 462: 65-72. Tel 7p11.2 Cen Balla P, et al. (2011) Histopathology 59: 376-89. Bernardes VM, et al. (2013) BMJ Open 3: e002077. SPEC EGFR Probe map (not to scale). Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Cappuzzo F, et al. (2005) J Natl Cancer Inst 97: 643-55. Ettl T, et al. (2012) Br J Cancer 106: 719-26. Ettl T, et al. (2014) Head Neck 36: 517-23. Gogas H, et al. (2010) Clin Breast Cancer 10: 230-7. Gonçalves A, et al. (2008) BMC Cancer 8: 169ff. Kondo I & Shimizu N (1983) Cytogenet Cell Genet 35: 9-14. Lauand C, et al. (2013) Cancer Cell Int 13: 38. Libermann TA, et al. (1985) J Cell Sci Suppl 3: 161-72. Merlino GT, et al. (1985) J Clin Invest 75: 1077-9. Murray S, et al. (2012) J Exp Clin Cancer Res 31: 77. Projetti F, et al. (2013) Human Pathology 44: 2116-25. Sassen A, et al. (2008) Breast Cancer Res 10: R2. Sassen A, et al. (2009) Breast Cancer Res 11: R50. Tovey SM, et al. (2004) Breast Cancer Res 6: 246-51.

Cancer cells with multiple copies of chromosome 7 and extra EGFR signals (green) in sputum sample from an NSCLC patient.

Prod. No. Product Label Tests* (Volume) Z-2033-50 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe / 5 (50 μl) Z-2033-200 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

74 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 74 FE010-1-20 ZytoLight ® SPEC CUX1/EZH2/CEN 7 Triple Color Probe

Background Probe Description Results The ZytoLight ® SPEC CUX1/EZH2/CEN 7 The SPEC CUX1/EZH2/CEN 7 Triple In a normal interphase nucleus, two or- Triple Color Probe is designed to detect Color Probe is a mixture of a green ange, two green, and two blue signals are losses of an entire chromosome 7 (mono- fluorochrome direct labeled SPEC CUX1 expected. In a cell with deletions affecting somy 7) and deletions of the long arm of probe hybridizing in the CDR at 7q22.1, the 7q22.1 and/or 7q36.1 locus, one chromosome 7 (del(7q)). an orange fluorochrome direct labeled or no copy of the green and/or orange In myeloid disorders, monosomy 7 or SPEC EZH2 probe hybridizing in the signal will be observed. Monosomy 7 will del(7q) are among the most common re- CDR at 7q36.1, and a blue fluorochrome result in a loss of a green, orange, and current chromosome abnormalities. These direct labeled CEN 7 probe specific for blue signal. aberrations occur in 8% of de novo acute the alpha satellite centromeric region of myeloid leukemia (AML), in 5-10% of de chromosome 7 (D7Z1). novo patients with myelodysplastic syndrome (MDS), and in approximately 50% of therapy-related myeloid neoplasms. My- eloid malignancies with monosomy 7 or CEN 7 (D7Z1) del(7q) respond poorly to chemotherapy and are associated with an unfavorable CUX1 prognosis. Several commonly deleted regions (CDRs) located on 7q have been identified in EZH2 MDS and AML, including CDRs at 7q22, Ideogram of chromosome 7 indicating the hybridization locations. 7q32-33, and 7q35-36. Bone marrow smear with deletion of the CUX1 gene Loss of one or more yet unidentified SWSS3109 SWSS3919 as indicated by one green signal in each nucleus. 5’ 3’ tumor suppressor gene(s) is thought to contribute to leukemic growth in myeloid CUX1 malignancies with -7/del(7q). CUX1 is a ~665 kb transcription factor encoded in the CDR at Cen 7q22.1 Tel 7q22 that exerts tumor suppressor activity SPEC CUX1 Probe map (not to scale). by regulating proliferative genes. Loss D7S2419 D7S688 SWSS2078 of CUX1 may thus contribute to disease 5’ 3’3’ 5’ pathogenesis. CUL1 EZH2 The CDR at 7q35-36 encodes nine genes ~245 kb including CUL1 and EZH2 which are the Cen 7q36.1 Tel most promising candidates due to known SPEC EZH2 Probe map (not to scale). function in and association with cancer. References Bone marrow smear with deletion of the EZH2 gene De Weer A, et al. (2010) PLoS One 5: e8676. as indicated by one orange signal in each nucleus. Döhner K, et al. (1998) Blood 92: 4031-5. McNerney ME, et al. (2013) Blood 121: 975-83. Specimens kindly provided by Paediatric Oncology/Haematology, Pellagatti A & Boultwood J (2015) Eur J Haematol 95: 3-15. Charité-Universitätsmedizin Berlin, Germany.

Prod. No. Product Label Tests* (Volume) Z-2214-50 ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

75 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 75 FE133-1-20 ZytoLight ® SPEC MET/CEN 7 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MET/CEN 7 Dual The SPEC MET/CEN 7 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of MET gene amplifications found in a direct labeled CEN 7 probe specific for In a cell with amplification of the MET variety of human tumors. the alpha satellite centromeric region gene locus, multiple copies of the green The MET gene (a.k.a. c-Met) is located of chromosome 7 (D7Z1) and a green signal or green signal clusters will be in the chromosomal region 7q31.2 and fluorochrome direct labeled SPEC MET observed. encodes a transmembrane tyrosine kinase probe specific for the MET gene located at receptor for the hepatocyte growth factor 7q31.2. (HGF). HGF and MET play an important role in angiogenesis and tumor growth. Activation or upregulation of MET was found in a number of carcinomas including lung, breast, colorectal, prostate, and gastric carcinomas as well as in gliomas, melanomas and some sarcomas. MET overexpression is known as a negative prognostic indicator in patients with various carcinomas, multiple myeloma, or Ideogram of chromosome 7 indicating the hybridization locations. glioma. Therefore, several inhibitors of the SPEC MET/CEN 7 Dual Color Probe hybridized STSG597800 REN64875 to normal interphase cells as indicated by two HGF/MET signaling pathway are being 5’ 3’ orange and two green signals in each nucleus. studied and developed as potent therapies to inhibit angiogenesis and tumor growth. MET Recently, it was shown that MET ampli- ~795 kb Cen 7q31.2 Tel fication leads to resistance to gefitinib or erlotinib in lung cancer by driving SPEC MET Probe map (not to scale). ERBB3-dependent activation of the PI3K pathway.

References Ach T, et al. (2013) Virchows Arch 462: 65-72. Cooper CS, et al. (1984) Nature 311: 29-32. Engelman JA, et al. (2007) Science 316: 1039-43. Ettl T, et al. (2014) Head Neck 36: 517-23. Garcia S, et al. (2007) Int J Oncol 31: 49-58. Hara T, et al. (1998) Lab Invest 78: 1143-53. Lacroix L, et al. (2014) PLoS One 1: e84319. Lee D, et al. (2015) Cancer Res Treat 47: 120-5. Preusser M, et al. (2014) Histopathology 65: 684-92. Schildhaus HU, et al. (2015) Clin Cancer Res 21: 907-15. Lung cancer cells with polysomy of chromosome 7 as indicated by four orange (CEN 7) and four green (MET) signals in the nuclei.

Prod. No. Product Label Tests* (Volume) Z-2087-50 ZytoLight SPEC MET/CEN 7 Dual Color Probe / 5 (50 μl) Z-2087-200 ZytoLight SPEC MET/CEN 7 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

76 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 76 FE024-1-20 ZytoLight ® SPEC BRAF Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC BRAF Dual Color The SPEC BRAF Dual Color Break Apart In an interphase nucleus lacking a rear- Break Apart Probe is designed for the Probe is a mixture of two direct labeled rangement involving the 7q34 band, two detection of rearrangements involving the probes hybridizing to the 7q34 band. The orange/green fusion signals are expected chromosomal region 7q34 harboring the orange fluorochrome direct labeled probe representing two normal (non-rearranged) BRAF (B-Raf proto-oncogene, serine/threo- hybridizes proximal, and the green fluo- 7q34 loci. A signal pattern consisting of nine kinase, a.k.a. BRAF1, NS7) gene. rochrome direct labeled probe hybridizes one orange/green fusion signal, one The BRAF gene encodes a protein-serine/ distal to the BRAF gene breakpoint region. orange signal, and a separate green sig- threonine kinase that participates in the nal indicates one normal 7q34 locus and MAPK cascade, which regulates a large one 7q34 locus affected by a transloca- variety of cell processes. tion or inversion. Isolated orange signals Various BRAF translocations were ob- are the result of deletions distal to the served in melanocytic nevi, pilocytic BRAF breakpoint region. astrocytomas, malignant melanoma, prostate and gastric cancer. The AKAP9-BRAF fusion resulting from paracentric inversion BRAF of chromosome 7q was found in radia- Ideogram of chromosome 7 tion-induced papillary thyroid carcinomas. indicating the hybridization locations. The fusion proteins contain the protein kinase domain but lack the autoinhibitory RH112764 D7S2227 SHGC-172032 SWSS3708 N-terminal portion of BRAF resulting in 3’ 5’ constitutive kinase activity. BRAF In addition, in pilocytic astrocytoma the ~495 kb ~700 kb FAM131B-BRAF fusion has been described Cen 7q34 Tel

resulting from interstitial deletion which SPEC BRAF Probe map (not to scale). removes the BRAF N-terminal inhibitory SPEC BRAF Dual Color Break Apart Probe domain. Moreover, pancreatic acinar cell hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. carcinoma - a rare subtype of pancreatic cancer with poor prognosis - shows a recurrent SND1-BRAF rearrangement. SND1-BRAF-transformed cells were shown to be sensitive to treatment with a MEK inhibitor. Hence, the detection of BRAF rearrangements by Fluorescence in situ Hybridiza- tion may represent a novel therapeutic target in various diseases. References Chmielecki J, et al. (2014) Cancer Discov 4: 1398-405. Ciampi R, et al. (2005) J Clin Invest 115: 94-101. Cin H, et al. (2011) Acta Neuropathol 121: 763-74. Dessars B, et al. (2007) J Invest Dermatol 127: 1468-70. Dougherty MJ, et al. (2010) Neuro Oncol 12: 621-30. Hutchinson KE, et al. (2013) Clin Cancer Res 19: 6696-702. Jones DT, et al. (2013) Nat Genet 45: 927-32. Miller VA, et al. (2014) J Clin Oncol 32 Suppl: Abstr. 11029. Palanisamy N, et al. (2010) Nat Med 16: 793-8.

Prod. No. Product Label Tests* (Volume) Z-2189-200 ZytoLight SPEC BRAF Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

77 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 77 FE095-1-20 ZytoLight ® SPEC BRAF/CEN 7 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC BRAF/CEN 7 Dual The SPEC BRAF/CEN 7 Dual Color Probe In a normal interphase nucleus, two orange Color Probe is designed for the detection is a mixture of a green fluorochrome direct and two green signals are expected. In a of amplifications involving the chromo- labeled SPEC BRAF probe specific for cell with amplification of the BRAF gene somal region 7q34 harboring the BRAF the BRAF gene at 7q34 and an orange locus or polysomy of chromosome 7, mul- gene (B-Raf proto-oncogene, serine/ fluorochrome direct labeled CEN 7 probe tiple copies of the green signal or large threonine kinase). The BRAF gene encodes specific for the alpha satellite centromeric green signal clusters will be observed. a protein-serine/threonine kinase that region of chromosome 7 (D7Z1). participates in the MAPK cascade, which regulates a large variety of cell processes. Activating mutations in BRAF are found in many tumor types, including malignant CEN 7 (D7Z1) melanoma, thyroid, colorectal, and ovarian carcinomas, lung adenocarcinoma, as well as in some sarcomas and gliomas. These mutations lead to constitutive BRAF activation of BRAF thereby promoting tumorigenesis. Ideogram of chromosome 7 indicating the hybridization locations. Copy number gains of mutated and non-mutated BRAF have been identified D7S2913 RH40727 3’ 5’ Normal interphase cells, in malignant melanoma (MM), follicular BRAF (green), CEN 7 (orange). thyroid tumors, astrocytoma, colorectal, BRAF and prostate cancer due to amplification ~525 kb Cen 7q34 Tel of the gene or polysomy of chromosome 7. These amplifications lead to an overexpres- SPEC BRAF Probe map (not to scale). sion of BRAF and to constitutive activation of the MAPK signaling pathway. Follicular carcinomas with BRAF copy number gain were observed to be more often invasive. Colorectal carcinoma or melanoma patients with BRAF V600E mutation were found to acquire resistance to MEK and BRAF inhibitors through amplification of the mutated BRAF gene. NSCLC tissue section with amplification of the BRAF gene (green). Hence, detection of BRAF amplifications by Fluorescence in situ Hybridization may be of therapeutic relevance for these cancer patients. References Ciampi R, et al. (2005) Endocr Pathol 16: 99-105. Corcoran RB, et al. (2010) Sci Signal 3: ra84. Dougherty MJ, et al. (2010) Neuro Oncol 12: 621-30. Little AS, et al. (2011) Sci Signal 4: ra17. Pfister S, et al. (2008) J Clin Invest 118: 1739-49. Ren G, et al. (2012) Genes Chromosomes Cancer 51: 1014-23. Roskoski R Jr. (2010) Biochem Biophys Res Commun 399: 313-7. Spittle C, et al. (2007) J Mol Diagn 9: 464-71. Tanami H, et al. (2004) Oncogene 23: 8796-804. Villanueva J, et al. (2013) Cell Rep 4: 1090-9. Willmore-Payne C, et al. (2006) Hum Pathol 37: 520-7.

Prod. No. Product Label Tests* (Volume) Z-2191-200 ZytoLight SPEC BRAF/CEN 7 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

78 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 78 FE094-1-20 ZytoLight ® SPEC NRG1/CD74 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC NRG1/CD74 The SPEC NRG1/CD74 TriCheck™ Probe In an interphase nucleus lacking a TriCheck™ Probe is designed to detect is a mixture of three direct labeled probes rearrangement involving the 8p12 and translocations involving the chromosomal hybridizing to the 8p12 and 5q32-q33.1 5q32-q33.1 bands, two orange/green region 8p12 harboring the NRG1 (neu- bands. The green fluorochrome direct fusion signals and two blue signals are regulin 1, a.k.a. HGL or GGF) gene and labeled probe hybridizes distal and the expected. A CD74-NRG1 fusion is indi- the chromosomal region 5q32 harboring orange fluorochrome direct labeled probe cated by one separate green signal, one the CD74 gene. hybridizes proximal to the NRG1 break- separate orange signal, and an addi- Using this probe it is possible to discrim- point region at 8p12. The blue fluoro- tional blue signal which colocalizes with inate between CD74-NRG1 fusions and chrome direct labeled probe hybridizes to the separated orange signal. An NRG1 translocations affecting NRG1, but not the CD74 gene region at 5q32-q33.1. rearrangement not involving CD74 is CD74, such as SLC3A2-NRG1 or VAMP2- indicated by separated orange and green NRG1 fusions. signals without an additional blue signal. NRG1 encodes a variety of growth factors NRG1 that are ligands for tyrosine kinase receptors of the ERBB family. Rearrangements of the NRG1 gene have been detected in various tumors, including breast cancer, lung cancer, and ovarian adenocarcinoma. NRG1 translocation-positive breast tumors Ideogram of chromosome 8 indicating the hybridization locations. show a more advanced pathological stage compared with translocation-negative tumors. NRG1 rearrangements in lung adenocarcinoma of never smokers were found

to result in, e.g., the fusion of CD74 to SPEC NRG1/CD74 TriCheck™ Probe hybridized to the EGF-like domain of NRG1and to be normal interphase cells as indicated by two orange/ green fusion signals and two blue signals per nucleus. associated with a shorter overall and dis- CD74 ease-free survival. Due to the involvement of NRG1 fusion proteins in oncogenesis and their association with ERBB receptors, Ideogram of chromosome 5 indicating the hybridization locations. NRG1 constitutes a good candidate for RH109547 RH111344 potential therapeutic applications, e.g., in D8S2226 D8S71 relation to lung tumor subtypes with so far 5’ 3’

no effective treatment. NRG1

Hence, detection of NRG1 rearrangements ~705 kb ~340 kb and CD74-NRG1 fusions by FISH may be Tel 8p12 Cen

of prognostic and therapeutic significance. SPEC NRG1 Probe map (not to scale). References Adélaïde J, et al. (2003) Genes Chromosomes Cancer 37: 333-45. BV166744 SHGC-53373 Fernandez-Cuesta L, et al. (2014) Cancer Discov 4: 415-22. 3’ 5’ Han JY, et al. (2015) Cancer Res 75: 614. Huang HE, et al. (2004) Cancer Res 64: 6840-4. CD74 Jung Y, et al. (2015) J Thorac Oncol 10: 1107-11. Pole JC, et al. (2006) Oncogene 25: 5693-706. ~1.0 Mb Cen 5q32-q33.1 Tel

SPEC CD74 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2194-200 ZytoLight SPEC NRG1/CD74 TriCheck Probe / / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

79 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 79 FE110-1-20 ZytoLight ® SPEC NRG1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NRG1 Dual Color The SPEC NRG1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 8p12 band, two translocations involving the chromosomal probes hybridizing to the 8p12 band. The orange/green fusion signals are expected region 8p12 harboring the NRG1 (neu- green fluorochrome direct labeled probe representing two normal (non-rearranged) regulin 1, a.k.a. HGL or GGF) gene. hybridizes distal and the orange fluoro- 8p12 loci. A signal pattern consisting NRG1 encodes a variety of growth factors chrome direct labeled probe hybridizes of one orange/green fusion signal, one that are ligands for tyrosine kinase recep- proximal to the NRG1 breakpoint region. orange signal, and a separate green tors of the ERBB family. Rearrangements signal in lung adenocarcinoma specimens of the NRG1 gene have been detected in indicates one normal 8p12 locus and one various tumors, including breast cancer, NRG1 8p12 locus affected by a translocation. lung cancer, and ovarian adenocarcinoma. NRG1 translocation-positive breast tumors show a more advanced pathological stage compared with translocation-negative tumors. NRG1 rearrangements in lung adenocar- Ideogram of chromosome 8 indicating the hybridization locations. cinomas of never smokers were found to result in the fusion of CD74 to the EGF-like RH109547 RH111344 D8S2226 D8S71 domain of NRG1. Several in vitro studies 5’ 3’ indicate that NRG1 fusion proteins lead to an increased activation of ERBB receptors NRG1 and are hence involved in tumor develop- ~705 kb ~340 kb Tel 8p12 Cen ment. SPEC NRG1 Dual Color Break Apart Probe Due to the involvement of NRG1 isoforms SPEC NRG1 Probe map (not to scale). hybridized on normal interphase cells as indicated by two orange/green fusion signals per nucleus. in oncogenesis and their association with ERBB receptors, NRG1 constitutes a good candidate for potential therapeutic applications, e.g., in relation to lung tumor subtypes with so far no effective treatment. Hence, detection of NRG1 rearrangements by Fluorescence in situ Hybridiza- tion represents a useful tool for studying carcinogenesis of various solid tumors and may be of prognostic and therapeutic significance. References Adélaïde J, et al. (2003) Genes Chromosomes Cancer 37: 333-45. Fernandez-Cuesta L, et al. (2014) Cancer Discov 4: 415-22. Huang HE, et al. (2004) Cancer Res 64: 6840-4. Lung cancer tissue section with rearrangement of the Pole JC, et al. (2006) Oncogene 25: 5693-706. NRG1 gene as indicated by extra orange signals.

Image kindly provided by Mc Leer A, Duruisseaux M, Wislez M, and colleagues, Grenoble and Paris, France.

Prod. No. Product Label Tests* (Volume) Z-2181-200 ZytoLight SPEC NRG1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

80 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 80 FE090-1-20 ZytoLight ® SPEC FGFR1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR1 Dual Color The SPEC FGFR1 Dual Color Break In an interphase nucleus of a normal Break Apart Probe is designed to detect Apart Probe is a mixture of two di- cell lacking a translocation involving the rearrangements involving the chromosom- rect labeled probes hybridizing to the 8p11.23-p11.22 band, two orange/ al region 8p11.23-p11.22 harboring the 8p11.23-p11.22 band. The orange green fusion signals are expected FGFR1 (fibroblast growth factor receptor fluorochrome direct labeled probe representing two normal (non-rear- 1, a.k.a. FLT2 and FLG) gene. hybridizes distal, the green fluorochrome ranged) 8p11.23-p11.22 loci. A signal Translocations affecting FGFR1 are direct labeled probe hybridizes proximal pattern consisting of one orange/green hallmarks of the 8p11 myeloproliferative to the FGFR1 gene breakpoint region at fusion signal, one orange signal, and syndrome (EMS), also known as stem 8p11.23-p11.22. a separate green signal indicates one cell leukemia/lymphoma syndrome, an normal 8p11.23-p11.22 locus and one aggressive stem cell myeloproliferative 8p11.23-p11.22 locus affected by a neoplasm that is associated with eosino- FGFR1 translocation. philia, poor prognosis, T-cell lymphoma, and frequent progression to acute myeloid leukemia. The most common translocation detected in EMS is t(8;13)(p11.2;q12.1) fusing FGFR1 to ZMYM2 (a.k.a. ZNF198). Ideogram of chromosome 8 indicating the hybridization locations. Several other rearrangements affecting the FGFR1 locus are also common in EMS, all RH110494 RH121775 RH44321 RH62114 of which result in fusion proteins compris- 3’ 5’ ing the tyrosine kinase domain of FGFR1 FGFR1 and a dimerization domain of a partner SPEC FGFR1 Dual Color Break Apart Probe ~605 kb ~600 kb hybridized to normal interphase cells as indicated protein. Due to dimerization these fusion Tel 8p11.22-p11.23 Cen by two orange/green fusion signals per nucleus. proteins show constitutive kinase activity. SPEC FGFR1 Probe map (not to scale). Currently, bone marrow or stem cell transplantation is the only curative treatment for patients with EMS. In vitro studies suggest that certain receptor tyrosine kinase inhibitors may provide a new therapeutic option. Detection of FGFR1 rearrangements using FISH may assist in the diagnosis of patients with this aggressive stem cell disorder. 8p11 myeloproliferative syndrome (EMS) tissue References section with translocation of the FGFR1 gene as Chase A, et al. (2007) Blood 110: 3729-34. indicated by one non-rearranged orange/green fusion Chase A, et al. (2013) Haematologica 98: 103-6. signal, one orange, and one separate green signal. Jackson CC, et al. (2010) Hum Pathol 41: 461-76. Sohal J, et al. (2001) Genes Chromosomes Cancer 32: 155-63.

Prod. No. Product Label Tests* (Volume) Z-2168-50 ZytoLight SPEC FGFR1 Dual Color Break Apart Probe / 5 (50 μl) Z-2168-200 ZytoLight SPEC FGFR1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

81 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 81 FE082-1-20 ZytoLight ® SPEC FGFR1/CEN 8 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR1/CEN 8 Dual The SPEC FGFR1/CEN 8 Dual Color In a normal interphase nucleus, two or- Color Probe is designed for the detection Probe is a mixture of an orange fluo- ange and two green signals are expected. of FGFR1 gene amplification frequently rochrome direct labeled CEN 8 probe In a cell with amplification of the FGFR1 observed in malignant tumors e.g. breast specific for the alpha satellite centromeric gene locus, multiple copies of the green and prostate cancer and oral squamous region of chromosome 8 (D8Z2) and a signal or green signal clusters will be cell carcinoma (OSCC). green fluorochrome direct labeled SPEC observed. The FGFR1 (fibroblast growth factor recep- FGFR1 probe specific for the FGFR1 gene tor 1) gene is located in the chromosomal at 8p11.23-p11.22. region 8p11.23-p11.22 and encodes a transmembrane receptor tyrosine kinase. Amplification of the FGFR1 gene, ob- FGFR1 CEN 8 (D8Z2) served in approximately 10% of all breast cancer samples, has revealed to be an independent prognostic factor for overall survival. FGFR1 is believed to emerge as a potential therapeutic target for lobular breast carcinomas. Ideogram of chromosome 8 indicating the hybridization locations. In prostate cancer, FGFR1 gene amplification seems to be an important step during RH32055 D8S414 SPEC FGFR1/CEN 8 Dual Color Probe hybridized to 3’ 5’ 3’ 5’ normal interphase cells as indicated by two orange the transmission to hormone resistance. and two green signals in each nucleus. In OSCC, FGFR1 gene amplification, WHSC1L1 FGFR1 observed in nearly 20% of all cases, is ~465 kb indicated to contribute to oral carcinogen- Tel 8p11.22-p11.23 Cen esis at an early stage of development. SPEC FGFR1 Probe map (not to scale). References Balko JM, et al. (2012) Mol Cancer Ther 11: 2301-5. Broom RJ, et al. (2012) Clin Genitourin Cancer 10: 202-6. Cihoric N, et al. (2014) Br J Cancer 110: 2914-22. Edwards J, et al. (2003) Clin Cancer Res 9: 5271-81. Elbauomy Elsheikh S, et al. (2007) Breast Cancer Res 9: R23. Fernanda Amary M, et al. (2014) Cancer Med 3: 980-7. Freier K, et al. (2007) Oral Oncology 43: 60-6. Kohler LH, et al. (2012) Virchows Arch 461: 49-57. Lacroix-Triki M, et al. (2010) J Pathol 222: 282-98. Lantuejoul S, et al. (2012) Oncologie 14: 530-7. Lee PL, et al. (1989) Science 245: 57-60. Lehnen NC, et al. (2013) Histopathology 63: 157-66. Pfeiffer M, et al. (2012) Nat Genet 44: 1104-10. Preusser M, et al. (2014) Lung Cancer 83: 83-9. Reis-Filho JS, et al. (2006) Clin Cancer Res 12: 6652-62. Schildhaus HU, et al. (2012) Mod Pathol 25: 1473-80. Lung carcinoma tissue section with interphase Schultheis AM, et al. (2014) Mod Pathol 27: 214-21. Seo AN, et al. (2014) Virchows Arch 465: 547-58. cells showing amplification of the FGFR1 Turner N, et al. (2010) Cancer Res 70: 2085-94. gene (green) and partly polysomy 8 (orange). Wetterskog D, et al. (2012) J Pathol 226: 84-96.

Prod. No. Product Label Tests* (Volume) Z-2072-50 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe / 5 (50 μl) Z-2072-200 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

82 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 82 FE020-1-20 ZytoLight ® SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC RUNX1/RUNX1T1 The SPEC RUNX1/RUNX1T1 Dual Color In a normal interphase nucleus, two or- Dual Color Dual Fusion Probe is designed Dual Fusion Probe is a mixture of a green ange and two green signals are expected. to detect the specific translocation involv- fluorochrome direct labeled RUNX1 probe A reciprocal translocation involving two ing the chromosomal region 21q22.12 covering the breakpoint region of the breakpoints splits the two signals and harboring the RUNX1 (a.k.a. AML1) gene RUNX1 gene and an orange fluorochrome generates a fusion signal on each of the and the chromosomal region 8q21.3 direct labeled RUNX1T1 probe covering chromosomes involved. The chromosomal harboring the RUNX1T1 (a.k.a. ETO, the breakpoint region of the RUNX1T1 regions which are not translocated are CBF2T1) gene. gene. This probe is approved to be used indicated by the single orange and green The balanced chromosomal translocation with a hybridization time of 2 hours on signal, respectively. t(8;21) is found in about 90% of acute cytological specimens. myeloid leukemia (AML) patients. AML is a heterogeneous clonal disorder of hematopoietic progenitor cells and one of the RUNX1 most common malignant myeloid disorders in adults. The runt related transcription factor 1 gene (RUNX1) and RUNX1 translocation partner 1 (RUNX1T1) gene are both involved in the transcriptional regulation of genes during normal hematopoiesis. RUNX1T1 The non-random translocation t(8;21) (q21.3;q22.1) is strongly associated SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Pro- be hybridized to normal interphase cells as indicated with the French-American-British (FAB) Ideograms of chromosomes 21 (above) and by two orange and two green signals in each nucleus. 8 (below) indicating the hybridization locations. phenotype M2 (AML-M2) and produces a chimeric gene consisting of the 5‘-region of D21S1705 RH104322 3’ 5’ the RUNX1 gene fused to the 3‘-region of the RUNX1T1 gene. The chimeric protein is RUNX1 thought to be associated with the nuclear ~1.3 Mb corepressor/histone deacetylase complex Cen 21q22.11-q22.12 Tel to block hematopoietic differentiation. SPEC RUNX1 Probe map (not to scale).

Fluorescence in situ Hybridization (FISH) RH122311 D8S2011 can provide important information for the 3’ 5’

management of patients with hematologic RUNX1T1

disorders. ~1.1 Mb References Cen 8q21.3-q22.1 Tel Dayyani F, et al. (2008) Blood 111: 4338-47. Bone marrow biopsy section with translocation Estey E & Döhner H (2006) Lancet 368: 1894-907. SPEC RUNX1T1 Probe map (not to scale). Gmidène A, et al. (2011) Med Oncol 28 Suppl 1: 509-12. affecting the RUNX1/RUNX1T1 locus as indicated by Licht D (2001) Oncogene 20: 5560-79. one separate orange signal, one separate green Vangala RK, et al. (2003) Blood 101: 270-7. signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2112-50 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe / 5 (50 μl) Z-2112-200 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe / 20 (200μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

83 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 83 FE047-1-20 ZytoLight ® SPEC MYC Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC MYC Dual Color The SPEC MYC Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 8q24.21 band two translocations involving the chromosomal probes hybridizing to the 8q24.21 band. orange/green fusion signals are expected region 8q24.21 harboring the MYC gene. The orange fluorochrome direct labeled representing two normal (non-rearranged) The MYC proto-oncogene (MYC proto-on- probe hybridizes proximal to the MYC 8q24.21 loci. A signal pattern consisting cogene, bHLH transcription factor, a.k.a. gene, the green fluorochrome direct labe- of one orange/green fusion signal, one CMYC) encodes a transcription factor led probe hybridizes distal to that gene. orange signal, and a separate green sig- essential for cell growth and proliferation The wide gap between the two probes nal indicates one normal 8q24.21 locus and is broadly implicated in tumorigene- of approximately 2 Mb allows for the and one 8q24.21 locus affected by an sis. Translocations involving the MYC gene detection of the t(2;8) translocation as well 8q24.21 translocation. are considered to be cytogenetic hall- as of t(8;14) and t(8;22). This probe is marks for Burkitt lymphoma but are also approved to be used with a hybridization found in other types of lymphomas. time of 2 hours on cytological specimens. The most frequent translocation involving the MYC gene region is t(8;14) (q24.21;q32.3) juxtaposing the MYC gene in 8q24.21 next to the IgH (immunoglobulin heavy chain) locus in 14q32.33. Further translocations affecting the MYC gene are t(8;22)(q24.21;q11.2) and t(2;8)(p11.2;q24.21), both of which involve one of the two immunoglobulin Ideogram of chromosome 8 SPEC MYC Dual Color Break Apart Probe indicating the hybridization locations. hybridized to normal interphase cells as indicated light chain loci. All three translocations by two orange/green fusion signals per nucleus. bring the MYC gene under the control of SHGC-130556 D8S1701 D8S1090 RH75025 a regulatory element from one of the im- 5’ 3’ munoglobulin loci resulting in constitutive overexpression of MYC. MYC ~475 kb ~560 kb References Boerma EG, et al. (2009) Leukemia 23: 225-34. Cen 8q24.21 Tel Dalla-Favera R, et al. (1982) Proc Natl Acad Sci U S A 79: 6497-501. Haralambieva E, et al. (2004) Genes Chromosomes Cancer 40: 10-8. SPEC MYC Probe map (not to scale). Veronese ML, et al. (1995) Blood 85: 2132-8.

Burkitt lymphoma tissue section with translocation affecting the 8q24.21 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2090-50 ZytoLight SPEC MYC Dual Color Break Apart Probe / 5 (50 μl) Z-2090-200 ZytoLight SPEC MYC Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

84 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 84 FE028-1-20 ZytoLight ® SPEC MYC/CEN 8 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MYC/CEN 8 Dual The SPEC MYC/CEN 8 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of MYC gene amplifications found in a direct labeled CEN 8 probe specific for In a cell with amplification of the MYC variety of human tumors. the alpha satellite centromeric region of gene locus, multiple copies of the green The MYC proto-oncogene (MYC proto-on- chromosome 8 (D8Z2) and a green fluo- signal or green signal clusters will be cogene, bHLH transcription factor, a.k.a. rochrome direct labeled SPEC MYC probe observed. CMYC) is located in the chromosomal specific for the MYC gene at 8q24.21. region 8q24.21 and encodes a transcription factor that can activate and repress transcription thereby regulating expression of numerous target genes that are essential for cell growth and proliferation. Deregulation of MYC is a common denom- inator in cancer. MYC amplification was found e.g. in breast, colon, kidney, lung, ovary, bladder, head and neck, and en- Ideogram of chromosome 8 indicating the hybridization locations. dometrial cancer. Several studies showed a correlation between gene amplification RH42764 SHGC-144185 5’3’ and disease progression or recurrence in SPEC MYC/CEN 8 Dual Color Probe hybridized to normal interphase cells as indicated by two breast cancer and other malignancies. MYC orange and two green signals in each nucleus. Malignant cutaneous angiosarcomas, for ~400 kb example, but not benign and atypical Cen 8q24.21 Tel vascular lesions occurring after radiother- SPEC MYC Probe map (not to scale). apy of breast cancer are characterized by amplification of the MYC gene. The presence of MYC amplification is thus of considerable diagnostic importance for the distinction of malignant from atypical postradiation vascular neoplasms of the skin. Since inactivation of MYC appears to References be effective in the treatment of neopla- Dalla-Favera R, et al. (1982) Proc Natl Acad Sci U S A 79: 6497-501. Fromont G, et al. (2013) Hum Pathol 44: 1617-23. sia MYC targeting therapies have been Mannuci S, et al. (2012) Adv Hematol 2012: 149780. Mentzel T, et al. (2012) Mod Pathol 25: 75-95. Breast cancer tissue section with developed some of which have entered Nesbit CE, et al. (1999) Oncogene 18: 3004-16. interphase cells showing partly polysomy 8 and Schraml P, et al. (1999) Clin Cancer Res 5: 1966-75. partly amplification of the MYC gene locus. clinical trials. Taub R, et al. (1982) Proc Natl Acad Sci U S A 79: 7837-41.

Prod. No. Product Label Tests* (Volume) Z-2092-50 ZytoLight SPEC MYC/CEN 8 Dual Color Probe / 5 (50 μl) Z-2092-200 ZytoLight SPEC MYC/CEN 8 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

85 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 85 FE025-1-20 ZytoLight ® SPEC MYC/IGH Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC MYC/IGH Dual Color The SPEC MYC/IGH Dual Color Dual In a normal interphase nucleus, two or- Dual Fusion Probe is designed to detect Fusion Probe is a mixture of an orange ange and two green signals are expected. the translocation t(8;14)(q24.21;q32.3) fluorochrome direct labeled MYC probe A reciprocal translocation involving two affecting the MYC gene in the chromo- spanning the known MYC breakpoints, breakpoints splits the two signals and somal region 8q24.21 and the IGH locus and a green fluorochrome direct labeled generates a fusion signal on each of the in 14q32.33. The MYC proto-oncogene IGH probe spanning the known break- chromosomes involved. The chromosomal (MYC proto-oncogene, bHLH transcription points of IGH. regions which are not translocated are factor, a.k.a. CMYC) encodes a transcrip- indicated by the single orange respective- tion factor essential for cell growth and ly green signal. proliferation and is broadly implicated in tumorigenesis. Translocations involving the MYC gene are considered to be cytogenetic hallmarks for Burkitt lymphoma (BL) but are also found in other types of lymphomas. The most frequent translocation involving the MYC gene region t(8;14) (q24.21;q32.3) can be found in approx. 80% of the BL cases and juxtaposes the MYC gene next to IGH (immunoglobulin heavy locus). Further translocations affecting the MYC gene Ideograms of chromosomes 8 (above) and SPEC MYC/IGH Dual Color Dual Fusion Probe are t(8;22)(q24.21;q11.2) and t(2;8) 14 (below) indicating the hybridization locations. hybridized to normal interphase cells as indicated by (p11.2;q24.21), both of which involve two orange and two green signals in each nucleus. SHGC-149687 8_3358F one of the two immunoglobulin light chain 5’ 3’ loci. All three translocations bring the MYC gene under the control of a regulato- MYC ry element from one of the immunoglobu- ~1.3 Mb Cen 8q24.21 Tel lin loci resulting in constitutive overexpression of MYC. SPEC MYC Probe map (not to scale).

The identification of MYC specific rear- BV448316 RH119884 3’ 5’ 3’ 5’ rangements is a critical part of the diag-

nostic work-up and management of JAG2 IGH

patients, identifying those who will benefit ~1.5 Mb from the intensive therapeutic regimens Cen 14q32.33 Tel used to treat BL. Fluorescence in situ SPEC IGH Probe map (not to scale). Hybridization (FISH) which allows the Burkitt lymphoma tissue section with t(8;14) as indi- correlation with immunochemistry can be cated by one separate orange signal, one separate green signal and two orange/green fusion signals References critical to patient management and is an May P, et al. (2010) Cancer Genet Cytogenet 198: 71-5. indicating the MYC/IGH translocation. Perkins AS & Friedberg JW (2008) Hematology Am Soc Hematol Educ Program: 341-8. approach commonly used. Veronese ML, et al. (1995) Blood 85: 2132-8.

Prod. No. Product Label Tests* (Volume) Z-2105-50 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe / 5 (50 μl) Z-2105-200 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

86 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 86 FE037-1-20 ZytoLight ® SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC CD274,PDCD1LG2/ The SPEC CD274, PDCD1LG2/CEN 9 In a normal interphase nucleus, two CEN 9 Dual Color Probe is designed for Dual Color Probe is a mixture of a green orange and two green signals are ex- the detection of CD274,PDCD1LG2 gene fluorochrome direct labeled SPEC CD274, pected. In a cell with amplification of the cluster amplifications observed in various PDCD1LG2 probe specific for the CD274 CD274,PDCD1LG2 gene cluster, multiple carcinomas, e.g. classical non-Hodgkin and PDCD1LG2 genes at 9p24.1 and an copies of the green signal or large green lymphoma and mediastinal large B-cell orange fluorochrome direct labeled CEN signal clusters will be observed. lymphoma. 9 probe specific for the classical satellite The CD274 (CD274 molecule, a.k.a. III region of chromosome 9 (D9Z3) at PDCD1LG1, PDL1) and PDCD1LG2 9q12. (programmed cell death 1 ligand 2, CD274, PDCD1LG2 a.k.a. PDL2, CD273) genes, which are separated by 42 kilobases, are located on chromosome 9p24.1. CEN 9 (D9Z3) The genes encode ligands for the PD-1 receptor of T-cells. CD274 is expressed by cancer cells of various tumor types, including melanoma, non-small cell lung cancer (NSCLC), breast cancer, and renal cell Ideogram of chromosome 9 indicating the hybridization locations. carcinomas. It is believed that interactions Primary mediastinal large B-cell lymphoma tissue section with amplification of the between the T-cell PD-1 receptor and its D9S791 RH110377 CD274,PDCD1LG2 gene region as indicated 5’ 3’ 5’ 3’ ligands CD274 or PDCD1LG2 expressed by green signal clusters in each nucleus.

by tumor cells prevent the immune system CD274 PDCD1LG2

from attacking the tumor cells. ~565 kb The blockade of the PD-1/CD274, Tel 9p24.1 Cen PDCD1LG2 pathway has yielded prom- SPEC CD274, PDCD1LG2 Probe map (not to scale). ising results in clinical trials conducted on tumors that express the PD-1 receptor. In early phase clinical trials compounds blocking PD-1 and CD274 have shown to be especially effective in advanced-stage NSCLC patients positive for CD274. Hence, targeting PD-1 or CD274, PDCD1LG2 represents a promising new treatment for this cancer entity. Consequently, the identification of CD274,PDCD1LG2 gene copy number References detected by Fluorescence in situ Hybridiza- Green MR, et al. (2012) Clin Cancer Res 18: 1611-8. Hao Y, et al. (2014) Clin Cancer Res 20: 2674-83. tion might be of prognostic and predictive Mamalis A, et al. (2014) Arch Dermatol Res 306: 511-9. Schalper KA, at al. (2014) Clin Cancer Res 20: 2773-82. relevance in diverse cancers. Velcheti V, et al. (2014) Lab Invest 94: 107-16.

Prod. No. Product Label Tests* (Volume) Z-2179-50 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe / 5 (50 μl) Z-2179-200 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

87 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 87 FE089-1-20 ZytoLight ® SPEC JAK2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC JAK2 Dual Color The SPEC JAK2 Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving rearrangements involving the chromo- probes hybridizing to the 9p24.2-p24.1 the 9p24.2-p24.1 bands, two orange/ somal region 9p24.1 harboring the JAK2 bands. The orange fluorochrome direct green fusion signals are expected rep- (Janus kinase 2, a.k.a. JTK10) gene. labeled probe hybridizes proximal to the resenting two normal (non-rearranged) The JAK (Janus kinase) family proteins, JAK2 gene at 9p24.1, the green fluoro- 9p24.2-p24.1 loci. A signal pattern con- which include JAK1, JAK2, JAK3, and chrome direct labeled probe hybridizes sisting of one orange/green fusion signal, TYK2, are cytoplasmic tyrosine kinases distal to the JAK2 gene at 9p24.2-p24.1. one orange signal, and a separate green that are essential in maintaining normal JAK2 signal indicates one normal 9p24.2-p24.1 hematopoiesis due to their involvement in locus and one 9p24.2-p24.1 locus affect- the JAK-STAT signaling pathway. Gain of ed by a translocation. function mutations, translocations, and amplifications involving JAK2, which lead to constitutive activation of the JAK2 kinase, have been described in various hematologic malignancies. Ideogram of chromosome 9 JAK2 translocations are associated with indicating the hybridization locations. a poor prognosis and have been found in myeloproliferative neoplasms (MPNs), RH62851 RH92501 D9S793 BV166680 including chronic myeloid leukemia (CML), 5’ 3’ as well as in other hematologic malignan- JAK2 cies, e.g., in acute lymphoblastic leukemia ~720 kb ~240 kb (ALL). Ph-like ALL patients with rearrange- Tel Cen 9p24.2-p24.1 SPEC JAK2 Break Apart Dual Color Probe hybridized ment of JAK2 were shown to have the SPEC JAK2 Probe map (not to scale). to normal interphase cells as indicated by two worst outcome compared to Ph-like ALL pa- orange/green fusion signals per nucleus and to metaphase chromosomes of a normal cell. tients carrying other genetic aberrations. Various different JAK2 fusion partners have been identified, with PMC1, BCR, and ETV6 being most common. In the revised 2016 WHO classification of myeloid neoplasms and acute leukemia, “myeloid/lymphoid neoplasms with PCM1-JAK2” are classified as a new provisional entity. Recent studies reported that after treatment with ruxolitinib, a JAK2 inhibitor, patients with JAK2-rearranged MPN achieved References hematologic remission. Arber DA, et al. (2016) Blood 127: 2391-405. He R, et al. (2016) Cancer Genet 209: 223-8. Hence, detection of JAK2 rearrangements Ho K, et al. (2010) J Assoc Genet Technol 36: 107-9. Lacronique V, et al. (1997) Science 278: 1309-12. by FISH may help in selecting patients Levavi H, et al. (2019) Acta Haematol 142: 105-12. Roberts KG, et al. (2014) N Engl J Med 371: 1005-15. eligible for therapy with JAK2 inhibitors. Rumi E, et al. (2015) Ann Hematol 94: 1927-8. Prod. No. Product Label Tests* (Volume) Z-2294-50 ZytoLight SPEC JAK2 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

88 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 88 FE163-1-20 ZytoLight ® SPEC CDKN2A/CEN 9 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC CDKN2A/CEN 9 The SPEC CDKN2A/CEN 9 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of an orange fluo- ange and two green signals are expected. detection of CDKN2A deletions frequently rochrome direct labeled CEN 9 probe In a cell with deletion of the CDKN2A observed in most tumor cell lines as well specific for the classical satellite III region gene locus, a reduced number of green as in primary human malignancies. of chromosome 9 (D9Z3) at 9q12 and a signals will be observed. Deletions The CDKN2A gene, often referred to as green fluorochrome direct labeled SPEC affecting only parts of the CDKN2A gene p16 or INK4a/ARF, is located in the chro- CDKN2A probe specific for the CDKN2A might result in a normal signal pattern with mosomal region 9p21.3. Using alternative gene at 9p21.3. green signals of reduced size. first exons and an alternative reading frame, the gene encodes for two distinct CDKN2A tumor suppressor proteins p16INK4a and p14ARF, both involved in cell cycle regulation. CDKN2A has been identified as a major susceptibility gene for melanoma. The tumor suppressor gene CDKN2A is inactivated by homozygous deletions with high frequency in a variety of human Ideogram of chromosome 9 indicating the hybridization locations. primary tumors e.g. bladder and renal cell D9S1749 RH98751 carcinoma, prostate and ovarian adeno- 5’ 3’ 3’ 5’ 3’5’ carcinoma, non-small cell lung cancer, MTAP sarcoma, glioma, mesothelioma, and SPEC CDKN2A/CEN 9 Dual Color Probe hybridized melanoma. Furthermore, deletion of the CDKN2A CDKN2B to normal interphase cells as indicated by two orange ~315 kb and two green signals in each nucleus. CDKN2A gene is found in up to 80% of Tel 9p21.3 Cen T-cell acute lymphoblastic leukemia cases SPEC CDKN2A Probe map (not to scale). and is associated with poor prognosis and relapse of the disease. References Cowan JM et al. (1988) J NatI Cancer Inst 80: 1159-64. Holley T, et al. (2012) PLoS One 7: e50586. Hussussian CJ, et al. (1994) Nat Genet 8: 15-21. Kamb A, et al. (1994) Science 264: 436-40. Nobori T, et al. (1994) Nature 368: 753-6. Quelle DE, et al. (1995) Cell 83: 993-1000. Rocco JW & Sidransky D (2001) Exp Cell Res 264: 42-55. Schoppmeyer K, et al. (1999) Neoplasia 1: 128-37. Schwarz S, et al. (2008) Cytometry A 73: 305-11. Sharpless NE (2005) Mutat Res 576: 22-38.

Glioblastoma tissue section with homozygous deletion of the CDKN2A gene as indicated by the loss of both green signals in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2063-50 ZytoLight SPEC CDKN2A/CEN 9 Dual Color Probe / 5 (50 μl) Z-2063-200 ZytoLight SPEC CDKN2A/CEN 9 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

89 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 89 FE012-1-20 ZytoLight ® SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe

Background Probe Description Results The ZytoLight ® SPEC CDKN2A/ The SPEC CDKN2A/CEN 3/7/17 In a normal interphase nucleus, two gold, CEN 3/7/17 Quadruple Color Probe is Quadruple Color Probe is a mixture of a two red, two green, and two blue signals designed for the simultaneous detection of gold fluorochrome direct labeled SPEC are expected. In a cell with deletion of the CDKN2A gene status and enumeration of CDKN2A probe specific for the CDKN2A CDKN2A gene locus, a reduced number chromosomes 3, 7, and 17 in tumor cells. gene at 9p21.3, a red fluorochrome direct of gold signals will be observed. In cells The tumor suppressor gene CDKN2A labeled CEN 3 probe specific for the with aneusomy of chromosomes 3, 7, or (a.k.a. p16 or p16INK4a) is located in alpha satellite centromeric region of chro- 17 more or less signals of the respective the chromosomal region 9p21.3 and mosome 3 (D3Z1), a green fluorochrome color will be visible. is inactivated by homozygous deletions direct labeled CEN 7 probe specific for with high frequency in a variety of human the alpha satellite centromeric region of primary tumors e.g. renal cell carcinoma, chromosome 7 (D7Z1), and a blue fluo- prostate and ovarian adenocarcinoma, rochrome direct labeled CEN 17 probe non-small cell lung cancer, sarcoma, glio- specific for the alpha satellite centromeric ma, mesothelioma, and melanoma. region of chromosome 17 (D17Z1). Additionally, non-random numerical D9S1749 RH98751 chromosome aberrations are frequently 5’ 3’ 3’ 5’ 3’5’ observed in a variety of solid tumors. MTAP

Hence, detection of these specific chromo- CDKN2A CDKN2B some aberrations in tumor cells can serve ~315 kb Tel 9p21.3 Cen as a valuable diagnostic aid in tumor SPEC CDKN2A Probe map (not to scale). Normal cytological specimen hybridized with SPEC classification and staging. For example, in CDKN2A/CEN 3/7/17 Quadruple Color Probe as papillary renal cell carcinoma trisomy 7 or indicated by two gold (CDKN2A), two red (CEN 3), two green (CEN 7), and two blue (CEN 17) signals. 17 is frequently found, while chromopho- bic RCC is characterized by widespread chromosomal losses.

References Barocas DA, et al. (2006) BJU Int 99: 290-5. Gallucci M, et al. (2005) J Clin Pathol 58: 367-71. Kamb A, et al. (1994) Science 264: 436-40. Sharpless NE (2005) Mutat Res 576: 22-38.

CDKN2A SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe hybridized to tumor cells showing a trisomy 9 as indicated by three CDKN2A signals (gold) in each nucleus.

Ideograms of chromosomes 9, 3, 7, and 17 indicating the hybridization locations.

Prod. No. Product Label Tests* (Volume) Z-2081-50 ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe / /•/• 5 (50 μl) Z-2081-200 ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe / /•/• 20 (200 μl)

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

90 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 90 FE029-1-20 ZytoLight ® SPEC NTRK2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NTRK2 Dual Color The SPEC NTRK2 Dual Color Break In an interphase nucleus of a normal Break Apart Probe is designed to detect Apart Probe is a mixture of two di- cell lacking a translocation involving the translocations involving the chromosomal rect labeled probes hybridizing to the 9q21.32-q21.33 band, two orange/ region 9q21.33 harboring the NTRK2 (neu- 9q21.32-q21.33 band. The green fluo- green fusion signals are expected rotrophic receptor tyrosine kinase 2, a.k.a. rochrome direct labeled probe hybridizes representing two normal (non-rear- TRKB) gene. proximal to the NTRK2 breakpoint region ranged) 9q21.32-q21.33 loci. A signal NTRK2 is a receptor tyrosine kinase (TK) at 9q21.32-q21.33, the orange fluoro- pattern consisting of one orange/green that upon brain-derived growth factor chrome direct labeled probe hybridizes fusion signal, one orange signal, and (BDGF) and neurotrophin 4/5 (NT-4/5) distal to the NTRK2 breakpoint region at a separate green signal indicates one binding phosphorylates itself and members 9q21.33. normal 9q21.32-q21.33 locus and one of the MAPK pathway. It plays a key role 9q21.32-q21.33 locus affected by a in central and peripheral nervous system translocation. development as well as in cell survival. Translocations affecting the NTRK2 gene have been reported in several cancer NTRK2 types, including glioblastomas, pilocytic astrocytomas, head and neck squamous cell carcinoma, and lung adenocarcinoma. NTRK2 rearrangements result in the fusion Ideogram of chromosome 9 indicating the hybridization locations. of the 3‘ end of the NTRK2 gene with the 5‘ end of different activating genes (AGBL4, RH122860 SHGC-146793 SHGC-112554 D9S919 PAN3, or AFAP1). All these fusion genes 5’ 3’ encode hybrid proteins comprising the TK domain of NTRK2 and the N-terminus of NTRK2 ~720 kb ~535 kb the partner proteins encoding dimerization SPEC NTRK2 Dual Color Break Apart Probe Cen 9q21.32-q21.33 Tel domains which results in ligand-independ- hybridized to normal interphase cells as indicated SPEC NTRK2 Probe map (not to scale). by two orange/green fusion signals per nucleus. ent TK activity. Currently, there are several ongoing clinical trials involving drugs with known inhibitory activity of NTRK-related kinases. Entrectinib and LOXO-101 represent two of these TRK inhibitors which have shown promising activity and good tolerability in patients with advanced solid tumors or NSCLC harboring NTRK1, 2, and 3 rearrangements. Hence, detection of NTRK2 translocations References by Fluorescence in situ Hybridization (FISH) Amatu A, et al. (2016) ESMO Open 1: e000023. Jones DTW, et al. (2013) Nat Genet 45: 927-32. may be of diagnostic and therapeutic Raez LE & Rolfo C (2016) Lung Cancer Manag 5: 1-4. Stransky N, et al. (2014) Nat Commun 5: 4846. relevance. Wu G, et al. (2014) Nat Genet 46: 444-50.

Prod. No. Product Label Tests* (Volume) Z-2205-50 ZytoLight SPEC NTRK2 Dual Color Break Apart Probe •/• 5 (50 μl) Z-2205-200 ZytoLight SPEC NTRK2 Dual Color Break Apart Probe •/• 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

91 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 91 FE117-1-20 ZytoLight ® SPEC NR4A3 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NR4A3 Dual Color The SPEC NR4A3 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 9q22.33-q31.1 translocations involving the chromosomal probes hybridizing to the 9q22.33-q31.1 band, two orange/green fusion signals region 9q22.33-q31.1 harboring the nu- band. The orange fluorochrome direct are expected representing two normal clear receptor subfamily 4, group A, mem- labeled probe hybridizes distal to the (non-rearranged) 9q22.33-q31.1 loci. A ber 3 (NR4A3; a.k.a. TEC, NOR1, CHN) NR4A3 gene and the green fluorochrome signal pattern consisting of one orange/ gene.Extraskeletal myxoid chondrosarco- direct labeled probe hybridizes proximal green fusion signal, one orange signal, ma (EMC) is a rare soft-tissue sarcoma of to that gene. and a separate green signal indicates chondroblastic origin that occurs primarily one normal 9q22.33-q31.1 locus and in adults. The tumor is characterized by one 9q22.33-q31.1 locus affected by a recurrent chromosomal translocations translocation. resulting in fusions of the NR4A3 gene to various N-terminal partners including EWSR1, RBP56, TCF12, and TFG. NR4A3 NR4A3 is a member of the steroid/thyroid receptor superfamily and acts as a transcriptional activator. The resulting chimeric Ideogram of chromosome 9 indicating the hybridization locations. proteins contain N-terminal parts of the various partners fused to the entire coding RH123392 BV448598 RH29424 RH69936 sequence of NR4A3. The most frequent 5’ 3’ reciprocal translocation is t(9;22)(q22.3- q31;q12.2) found in about 70% of EMC NR4A3 generating a EWSR1-NR4A3 fusion gene ~490 kb ~430 kb Cen 9q22.33-q31.1 Tel in which the 3’-terminal part of EWSR1 is SPEC NR3A3 Dual Color Break Apart Probe SPEC NR4A3 Probe map (not to scale). replaced by the entire NR4A3 gene. hybridized to normal interphase cells as indicated by two orange/green fusion signal per nucleus. EMC is histologically characterized by a mixture of cellular and myxoid stromal components, making it difficult to distinguish it from other benign or malignant mesenchymal tumors. Since chromosomal translocations of EWSR1 are found in several different neoplasias while NR4A3 rearrangements have been exclusively detected in EMC, assessment of NR4A3 rearrangements by Fluorescence in situ Hy- bridization might represent a helpful tool for the differential diagnosis of EMC.

References Benini S, et al. (2014) J Mol Diagn 16: 314-23. Extraskeletal myxoid chondrosarcoma tissue Labelle Y, et al. (1995) Hum Mol Genet 4: 2219-26. section with translocation affecting the 9q22.33-q31.1 Nogushi H, et al. (2010) Hum Pathol 41: 336-42. locus as indicated by one orange/green fusion Ohkura N, et al. (1994) Biochem Biophys Res Commun 205: 1959-65. Panagopoulos I, et al. (2002) Genes Chromosomes Cancer 35: 340-52. (non-rearranged) signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2145-50 ZytoLight SPEC NR4A3 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

92 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 92 FE068-1-20 ZytoLight ® SPEC ABL1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ABL1 Dual Color The SPEC ABL1 Dual Color Break In an interphase nucleus of a normal Break Apart Probe is designed to detect Apart Probe is a mixture of two di- cell lacking a translocation involving the rearrangements involving the chromosom- rect labeled probes hybridizing to the 9q34.11-q34.13 band, two orange/ al region 9q34.12 harboring the ABL1 9q34.11-q34.13 band. The green green fusion signals are expected rep- (ABL proto-oncogene 1, non-receptor fluorochrome direct labeled probe resenting two normal (non-rearranged) tyrosine kinase, a.k.a. ABL) gene. hybridizes proximal to the ABL1 gene 9q34.11-q34.13 loci. A signal pat- Chromosomal rearrangements involving at 9q34.11-q34.12, the orange flu- tern consisting of one orange/green ABL1 occur in various hematological ma- orochrome direct labeled probe hy- fusion signal, one orange signal, and lignancies leading to fusions of the ABL1 bridizes distal to the ABL1 gene at a separate green signal indicates one gene to different fusion partners. 9q34.12-q34.13. normal 9q34.11-q34.13 locus and one The translocation t(9;22)(q34.1;q11.2) re- 9q34.11-q34.13 locus affected by a trans- sults in BCR/ABL1 fusion and is observed location. in approx. 90% of patients with chronic Amplifications of the NUP214-ABL1 fusion myeloid leukemia (CML) and in approx. genes will result in multiple orange signals 25% of adults with acute lymphoblastic or orange signal clusters. leukemia (ALL). The rearrangements are cytogenetically characterized by the pres- ABL1 ence of the Philadelphia (Ph) chromosome. Other ABL1 fusion partners include, e.g., Ideogram of chromosome 9 indicating the hybridization locations. ETV6 and NUP214. The kinase domain of ABL1 is retained in all chimeric proteins. RH63026 RH35119 RH74683 D9S1991 The NUP214-ABL1 is the second most 5’ 3’ 5’ 3’ prevalent ABL1 fusion gene in malignant ABL1 NUP214 hemopathies, with a frequency of 5% in ~710 kb ~420 kb T-cell ALL. NUP214-ABL1 fusion genes are Cen 9q34.11-q34.13 Tel often found amplified on episomes. SPEC ABL1 Dual Color Break Apart Probe SPEC ABL1 Probe map (not to scale). hybridized to normal interphase cells as indicated Tyrosine kinase inhibitors, such as imatin- by two orange/green fusion signals per nucleus. ib, suppress the constitutive kinase activity of ABL1 fusion proteins. Therefore, these drugs may have potential in the treatment of patients with ABL1 fusions.

References De Braekeleer E, et al. (2011) Eur J Haematol 86: 361-71. De Klein A, et al. (1982) Nature 300: 765-7. Graux C, et al. (2009) Leukemia 23: 125-33. Lim TH, et al. (2005) Ann Acad Med Singapore 34: 533-8. Primo D, et al. (2003) Leukemia 17: 1124-9. Rieder H, et al. (1998) Leukemia 12: 1473-81. Sessarego M, et al. (2000) Haematologica 85: 35-9. Zheng X, et al. (2009) PLoS One 4: e7661.

Bone marrow biopsy section with translocation affecting the 9q34.11-q34.13 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2199-50 ZytoLight SPEC ABL1 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

93 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 93 FE100-1-20 ZytoLight ® SPEC BCR/ABL1 Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC BCR/ABL1 Dual The SPEC BCR/ABL1 Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed for Fusion Probe is a mixture of a green ange and two green signals are expected. the detection of the specific transloca- fluorochrome direct labeled BCR probe A reciprocal translocation involving two tions involving the chromosomal region spanning the minor and major breakpoint breakpoints splits the two signals and 9q34.12 harboring the ABL1 (a.k.a cluster of the BCR gene and an orange generates a fusion signal on each of the ABL) gene, and the chromosomal region fluorochrome direct labeled ABL1 probe chromosomes involved. The chromosomal 22q11.23, harboring the BCR (a.k.a. spanning the breakpoint region of the regions which are not translocated are BCR1) gene. Rearrangements involving ABL1 gene. This probe is approved to be indicated by the single orange respective- t(9;22)(q34.1;q11.2) are observed in used with a hybridization time of 2 hours ly green signal. approx. 90% of patients with chronic on cytological specimens. myeloid leukemia (CML) and in approx. 25% of adults with acute lymphoblastic leukemia (ALL). The rearrangements are cytogenetically characterized by the presence of the Philadelphia (Ph) chromosome. The translocation frequently results in the formation of a chimeric BCR/ABL1 fusion gene on the derivative chromosome 22. The gene product is a BCR/ABL1 protein with abnormal tyrosine kinase activity. In normal cells, ABL1 kinase activity is finely ABL1 SPEC BCR/ABL1 Dual Color Dual Fusion Probe regulated in response to growth factors hybridized to normal interphase cells as indicated by and other stimuli. The BCR/ABL1 fusion Ideograms of chromosomes 22 (above) and two orange and two green signals in each nucleus. 9 (below) indicating the hybridization locations. protein leads to constitutive activation of

down-stream signaling pathways, includ- D22S1114 D22S1259 ing Ras, Jak/Stat and PI-3 kinase. In rare 5’ 3’

cases the BCR/ABL1 fusion gene is locat- BCR

ed on chromosomal sites other than the Ph ~1.4 Mb chromosome. Cen 22q11.22-q11.23 Tel Fluorescence in situ Hybridization (FISH) SPEC BCR Probe map (not to scale). allows for the identification of rearrange- SHGC-147754 RH69984 ments that could otherwise not be detected 5’ 3’ by conventional karyotyping. ABL1 References Hehne S, et al. (2012) Pathol Res Pract 208: 510-7. ~880 kb Lim TH, et al. (2005) Ann Acad Med Singapore 34: 533-8. Cen Tel Primo D, et al. (2003) Leukemia 17: 1124-9. 9q34.11-q34.13 Bone marrow biopsy tissue section with translocation Rieder H, et al. (1998) Leukemia12: 1473-81. affecting the BCR/ABL1 loci as indicated by one SPEC ABL1 Probe map (not to scale). Sessargeo M, et al. (2000) Haematologica 85: 35-9. separate orange signal, one separate green Zheng X, et al. (2009) PLoS One 4: e7661. signal and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2111-50 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe / 5 (50 μl) Z-2111-200 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

94 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 94 FE036-1-20 ZytoLight ® SPEC NUP214 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NUP214 Dual Color The SPEC NUP214 Dual Color Break In an interphase nucleus lacking a rear- Break Apart Probe is designed for the Apart Probe is a mixture of two di- rangement involving the 9q34.12-q34.13 detection of translocations involving the rect labeled probes hybridizing to the band, two orange/green fusion signals chromosomal region 9q34.13 harboring 9q34.12-q34.13 band. The orange fluo- are expected representing two normal the NUP214 (nucleoporin 214, a.k.a. rochrome direct labeled probe hybridizes (non-rearranged) 9q34.12-q34.13 loci. A CAN, CAIN) gene. proximal and the green fluorochrome signal pattern consisting of one orange/ Rearrangements of the NUP214 gene direct labeled probe hybridizes distal to green fusion signal, one orange signal, have been implicated in the pathogenesis the NUP214 gene. and a separate green signal indicates of several types of hematologic malignan- one normal 9q34.12-q34.13 locus and cies, including T-cell acute lymphoblastic one 9q34.12-q34.13 locus affected by a leukemia (T-ALL), acute myeloid leukemia translocation. (AML), and also myelodysplastic syndrome Isolated green signals are the result of (MDS). Several fusion partners have been deletions proximal to the NUP214 break- identified for NUP214. The most common point region. are the DEK, SET, and the tyrosine kinase NUP214 encoding gene ABL1. The translocation t(6;9)(p22.3;q34.1) re- Ideogram of chromosome 9 indicating the hybridization locations. sults in a DEK-NUP214 fusion and defines a specific subcategory of AML according SHGC-130535 RH26603 ABL1_685 SHGC-154727 to the World Health Organization 2008 5’ 3’ classification. The SET-NUP214 fusion is associated with NUP214 T-ALL, less frequently with AML, and acute ~290 kb ~555 kb Cen 9q34.12-q34.13 Tel undifferentiated leukemia and can result SPEC NUP214 Probe map (not to scale). from either a translocation or a deletion. NUP214-ABL1 fusions are exclusively as-

sociated with T-ALL patients. These patients SPEC NUP214 Dual Color Break Apart Probe may be considered for a targeted therapy hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus and to with specific tyrosine kinase inhibitors. The metaphase chromosomes of a normal cell. fusion is often located on amplified episomes and is cytogenetically cryptic but can be detected by FISH. Malignancies with NUP214 rearrangements are associated with a poor prognosis indicating the usefulness of NUP214 also as a prognostic biomarker.

References Fahrenkrog B (2014) New J Sci 2014: 468306. Takeda A & Yaseen NR (2014) Semin Cancer Biol 27: 3-10. von Lindern M, et al. (1992) Baillieres Clin Haematol 5: 857-79. Zhou MH & Yang QM (2014) Oncol Lett 8: 959-62.

Prod. No. Product Label Tests* (Volume) Z-2265-50 ZytoLight SPEC NUP214 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

95 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 95 FE135-1-20 ZytoLight ® SPEC KIF5B Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC KIF5B Dual Color The SPEC KIF5B Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cation involving the 10p11.22 band, two translocations involving the chromosomal probes hybridizing to the 10p11.22 band. orange/green fusion signals are expected region 10p11.22 harboring the KIF5B The orange fluorochrome direct labeled representing two normal (non-rearranged) (kinesin family member 5B) gene. probe hybridizes distal, the green fluoro- 10p11.22 loci. A signal pattern consisting About 5% of all non-small cell lung cancer chrome direct labeled probe hybridizes of one orange/green fusion signal, one cases are positive for the ALK-EML4 fusion proximal to the KIF5B gene. orange signal, and a separate green sig- as a result of an inversion in chromosome nal indicates one normal 10p11.22 locus 2. However, not in all cases showing an and one 10p11.22 locus affected by a aberration of the ALK gene the EML4-ALK KIF5B translocation. fusion transcript could be detected. KIF5B was identified as a novel fusion partner for ALK in ALK-positive lung cancer. KIF5B is a ubiquitously expressed microtubule-based motor protein involved in orga- Ideogram of chromosome 10 nelle transport. The translocation t(2;10) indicating the hybridization locations. (p23;p11.2) results in the fusion of the first domains of KIF5B including the motor HUMUT564 RH51311 GDB:209150 D10S2354 domain and the coiled-coil domain with 3’ 5’ the tyrosine kinase domain of ALK. Over- KIF5B expression of the aberrant KIF5B/ALK ~465 kb ~890 kb fusion transcript can lead to enhanced cell Tel 10p11.22 Cen

proliferation, migration, and invasion. SPEC KIF5B Probe map (not to scale). SPEC KIF5B Dual Color Break Apart Probe hybridized A further aberration affecting the KIF5B to normal interphase cells as indicated by two orange/green fusion signals per nucleus. gene is inv(10)(p11.2q11.2). This inversion was detected in adenocarcinomas of the lung and results in the fusion of KIF5B with the ret proto-oncogene (RET). The fusion transcript again comprises the coiled-coil domain of KIF5B and the tyrosine kinase domain of RET. In accordance with the EML4-ALK fusion the development of specific agents targeting KIF5B-RET might provide a new therapeutic strategy for lung adenocarcinomas.

References Gautschi O, et al. (2013) J Thorac Oncol 8: e43-4. Ju YS, et al. (2012) Genome Res 22: 436-45. Kohno T, et al. (2012) Nat Med 18: 375-7. NSCLC tissue section with tetrasomy Takeuchi K, et al. (2009) Clin Cancer Res 15: 3143-9. of chromosome 10 in some cells and an Takeuchi K, et al. (2012) Nat Med 18: 378-81. unbalanced translocation affecting KIF5B as Wong DW, et al. (2011) Cancer 117: 2709-18. indicated by one or two extra green signals.

Prod. No. Product Label Tests* (Volume) Z-2131-50 ZytoLight SPEC KIF5B Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

96 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 96 FE055-1-20 ZytoLight ® SPEC RET Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC RET Dual Color Break The SPEC RET Dual Color Break Apart In an interphase nucleus lacking a translo- Apart Probe is designed to detect translo- Probe is a mixture of two direct labeled cation involving the 10q11.21 band, two cations involving the chromosomal region probes hybridizing to the 10q11.21 orange/green fusion signals are expected 10q11.21 harboring the RET (ret proto-on- band. The orange fluorochrome direct representing two normal (non-rearranged) cogene) gene. labeled probe hybridizes proximal to the 10q11.21 loci. A signal pattern consisting RET encodes a tyrosine kinase (TK) receptor. RET gene, the green fluorochrome direct of one orange/green fusion signal, one Translocations involving RET were first labeled probe hybridizes distal to that orange signal, and a separate green signal described in papillary thyroid carcinoma gene. indicates one normal 10q11.21 locus and (PTC) where somatic rearrangements result one 10q11.21 locus affected by a translo- in the fusion of its TK catalytic domain with cation or inversion. Isolated green signals an N-terminal dimerization domain encod- are the result of deletions proximal to the ed by various fusion partner genes. RET RET breakpoint region. More recently, recurrent inversions [inv (10)(p11.2q11.2)] fusing the coiled-coil domains of the kinesin family member 5B (KIF5B) gene to the RET kinase domain Ideogram of chromosome 10 have been detected in lung adenocarcino- indicating the hybridization locations. ma. The resulting KIF5B-RET fusion protein can form homodimers through the coiled- D10S469 SHGC-105605 D10S2320 BV208590 coil domains of KIF5B, causing an aberrant 5’ 3’ activation of the TK of RET, a mechanism RET known from KIF5B-ALK fusions which is also ~170 kb ~275 kb found in lung adenocarcinoma. Cen 10q11.21 Tel Since in vitro studies showed transforming SPEC RET Probe map (not to scale). SPEC RET Dual Color Break Apart Probe hybridized activity of KIF5B-RET which could be sup- to normal interphase cells as indicated by two orange/green fusion signals per nucleus. pressed by a TK inhibitor, it was assumed that the chimeric oncogene might be a promising molecular target for the treatment of lung cancer. The same holds true for the very recently discovered BCR-RET and FGFR1OP-RET fusion genes in chronic myelomonocytic leukemia (CMML) generated by two balanced translocations t(10;22)(q11.2;q11.2) and t(6;10)(q27;q11.2), respectively.

References Ballerini P, et al. (2012) Leukemia 26: 2384-9. Gautschi O, et al. (2013) J Thorac Oncol 8: e43-4. Ju YS, et al. (2012) Genome Res 22: 436-45. Human thyroid tumor cell line (TPC-1) with transloca- Kohno T, et al. (2012) Nat Med 18: 375-7. tion affecting the 10q11.21 locus as indicated by Lee SE, et al. (2015) Mod Pathol 28: 468-79. one orange/green fusion (non-rearranged) signal, Nikiforov YE (2002) Endocr Pathol 13: 3-16. one orange signal, and one separate green signal. Takahashi M, et al. (1985) Cell 42: 581-8. Takeuchi K, et al. (2012) Nat Med 18: 378-81.

Prod. No. Product Label Tests* (Volume) Z-2148-50 ZytoLight SPEC RET Dual Color Break Apart Probe •/• 5 (50 μl) Z-2148-200 ZytoLight SPEC RET Dual Color Break Apart Probe •/• 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

97 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 97 FE065-1-20 ZytoLight ® SPEC PTEN/CEN 10 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC PTEN/CEN 10 Dual The SPEC PTEN/CEN 10 Dual Color In a normal interphase nucleus, two or- Color Probe is designed for the detection Probe is a mixture of an orange fluoro- ange and two green signals are expected. of PTEN deletions frequently observed chrome direct labeled CEN 10 probe In a cell with deletions of the PTEN gene in many tumor types, including renal, specific for the alpha satellite centromeric locus, a reduced number of green signals melanoma, endometrial, breast, prostate, region of chromosome 10 (D10Z1) and a will be observed. Deletions affecting only lung, bladder, and thyroid cancer but also green fluorochrome direct labeled SPEC parts of the PTEN gene might result in in hematological neoplasms. PTEN probe specific for the chromosomal normal signal pattern with green signals The tumor suppressor gene PTEN (phos- region 10q23.2-q23.31 harboring the of reduced size. phatase and tensin homolog deleted on PTEN gene. chromosome ten), often referred to as MMAC1 (mutated in multiple advanced cancers 1), is located on 10q23.31 and encodes a 47 kDa dual-specificity phosphatase that has both lipid and protein phosphatase activity. Its inactivation results in constitutive activation of the PI3K/AKT pathway and in subsequent increase in Ideogram of chromosome 10 protein synthesis, cell cycle progression, indicating the hybridization locations. migration, and survival. RH17273 RH93626 Deletions affecting the long arm of chro- 5’ 3’ mosome 10 have been detected in 30 to SPEC PTEN/CEN 10 Dual Color Probe hybridized to PTEN 50% of early and advanced stage spo- normal interphase cells as indicated by two orange ~315 kb and two green signals in each nucleus. radic melanomas and about 40 to 70% Cen 10q23.2-q23.31 Tel of prostate cancers. In both tumor entities SPEC PTEN Probe map (not to scale). loss of PTEN has been associated with poor clinical outcome. Currently, several drugs targeting the PI3K/AKT pathway for the therapy of solid tumors have entered clinical trials. References Ach T, et al. (2013) Virchows Arch 462: 65-72. Dahia PLM, et al. (1999) Hum Mol Genet 8: 185-93. Ettl T, et al. (2012) Br J Cancer 106: 719-26. Ettl T, et al. (2014) Head Neck 36: 517-23. Healy E, et al. (1998) Oncogene 16: 2213-8. Li J, et al. (1997) Science 275: 1943-7. Swoboda A, et al. (2011) Genes Chromosomes Cancer 50: 680-8. Weng LP, et al. (2001) Hum Mol Genet 10: 599-604. Yoshimoto M, et al. (2006) Cancer Genet Cytogenet 169: 128-37. Yoshimoto M, et al. (2007) Br J Cancer 97: 678-85. Melanoma tissue section with chromosome 10 monosomy as indicated by one orange and one green signal in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2078-50 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe / 5 (50 μl) Z-2078-200 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

98 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 98 FE018-1-20 ZytoLight ® SPEC FGFR2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR2 Dual Color The SPEC FGFR2 Dual Color Break In an interphase nucleus of a normal Break Apart Probe is designed to detect Apart Probe is a mixture of two di- cell lacking a translocation involving the rearrangements involving the chromosom- rect labeled probes hybridizing to the 10q26.13 band, two orange/green al region 10q26.13 harboring the FGFR2 10q26.12-q26.13 band. The orange fluo- fusion signals are expected representing (fibroblast growth factor receptor ,2 a.k.a. rochrome direct labeled probe hybridizes two normal (non-rearranged) 10q26.13 BEK) gene. distal to the FGFR2 gene at 10q26.13, loci. A signal pattern consisting of one Translocations and inversions affecting the green fluorochrome direct labeled orange/green fusion signal, one orange FGFR2 have been detected in several solid probe hybridizes proximal to the FGFR2 signal, and a separate green signal tumors, including e.g. breast cancer, lung gene at 10q26.12-q26.13. indicates one normal 10q26.13 locus cancer, and the intrahepatic subtype of and one 10q26.13 locus affected by a cholangiocarcinoma. translocation. Several partner genes have been described to be fused to FGFR2 after rearrangement. The resulting fusion genes are predicted to encode chimeric proteins carrying the kinase domain of FGFR2. Most of the currently FGFR2 known FGFR2 fusion products are likely to Ideogram of chromosome 10 exhibit oligomerization capability resulting indicating the hybridization locations. in kinase activation. RH113319 SHGC-146065 In prostate cancer FGFR2 was found to be RH37911 SHGC-152845 fused to the promoter region of SLC45A3 3’ 5’

predicted to result in signal activation by FGFR2

overexpression of the FGFR2 protein. ~535 kb ~355 kb Recent studies indicate the involvement of Cen 10q26.12-q26.13 Tel SPEC FGFR2 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated FGFR2 fusion proteins in tumorigenesis. SPEC FGFR2 Probe map (not to scale). by two orange/green fusion signals per nucleus. Moreover, in vitro studies suggest that certain FGFR tyrosine kinase inhibitors may provide a new therapeutic option for patients showing FGFR2 rearrangement. Hence, detection of FGFR2 rearrangements using FISH may help to identify patients which might respond to FGFR2 kinase targeting therapies.

References Arai Y, et al. (2014) Hepatology 59: 1427-34. Seo JS, et al. (2012) Genome Res 22: 2109-19. Wu YM, et al. (2013) Cancer Discov 3: 636-47.

Cholangiocellular adenocarcinoma tissue section with translocation of the FGFR2 gene as indicated by one non-rearranged orange/green fusion signal, one orange, and one separate green signal.

Kindly provided by Prof. Dr. Büttner, Cologne, Germany.

Prod. No. Product Label Tests* (Volume) Z-2169-200 ZytoLight SPEC FGFR2 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

99 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 99 FE083-1-20 ZytoLight ® SPEC FGFR2/CEN 10 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC FGFR2/CEN 10 The SPEC FGFR2/CEN 10 Dual Color In a normal interphase nucleus, two Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- orange and two green signals are ex- detection of FGFR2 gene amplifications chrome direct labeled CEN 10 probe pected. Nuclei with amplification of the frequently observed in breast cancer as specific for the alpha satellite centromeric FGFR2 gene locus 10q26.12-q26.13, or well as in gastric cancer. region of chromosome 10 (D10Z1) and a aneuploidy of chromosome 10 will show The FGFR2 (fibroblast growth factor gene green fluorochrome direct labeled SPEC multiple copies of the green signal or 2, a.k.a. BEK) gene is located on chro- FGFR2 probe specific for the chromosomal large green signal clusters. mosome 10q26.13 and encodes splice region 10q26.12-q26.13 harboring the variants of the receptor tyrosine kinases FGFR2 gene. FGFR2b and FGFR2c. Amplification of the FGFR2 gene leads to overexpression of the FGFR2 protein and subsequently to signal activation. Addition- ally, during the amplification process the C-terminal deletion of FGFR2 can occur due to exclusion of the last exon from FGFR2 the FGFR2 amplicon. Both, overexpres- Ideogram of chromosome 10 sion and deletion of the last exon result indicating the hybridization locations. in FGFR2 signaling activation based on SHGC-112517 RH76635 constitutive phosphorylation of the FRS2 3’ 5’ SPEC FGFR2/CEN 10 Dual Color Probe adaptor molecule. hybridized to normal interphase cells as indicated by two orange and two green signals in each nucleus. The process of ligand independent FGFR2 FGFR2 signaling leads to a more severe malig- ~775 kb Cen 10q26.12-q26.13 Tel nant phenotype of these tumors. Moreo- SPEC FGFR2 Probe map (not to scale). ver, high FGFR2 expression is correlated with poor overall survival (OS) and poor disease-free survival (DFS) rates in breast cancer patients. Consequently, FGFR2 gene amplification detected by Fluores- cence in situ Hybridization might be used as a prognostic marker e.g. in breast cancer. References Azuma K, et al. (2011) Biochem Biophys Res Commun 407: 219-24. Chang J, et al. (2015) Oncotarget 6: 2009-22. Katoh M (2010) Expert Rev Anticancer Ther 10: 1375-9. Breast cancer tissue section with amplification Katoh Y & Katoh M (2009) Int J Mol Med 23: 307-11. of the FGFR2 gene as indicated by Moffa AB, et al. (2004) Mol Cancer Res 2: 643-52. Sun S, et al. (2012) J Surg Oncol 105: 773-9. green signal clusters in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2122-200 ZytoLight SPEC FGFR2/CEN 10 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

100 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 100 FE046-1-20 ZytoLight ® SPEC CARS Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CARS Dual Color The SPEC CARS Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 11p15.4-p15.5 translocations involving the chromosomal probes hybridizing to the 11p15.4-p15.5 band, two orange/green fusion signals region 11p15.4 harboring the CARS band. The orange fluorochrome direct are expected representing two normal (cysteinyl-tRNA-synthetase) gene detected labeled probe hybridizes proximal to the (non-rearranged) 11p15.4-p15.5 loci. A in inflammatory myofibroblastic tumors CARS gene and the green fluorochrome signal pattern consisting of one orange/ (IMT). direct labeled probe hybridizes distal to green fusion signal, one orange signal, IMT are neoplastic mesenchymal prolifera- that gene. and a separate green signal indicates tions that occur predominantly in children one normal 11p15.4-p15.5 locus and and young adults. Cytogentic studies of CARS one 11p15.4-p15.5 locus affected by a IMT show various complex karyotypic ab- translocation. normalities, frequently involving the short arm of chromosome 2 harboring the ALK gene locus in 2p23.1-p23.2. The ALK (ALK receptor tyrosine kinase, a.k.a. CD246) gene encodes a receptor tyrosine Ideogram of chromosome 11 kinase and was frequently identified as indicating the hybridization locations. a fusion partner of various hybrid genes predominantly in anaplastic large cell lym- RH120245 RH92244 SHGC-14772 D11S2431 phoma, and more recently, in non-small 3’ 5’ cell lung cancer. However, also in IMT CARS several different ALK fusion genes have ~410 kb ~270 kb been identified including CARS-ALK. Tel 11p15.4-p15.5 Cen

CARS encodes a class 1 aminoacyl-tRNA SPEC CARS Probe map (not to scale). SPEC CARS Dual Color Break Apart Probe hybridized synthetase and is ubiquitously expressed. to normal interphase cells as indicated by two orange/green fusion signals per nucleus. The translocation results in the fusion of the active promoter as well as the first domains of CARS to the receptor tyrosine kinase domain of ALK. Thus, CARS is predicted to mediate homodimerization of the chimeric product resulting in constitutive ALK kinase activation. The detection of translocations affecting CARS and ALK by Fluorescence in situ Hy- bridization might represent a valuable tool to identify a subpopulation of IMT likely to respond to ALK kinase targeting therapies.

References Butrynski JE, et al. (2010) N Engl J Med 363: 1727-33. Cools J, et al. (2002) Genes Chromosomes Cancer 34: 354-62. Cruzen ME, et al. (1993) Genomics 15: 692-3. Debelenko LV, et al. (2003) Lab Invest 83: 1255-65.

Prod. No. Product Label Tests* (Volume) Z-2137-50 ZytoLight SPEC CARS Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

101 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 101 FE057-1-20 ZytoLight ® SPEC NUP98 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NUP98 Dual Color The SPEC NUP98 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed for the Probe is a mixture of two direct labeled location involving the 11p15.4-p15.5 detection of translocations involving the probes hybridizing to the 11p15.4-p15.5 band, two orange/green fusion signals chromosomal region 11p15.4 harbor- band. The orange fluorochrome direct la- are expected representing two normal ing the NUP98 (nucleoporin 98, a.k.a. beled probe hybridizes proximal and the (non-rearranged) 11p15.4-p15.5 loci. A NUP96) gene. green fluorochrome direct labeled probe signal pattern consisting of one orange/ The nucleoporin NUP98, a component of hybridizes distal to the NUP98 gene. green fusion signal, one orange signal, the nuclear pore complex, is involved in and a separate green signal indicates NUP98 nucleocytoplasmic transport and exhibits one normal 11p15.4-p15.5 locus and multiple roles in RNA export from and one 11p15.4-p15.5 locus affected by a protein import into the nucleus. Rearrange- translocation or inversion. ments of the NUP98 gene have been implicated in the pathogenesis of several types of hematologic malignancies, including de novo and therapy-related Ideogram of chromosome 11 acute myeloid leukemia (AML), and also indicating the hybridization locations. myelodysplastic syndrome (MDS), chronic myelogenous leukemia (CML), and T-cell D11S2431 D11S4525 D11S3313 RH35854 acute lymphoblastic leukemia (T-ALL). 3’ 5’ NUP98 rearrangements result in the NUP98 fusion of the N-terminal region of NUP98, ~590 kb ~315 kb which is rich in phenylalanine-glycine (FG) Tel 11p15.4-p15.5 Cen repeats, to one of 29 different proteins. SPEC NUP98 Probe map (not to scale). Many of the NUP98 fusion partners are SPEC NUP98 Dual Color Break Apart Probe transcription factors of the homeobox hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus and to family. NUP98 fusions cause aberrant metaphase chromosomes of a normal cell. differentiation and increased proliferation when expressed in primary human hematopoietic cells. Malignancies with NUP98 rearrangements are associated with a poor prognosis and a poor treatment outcome indicating the usefulness of NUP98 as a prognostic biomarker. References Borrow J, et al. (1996) Nat Genet 12: 159-67. Fahrenkrog B (2014) New J Sci 2014: 468306. Takeda A & Yaseen NR (2014) Semin Cancer Biol 27: 3-10.

Prod. No. Product Label Tests* (Volume) Z-2266-50 ZytoLight SPEC NUP98 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

102 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 102 FE134-1-20 ZytoLight ® SPEC WT1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC WT1 Dual Color The SPEC WT1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 11p13 band, two translocations involving the chromosomal probes hybridizing to the 11p13 band. orange/green fusion signals are expected region 11p13 harboring the WT1 (Wilms The orange fluorochrome direct labeled representing two normal (non-rearranged) tumor 1) gene. probe hybridizes distal and the green fluo- 11p13 loci. A signal pattern consisting The WT1 gene is located on 11p13 and rochrome direct labeled probe hybridizes of one orange/green fusion signal, one encodes a zinc finger DNA-binding pro- proximal to the WT1 gene. orange signal, and a separate green signal tein that acts as a transcriptional activator indicates one normal 11p13 locus and one or repressor depending on the cellular 11p13 locus affected by a translocation. or chromosomal context. Inactivating WT1 mutations in the tumor suppressor gene WT1 have been identified in patients with Wilms’ tumor and in a subset of sporadic cancers. However, in desmoblastic small round cell Ideogram of chromosome 11 tumors (DSRCT) recurrent translocations indicating the hybridization locations. affecting the WT1 gene have been found. RH122877 RH121469 DSRCT is a highly aggressive mesen- D11S4662 SHGC-14132 chymal tumor that primarily affects male 3’ 5’

adolescents and young adults. The translo- WT1

cation t(11;22)(p13;q12.2) is detectable ~795 kb ~805 kb in virtually all DSRCT tested and results Tel 11p13 Cen SPEC WT1 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated in the fusion of the potent transcriptional SPEC WT1 Probe map (not to scale). by two orange/green fusion signals per nucleus. activator domain of the EWSR1 gene and the DNA-binding zinc-finger domains of the WT1 gene. The EWSR1-WT1 chimeric protein acts as an oncogenic transcription factor as evidenced by its ability to trans- form cells in vitro. While EWSR1 rearrangements are present in about 90% of DSRCT but are also frequently found in other small round blue cell neoplasms as e.g. Ewing sarcoma, WT1 translocations are exclusively found in DSRCT. Hence, detection of the t(11;22) by Fluorescence in situ Hybridization rep- Desmoblastic small round cell tumor tissue section with resents a valuable tool for the differential translocation affecting the 11p13 locus as indicated diagnosis of DSRCT. by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal. References Gerald WL, et al. (1995) Proc Natl Acad Sci U S A 92: 1028-32. Kim J, et al. (1998) Oncogene 16: 1973-9. Ladanyi M & Gerald W (1994) Cancer Res 54: 2837-40. Wang ZY, et al. (1993) J Biol Chem 268: 9172-5.

Prod. No. Product Label Tests* (Volume) Z-2142-50 ZytoLight SPEC WT1 Dual Color Break Apart Probe / 5 (50 µl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

103 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 103 FE066-1-20 ZytoLight ® SPEC SPI1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC SPI1 Dual Color The SPEC SPI1 Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to Probe is a mixture of two direct labeled cell lacking a translocation involving the detect rearrangements involving the probes hybridizing to the 11p11.2 11p11.2 band, two orange/green fusion chromosomal region 11p11.2 harboring band. The green fluorochrome direct signals are expected representing two the SPI1 (Spi-1 proto-oncogene, a.k.a. labeled probe hybridizes proximal to normal (non-rearranged) 11p11.2 loci. A PU.1, SPI-A) gene. the SPI1 gene at 11p11.2, the orange signal pattern consisting of one orange/ SPI1 is a member of the ETS family of fluorochrome direct labeled probe green fusion signal, one orange signal, transcription factors and is essential for hybridizes distal to the SPI1 gene at and a separate green signal indicates one the normal development of hematopoietic 11p11.2. normal 11p11.2 locus and one 11p11.2 stem cells. SPI1 rearrangements were locus affected by a translocation. detected in some pediatric T-cell acute lymphoblastic leukemia (T-ALL) cases SPI1 resulting in the fusion of the N-terminal region of the fusion partner (STMN1, TCF7, or BCL11B) to the C-terminal DNA binding ETS domain of the SPI1 protein. Hence, the resulting fusion proteins retain Ideogram of chromosome 11 the transcriptional activity inherent to SPI1. indicating the hybridization locations. SPI1 fusion positive cases show markedly elevated SPI1 expression, most likely RH76662 RH98825 RH103806 RH91838 because the fusion gene comes under the 3’ 5’ transcriptional control of the heterologous SPI1 promoter of the respective partner gene. ~485 kb ~445 kb SPEC SPI1 Dual Color Break Apart Probe hybridized Overexpression of SPI1 is thought to Tel 11p11.2 Cen to normal interphase cells as indicated by two contribute to T-cell leukemogenesis. orange/green fusion signals in each nucleus and SPEC SPI1 Probe map (not to scale). to metaphase chromosomes of a normal cell. Moreover, T-ALL patients with SPI1 fusion show a uniformly poor overall survival and seem to be incurable with current standard chemotherapy. This underscores the importance of detecting this subset of patients by FISH so that they may receive more intensive or alternative therapies.

References Homminga I, et al. (2011) Cancer Cell 19: 484-97. Liu Y, et al. (2017) Nat Genet 49: 1211-8. Seki M, et al. (2017) Nat Genet 49: 1274-81.

Bone marrow smear with translocation of the SPI1 gene as indicated by one non-rearranged orange/green fusion signal, one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2291-50 ZytoLight SPEC SPI1 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

104 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 104 FE158-1-20 ZytoLight ® SPEC CCND1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CCND1 Dual Color The SPEC CCND1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 11q13.2-q13.3 translocations involving the chromosomal probes hybridizing to the 11q13.2-q13.3 band, two orange/green fusion signals region 11q13.3 harboring the CCND1 band. The orange fluorochrome direct are expected representing two normal gene. The CCND1 gene (cyclin D1, a.k.a. labeled probe hybridizes distal to and (non-rearranged) CCND1 loci. A signal BCL1 or PRAD1) encodes a regulatory covers the CCDN1 gene, while the green pattern consisting of one orange/green subunit of cyclin-dependent kinases. fluorochrome direct labeled probe hybrid- fusion signal, one orange signal, and Translocations involving the chromosomal izes proximal to that gene. a separate green signal indicates one region t(11;14) (q13.3;q32.3) are consid- normal CCND1 locus and one CCND1 ered to be characteristic for mantle cell locus affected by an 11q13.2-q13.3 trans- lymphomas (MCL) but have also been location. identified in other lymphoproliferative disorders (LPDs), such as B-prolymphocytic CCND1 leukemia, and, less frequently, in plasma cell myelomas, B-cell chronic lymphocytic leukemia, and in splenic lymphomas with Ideogram of chromosome 11 villous lymphocytes (SLVL). indicating the hybridization locations. The t(11;14) rearrangement often leads to RH47914 PMC133966P1 overexpression of the CCND1 protein. De- RH46970 RH48109 termination of translocations involving the 5’ 3’ 5’ 3’

chromosomal region 11q13.3 can also MYEOV CCND1 help to distinguish MCL from other chronic ~455 kb ~580 kb SPEC CCND1 Dual Color Break Apart Probe lymphoproliferative disorders. Since the Cen 11q13.2-q13.3 Tel hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. course of MCL is aggressive, and its re- SPEC CCND1 Probe map (not to scale). sponse to chemotherapy is poor, differential diagnosis is clinically important. Additionally, it was also shown that a renal oncocytoma (RO) specific breakpoint is located in band 11q13.3, involving the CCND1 locus. The histologic features of RO may overlap with those of chromophobe renal cell carcinoma (ChRCC). References Bentz JS, et al. (2004) Cancer 102: 124-31. Fluorescence in situ Hybridization (FISH) Bosch F, et al. (1997) Cancer 82: 567-75. Sinke RJ, et al. (1997) Cancer Genet Cytogenet 96: 95-101. can be used as a diagnostic tool for differ- Sukov WR, et al. (2007) Hum Pathol 40: 1296-303. Tarsitano M, et al. (2009) Cancer Genet Cytogenet 195: 164-7. entiation of RO from ChRCC. Vaandrager JW, et al. (1996) Blood 4: 1177-82. Bone marrow biopsy section with translocation affecting the 11q13.2-q13.3 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2108-50 ZytoLight SPEC CCND1 Dual Color Break Apart Probe / 5 (50 μl) Z-2108-200 ZytoLight SPEC CCND1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

105 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 105 FE041-1-20 ZytoLight ® SPEC CCND1/CEN 11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC CCND1/CEN 11 The SPEC CCND1/CEN 11 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- ange and two green signals are expected. detection of CCND1 gene amplification chrome direct labeled CEN 11 probe In a cell with amplification of the CCND1 frequently observed in breast cancer and specific for the alpha satellite centromeric gene locus, multiple copies of the green other human tumors. region of chromosome 11 (D11Z1) and a signal or large green signal clusters will The cyclin D1 gene CCND1 (a.k.a. BCL1 green fluorochrome direct labeled SPEC be observed. or PRAD1) is located in the chromosomal CCND1 probe specific for the CCND1 region 11q13.3 and encodes a regula- gene at 11q13.3. tory subunit of cyclin-dependent kinases that promote progression through the cell cycle. The proto-oncogene CCND1 is amplified CEN 11 (D11Z1) in a number of solid tumors including CCND1 approx. 20% of all human breast cancer cases and about 30% of squamous cell carcinomas of the esophagus and the Ideogram of chromosome 11 head and neck region. Amplification of indicating the hybridization locations. chromosomal material from 11q13.3 har- SHGC-149381 RH48109 boring the CCND1 gene is discussed as a 5’ 3’ SPEC CCND1/CEN 11 Dual Color Probe hybridized prognostic marker in terms of metastasis, to normal interphase cells as indicated by two CCND1 orange and two green signals in each nucleus. tumor recurrence, and survival for several tumor entities. In gastrointestinal stromal ~825 kb Cen 11q13.3 Tel tumors (GIST), CCND1 amplification was SPEC CCND1 Probe map (not to scale). found in 16% of high-risk tumors and was absent in low- or intermediate-risk tumors indicating the prognostic relevance of this genetic alteration in GIST.

References Al-Kuraya K, et al. (2004) Cancer Res 64: 8534-40. Courjal F, et al. (1997) Cancer Res 57: 4360-7. Motokura T, et al. (1991) Nature 350: 512-5. Ormandy CJ, et al. (2003) Breast Cancer Res Treat 78: 323-35. Schuuring E (1995) Gene 159: 83-96. Tornillo L, et al. (2005) Lab Invest 85: 921-31. Xiong Y, et al. (1991) Cell 65: 691-9.

Polysomy of chromosome 11 as indicated by three orange (CEN 11) and three green (CCND1) signals in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2071-50 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe / 5 (50 μl) Z-2071-200 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

106 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 106 FE013-1-20 ZytoLight ® SPEC CCND1/IGH Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC CCND1/IGH Dual The SPEC CCND1/IGH Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed to de- Fusion Probe is a mixture of an orange flu- ange and two green signals are expected. tect translocation t(11;14)(q13.3;q32.3) orochrome direct labeled CCND1 probe A reciprocal CCND1/IGH translocation frequently found in mantle cell lymphomas. spanning the major translocation cluster leads to two orange/green fusion signals The translocation juxtaposes the CCND1 (MTC) region comprising about 120 kb indicating both rearranged chromosomes. gene (cyclin D1, a.k.a. PRAD1 and BCL1) upstream of CCND1 and a green fluoro- Additionally, the non-rearranged chromo- next to the IGH (immunoglobulin heavy chrome direct labeled IGH probe spanning somes are indicated by one orange signal locus, a.k.a. IGH@) locus and results in the breakpoint cluster region of IGH. and a separate green signal, respectively. constitutive overexpression of CCND1. The translocation t(11;14)(q13.3;q32.3) that involves the CCND1 and IGH gene regions is detected in up to 95% of patients with mantle cell lymphomas CCND1 (MCL) and is considered to be the genetic hallmark of this subtype of low-grade peripheral B-cell neoplasms. However, the t(11;14) has also been identified in other lymphoproliferative disorders (LPDs), such as B-prolymphocytic leukemia (PLL), and, less frequently, in plasma cell myelomas, B-cell chronic lymphocytic leukemia, and SPEC CCND1/IGH Dual Color Dual Fusion Probe in splenic lymphomas with villous lympho- hybridized to normal interphase cells as indicated by two orange and two green signals in each nucleus. cytes (SLVL). Ideograms of chromosomes 11 (above) and 14 (below) indicating the hybridization locations. Since the course of MCL is aggressive, and its response to standard chemother- RH51714 RH48109 5’3’ 5’3’ apy is poor, differential diagnosis from MTC other chronic lymphoproliferative disorders MYEOV CCND1 via detection of the t(11;14) translocation ~1.5 Mb might be of great clinical importance. Cen 11q13.3 Tel

References SPEC CCND1 Probe map (not to scale). Bentz JS, et al. (2004) Cancer 102: 124-31. Li JY, et al. (1999) Am J Pathol 154: 1449-52. BV448316 RH119884 Siebert R, et al. (1998) Ann Oncol 9: 519-26. 3’5’ 3’ 5’ Vaandrager JW, et al. (1996) Blood 88: 1177-82.

JAG2 IGH

~1.5 Mb Cen 14q32.33 Tel Section of an iliac crest biopsy with translocation SPEC IGH Probe map (not to scale). affecting the CCND1/IGH loci as indicated by one separate orange signal, one separate green signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2125-50 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe / 5 (50 μl) Z-2125-200 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

107 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 107 FE052-1-20 ZytoLight ® SPEC MAML2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC MAML2 Dual Color The SPEC MAML2 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 11q21 band two the translocation t(11;19)(q21;p13.1) probes hybridizing to the 11q21 band. orange/green fusion signals are expected specific for mucoepidermoid carcinomas. The green fluorochrome direct labeled representing two normal (non-rearranged) The mucoepidermoid carcinoma is the probe hybridizes distal to the MAML2 11q21 loci. A signal pattern consisting of most common malignant tumor of the gene, the orange fluorochrome direct one orange/green fusion signal, one or- salivary gland. With about 30-50% labeled probe hybridizes proximal to that ange signal, and a separate green signal of all cases, the translocation t(11;19) gene. indicates one normal 11q21 locus and (q21;p13.1) is the most frequent chro- one 11q21 locus affected by the trans- mosomal aberration in mucoepidermoid location specific for mucoepidermoid carcinomas. In some cases the t(11;19) carcinomas. is the sole chromosomal anomaly and in other cases the t(11;19) was found either as a more complex translocation involving MAML2 other chromosomes or together with other abnormalities. Ideogram of chromosome 11 References indicating the hybridization locations. Bishop JA, et al. (2014) Head Neck Pathol 8: 287-90. Camelo-Piragua SI, et al. (2009) Hum Pathol 40: 887-92. Chiosea SI, et al. (2012) Laryngoscope 122: 1690-4. RH119986 D11S3037 El-Naggar A, et al. (1996) Cancer Genet Cytogenet 87: 29-33. RH2958 RH106430 Jee KJ, et al. (2013) Mod Pathol 26: 213-22. 3’ 5’ Lei Y & Chiosea SI (2012) Head Neck Pathol 6: 166-70. Noda H, et al. (2013) Cancer Sci 104: 85-92. Nordkvist A, et al. (1994) Cancer Genet Cytogenet 74: 77-83. MAML2 Rotellini M, et al. (2012) J Oral Pathol Med 41: 615-20. Schwarz S, et al. (2011) Histopathology 58: 557-70. ~370 kb ~680 kb Schwarz S, et al. (2011) Int J Clin Exp Pathol 4: 336-48. Winnes M, et al. (2007) Genes Chromosomes Cancer 46: 559-63. Cen 11q21 Tel Zhu F, et al. (2014) PLoS One 9: e94399. SPEC MAML2 Probe map (not to scale). SPEC MAML2 Dual Color Break Apart Probe hybridized to abnormal nuclei containing two normal chromosomes 11 as indicated by two orange/green signal pairs and a derivative chromosome 11 with a translocation involving the 11q21 band as indicated by one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2014-50 ZytoLight SPEC MAML2 Dual Color Break Apart Probe / 5 (50 μl) Z-2014-200 ZytoLight SPEC MAML2 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

108 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 108 FE002-1-20 ZytoLight ® SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC BIRC3/MALT1 Dual The SPEC BIRC3/MALT1 Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed to Fusion Probe is a mixture of a green direct ange and two green signals are expected. detect translocations involving the chro- labeled BIRC3 probe spanning the BIRC3 A reciprocal translocation involving two mosomal region 11q22.2 harboring the gene region at 11q22.1-q22.2 and an breakpoints splits the two signals and BIRC3 (baculoviral IAP repeat containing orange direct labeled MALT1 probe generates a fusion signal on each of the 3, a.k.a. API2) gene and the chromosomal spanning the MALT1 gene region at chromosomes involved. The chromosomal region 18q21.32 harboring the MALT1 18q21.31-q21.32. regions which are not translocated are (MALT1 paracaspase, a.k.a. MLT) gene. indicated by the single orange and green The recurrent translocation t(11;18) signal, respectively. (q22.2;q21.3) is frequently found in muco- sa-associated lymphoid tissue (MALT) lymphoma which represents the most common extranodal B-cell tumor and accounts for BIRC3 5-10% of all non-Hodgkin lymphoma. The translocation results in the expression of chimeric fusion transcripts comprising the N-terminal end of the apoptosis inhibitor BIRC3 which is highly expressed in adult lymphoid tissue and C-terminal parts of the MALT1 MALT1 protease. Ideograms of chromosomes 11 (above) and 18 (be- The BIRC3/MALT1 fusion protein was low) indicating the hybridization locations. shown to induce proteolytic cleavage of SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe hybridized to normal interphase cells as indicated by RH122451 D11S1192 NF-kappa-B-inducing kinase (NIK) ultimate- 5’ 3’ two orange and two green signals in each nucleus. ly resulting in constitutive non-canonical NF-kappa-B signaling, enhanced B-cell BIRC3 adhesion, and apoptosis resistance. It is ~825 kb Cen 11q22.1-q22.2 Tel assumed that disruption of the BIRC3-NIK SPEC BIRC3 Probe map (not to scale). interaction and/or blocking of MALT1 protease or NIK kinase activity could KAM39 D18S529 5’ 3’ represent new treatment approaches for refractory t(11;18)-positive MALT lymphoma. MALT1 References ~705 kb Dierlamm J, et al. (1999) Blood 93: 3601-9. Cen Tel Dierlamm J, et al. (2000) Blood 96: 2215-8. 18q21.31-q21.32 Morgan JA, et al. (1999) Cancer Res 59: 6205-13. Rosebeck S, et al. (2011) Science 331: 468-72. SPEC MALT1 Probe map (not to scale).

MALT lymphoma tissue section with translocation affecting the BIRC3/MALT1 loci as indicated by one separate orange signal, one separate green signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2146-50 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe / 5 (50 μl) Z-2146-200 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

109 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 109 FE064-1-20 ZytoLight ® SPEC ATM/CEN 11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ATM/CEN 11 Dual The SPEC ATM/CEN 11 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detec- is a mixture of an orange fluorochrome ange and two green signals are expected. tion of deletions affecting the ATM (ATM direct labeled SPEC ATM probe specific In a cell with deletion of the ATM gene serine/threonine kinase, a.k.a. ATA, TEL1) for the ATM gene at 11q22.3 and a locus, one or no copy of the orange signal gene. green fluorochrome direct labeled CEN will be observed. The ATM gene is located on 11q22.3 and 11 probe specific for the alpha satellite encodes a protein kinase. This kinase reg- centromeric region of chromosome 11 ulates the response to DNA double strand (D11Z1). breaks by triggering signaling, which synchronizes DNA repair, cell-cycle arrest, and apoptosis. In chronic lymphocytic leukemia (CLL), CEN 11 (D11Z1) recurrent alterations include deletions at chromosome 13q14, 11q22.3, 17p13 and 6q21, trisomy of chromosome 12 ATM and IGH translocation. It was shown that compared to the absence of cytogenetic Ideogram of chromosome 11 indicating the hybridization locations. abnormalities, 17p13 deletion conferred SPEC ATM/CEN 11 Dual Color Probe hybridized to normal interphase cells as indicated by two the worst prognosis, followed by 11q22.3 RH75994 RH111454 orange and two green signals in each nucleus and to deletion and trisomy 12, whereas 13q14 5’ 3’ metaphase chromosomes of a normal cell.

deletion as a sole abnormality was found ATM

to be associated with a good prognosis. ~425 kb Chromosome 11q22.3 deletion is detect- Cen 11q22.3 Tel ed in approximately 20% of all CLL pa- SPEC ATM Probe map (not to scale). tients at diagnosis. These patients exhibit rapid disease progression and shorter treatment-free and overall survival times. Moreover, CLL subsets with 11q deletion are associated with an elevation of gene copy number alterations representing genomic instability. Hence, FISH can be used for the detection CLL with deletion affecting the ATM locus as of 11q22.3 deletions in CLL to predict indicated by one orange signal in each nucleus. disease progression and overall survival.

References Döhner H, et al. (2000) N Engl J Med 343: 1910-6. Stankovic T & Skowronska A (2014) Leuk Lymphoma 55: 1227-39. Zenz T, et al. (2010) Best Pract Res Clin Haematol 23: 71-84.

Prod. No. Product Label Tests* (Volume) Z-2297-50 ZytoLight SPEC ATM/CEN 11 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

110 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 110 FE162-1-20 ZytoLight ® SPEC ATM/CEN 12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ATM/CEN 12 Dual The SPEC ATM/CEN 12 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detec- is a mixture of an orange fluorochrome ange and two green signals are expected. tion of deletions affecting the ATM (ATM direct labeled SPEC ATM probe specific In a cell with deletion of the ATM gene serine/threonine kinase, a.k.a. ATA, TEL1) for the ATM gene at 11q22.3 and a green locus, one or no copy of the orange signal gene as well as for the enumeration of fluorochrome direct labeled CEN 12 will be observed. In a cell with trisomy or chromosome 12. probe specific for the alpha satellite centro- polysomy 12, three or more copies of the The ATM gene is located on 11q22.3 and meric region of chromosome 12 (D12Z3). green signal will be observed, respectively. encodes a protein kinase. This kinase regulates the response to DNA double strand breaks by triggering signaling, which synchronizes DNA repair, cell-cycle arrest and apoptosis. In chronic lymphocytic leukemia (CLL), recurrent alterations include deletions at ATM chromosome 13q14, 11q22.3, 17p13 and 6q21, trisomy of chromosome 12 and IGH translocation. It was shown that compared to the absence of cytogenetic abnormalities, 17p13 deletion conferred the worst prognosis, followed by 11q22.3 SPEC ATM/CEN 12 Dual Color Probe hybridized to normal interphase cells as indicated by two orange deletion and trisomy 12, whereas 13q14 and two green signals in each nucleus and deletion as a sole abnormality was found to metaphase chromosomes of a normal cell. Ideograms of chromosomes 11 (above) and 12 to be associated with a good prognosis. (below) indicating the hybridization locations. Trisomy 12 and 11q22.3 deletion are both detected in about 20% of CLL cases. RH75994 RH111454 5’ 3’ Patients with 11q deletion exhibit rapid disease progression and shorter treat- ATM ment-free and overall survival times. More- ~425 kb over, CLL subsets with 11q deletion are Cen 11q22.3 Tel associated with an elevation of gene copy SPEC ATM Probe map (not to scale). number alterations representing genomic instability. Trisomy 12 is associated with a median

survival and an atypical morphology. CLL with deletion of the ATM gene and Hence, FISH can be used to predict amplification affecting the centromeric region of chromosome 12 as indicated by one orange disease progression and overall survival in signal and five or more green signals in each nucleus. CLL patients.

References Döhner H, et al. (2000) N Engl J Med 343: 1910-6. Glassman AB & Hayes KJ (2005) Cancer Genet Cytogenet 158: 88-91. Stankovic T & Skowronska A (2014) Leuk Lymphoma 55: 1227-39. Zenz T, et al. (2010) Best Pract Res Clin Haematol 23: 71-84.

Prod. No. Product Label Tests* (Volume) Z-2296-50 ZytoLight SPEC ATM/CEN 12 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

111 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 111 FE161-1-20 ZytoLight ® SPEC TP53/ATM Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC TP53/ATM Dual The SPEC TP53/ATM Dual Color Probe is Using the SPEC TP53/ATM Dual Color Color Probe is designed for the detection a mixture of an orange fluorochrome di- Probe in a normal interphase nucleus, of deletions affecting the genes TP53 and rect labeled SPEC TP53 probe hybridizing two orange and two green signals are ATM. to the TP53 gene in the chromosomal re- expected. In a cell with deletions affecting CLL (chronic lymphocytic leukemia) is the gion 17p13.1 and a green fluorochrome the TP53 gene locus, a reduced number of most common form of leukemia in Western direct labeled SPEC ATM probe specific orange signals will be observed. Deletions population. for the ATM gene at 11q22.3. affecting only parts of the TP53 locus TP53 (tumor protein p53, a.k.a. p53) might result in a normal signal pattern TP53 gene deletions have been detected in with orange signals of reduced size. In a patients with CLL, multiple myeloma (MM), cell with ATM gene deletions, a reduced and acute myeloid leukemia (AML). In CLL number of green signals will be observed. patients, allelic loss of the short arm of Deletions affecting only parts of the ATM chromosome 17 is associated with treat- locus might result in a normal signal pat- ment failure with alkylating agents and tern with green signals of reduced size. short survival times. The ATM (ATM serine/threonine kinase) gene is located on 11q22.3 and encodes a protein kinase which is involved in cell cycle regulation, including TP53 activa- ATM tion. CLL patients with 11q deletion exhibit rapid disease progression and inferior Ideograms of chromosomes 17 (above) and 11 survival. (below) indicating the hybridization locations.

Hence, in combination with further bio- RH76489 SGC30770 3’ 5’ logical markers, morphology and clinical SPEC TP53/ATM Dual Color Probe hybridized to information FISH is a valuable tool to TP53 bone marrow biopsy section with deletions of the ATM and the TP53 genes as indicated by one predict disease progression and overall ~165 kb green and one orange signal in each nucleus. survival in CLL patients. Tel 17p13.1 Cen

References SPEC TP53 Probe map (not to scale). Pettitt AR, et al. (2001) Blood 98: 814-22. Ripollés L, et al. (2006) Cancer Genet Cytogenet 171: 57-64. Shanafelt TD, et al. (2006) Ann Intern Med 145: 435-47. RH75994 RH111454 Stilgenbauer S, et al. (2002) Leukemia 16: 993-1007. 5’ 3’

ATM

~425 kb Cen 11q22.3 Tel

SPEC ATM Probe map (not to scale).

SPEC TP53/ATM Dual Color Probe hybridized to bone marrow biopsy section with deletion of the ATM gene as indicated by one green signal in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2159-50 ZytoLight SPEC TP53/ATM Dual Color Probe / 5 (50 μl) Z-2159-200 ZytoLight SPEC TP53/ATM Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

112 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 112 FE075-1-20 ZytoLight ® SPEC D13S319/13q34/CEN 12 Triple Color Probe

Background Probe Description Results The ZytoLight ® SPEC D13S319/13q34/ The SPEC D13S319/13q34/CEN 12 Using the SPEC D13S319/13q34/ CEN 12 Triple Color Probe is designed for Triple Color Probe is a mixture of an CEN 12 Triple Color Probe in a normal the detection of D13S319 deletions as well orange fluorochrome direct labeled SPEC interphase nucleus, two orange, two as for the enumeration of chromosome 12. D13S319 probe specific for the D13S319 green, and two blue signals are expect- CLL (chronic lymphocytic leukemia) is the locus at 13q14.2, a blue fluorochrome ed. In a cell with deletions affecting the most common form of leukemia in Western direct labeled SPEC 13q34 probe specific D13S319 locus, a reduced number of population. for the chromosomal region 13q34 and a orange signals will be observed. Deletions The most frequent aberration in CLL is green fluorochrome direct labeled CEN 12 affecting only parts of the D13S319 locus the deletion of 13q14 which involves the probe specific for the alpha satellite centro- might result in a normal signal pattern D13S319 locus and which is associated meric region of chromosome 12 (D12Z3). with orange signals of reduced size. In a with a favorable prognosis if occurring as The SPEC 13q34 probe is specific for the cell with trisomy or polysomy 12, three or the sole genetic aberration. Deletions of LAMP1 (lysomal associated membrane more copies of the green signal will be the long arm of chromosome 13 are also protein 1) gene region in 13q34. Due to observed, respectively. frequently detected in patients with aggres- cross-hybridizations of chromosome 13 al- sive non-Hodgkin lymphoma (NHL) and pha satellites to other centromeric regions, have been found to represent an adverse probes specific for 13q34 are frequently prognostic factor in MM. used for chromosome 13 copy number Trisomy 12 represents another frequent detection. chromosomal aberration in CLL, detected in about 20% of CLL cases. Trisomy 12 as single aberration is associated with an D13S319 intermediate prognostic outcome. Hence, in combination with further bio- 13q34 SPEC D13S319/13q34/CEN 12 Triple Color Probe logical markers, morphology and clinical hybridized to bone marrow biopsy section with deletion of the D13S319 locus as indicated by one information FISH is a valuable tool to orange signal and two blue signals in each nucleus. predict disease progression and overall survival in CLL patients. References Chang H, et al. (1999) Leukemia 13: 105-9. Dal Bo M, et al. (2011) Genes Chromosomes Cancer 50: 633-43. Ouillette P, et al. (2011) Clin Cancer Res 21: 6778-90. Ripollés L, et al. (2006) Cancer Genet Cytogenet 171: 57-64. Shanafelt TD, et al. (2006) Ann Intern Med 145: 435-47. Stilgenbauer S, et al. (2002) Leukemia 16: 993-1007. Ideograms of chromosomes 13 (above) and 12 D13S319 RH45599 D13S1474 (below) indicating the hybridization locations.

3’ 5’ 5’ 3’ SHGC111313 RH39633 5’3’ 5’ 3’

DLEU2 DLEU1 PROZ LAMP1 MIR16-1MIR15A SPEC D13S319/13q34/CEN 12 Triple Color ~ 630 kb ~175 kb Probe hybridized to bone marrow biopsy section Cen 13q14.2 Tel Cen 13q34 Tel with trisomy of chromosome 12 as indicated by three green signals in each nucleus. SPEC D13S319 Probe map (not to scale). SPEC 13q34 Probe map (not to scale). Prod. No. Product Label Tests* (Volume) Z-2160-50 ZytoLight SPEC D13S319/13q34/CEN 12 Triple Color Probe / / 5 (50 μl) Z-2160-200 ZytoLight SPEC D13S319/13q34/CEN 12 Triple Color Probe / / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

113 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 113 FE128-1-20 ZytoLight ® SPEC D13S319/13q34 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC D13S319/13q34 The SPEC D13S319/13q34 Dual Color Using the SPEC D13S319/13q34 Dual Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- Color Probe in a normal interphase nucle- detection of deletions on the long arm of chrome direct labeled SPEC D13S319 us, two orange and two green signals are chromosome 13. probe specific for the D13S319 locus at expected. In a cell with deletions affecting Chronic lymphocytic leukemia (CLL) is 13q14.2 and a green fluorochrome direct the D13S319 locus, a reduced number of the most common form of leukemia in the labeled SPEC 13q34 probe specific for orange signals will be observed. Deletions Western population. It is characterized the chromosomal region 13q34. affecting only parts of the D13S319 locus by a marked variable outcome, from an The SPEC 13q34 probe is specific for the might result in a normal signal pattern indolent clinical course to more aggres- LAMP1 (lysosome-associated membrane with orange signals of reduced size. If sive forms, depending on the aberration. protein 1) gene region in 13q34. Due to deletions affect the D13S319 locus as well The most frequent aberration in CLL is the cross-hybridizations of chromosome 13 al- as the 13q34 locus, this might result in deletion of 13q14.2 which involves the pha satellites to other centromeric regions, a reduced number of orange and green D13S319 locus and which is associated probes specific for 13q34 are frequently signals. with a favorable prognosis if occurring as used for chromosome 13 copy number the sole genetic aberration. detection. Deletions of the long arm of the chromosome 13 are also frequently detected in patients with aggressive non-Hodgkin D13S319 lymphoma (NHL) and have been found to represent an adverse prognostic factor in multiple myeloma (MM). The telomeric 13q34 region 13q34 is used as control region, to Ideogram of chromosome 13 distinguish between an interstitial deletion indicating the hybridization locations.

and a complete loss of the 13q arm. D13S319 RH45599 D13S1474 In combination with further biological 3’ 5’ 5’ 3’ SPEC D13S319/13q34 Dual Color Probe markers, morphology, and clinical infor- hybridized to normal interphase cells as indicated by mation, Fluorescence in situ Hybridization DLEU2 DLEU1 two green and two orange signals in each nucleus.

(FISH) can be a valuable tool to predict MIR16-1MIR15A disease progression and overall survival in ~175 kb Cen 13q14.2 Tel CLL patients. SPEC D13S319 Probe map (not to scale). References Chang H, et al. (1999) Leukemia 13: 105-9. Dal Bo M, et al. (2011) Genes Chromosomes Cancer 50: 633-43. SHGC111313 RH39633 Liu Y, et al. (1998) Blood 86: 1911-15. 5’ 3’ 5’ 3’ La Starza R, et al. (2018) Mol Cytogenet 11: 6. Ouillette P, et al. (2011) Clin Cancer Res 21: 6778-90. Stilgenbauer S, et al. (1998) Oncogene 16: 1891-7. PROZ LAMP1 ~630 kb Cen 13q34 Tel

SPEC 13q34 Probe map (not to scale).

Bone marrow biopsy section with deletion of the D13S319 locus as indicated by one orange and two green signals in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2280-50 ZytoLight SPEC D13S319/13q34 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

114 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 114 FE152-1-20 ZytoLight ® SPEC 11q gain/loss Triple Color Probe

Background Probe Description Results The ZytoLight ® SPEC 11q gain/loss Triple The SPEC 11q gain/loss Triple Color In a normal interphase nucleus, two Color Probe is designed to detect 11q Probe is a mixture of a green fluoro- green, two orange, and two blue signals alterations. chrome direct labeled probe hybridizing are expected. In a cell with amplification A subset of lymphomas with gene expres- in the MGR at 11q23.3, an orange fluo- at 11q23.3 and deletion at 11q24.3, sion and pathological characteristics of rochrome direct labeled probe hybridizing multiple copies of the green signals and a Burkitt lymphomas (BL) but absence of MYC in the MLR at 11q24.3, and a blue fluo- reduced number of orange signals will be translocation has been recently described rochrome direct labeled CEN 11 probe observed. which carries 11q proximal gains and telo- specific for the alpha satellite centromeric meric losses. It is assumed that this aberra- region of chromosome 11 (D11Z1). tion leads to co-deregulation of oncogenes and tumor suppressor genes which are located in the affected chromosomal regions. The current WHO classification introduced CEN 11 (D11Z1) this new provisional entity as Burkitt-like lymphoma with 11q aberration. The minimal region of gain (MGR) and loss (MLR) was 11q23.3 defined at 11q23.3 and at 11q24.1-q25, 11q24.3 Ideogram of chromosome 11 respectively, based on the studies by indicating the hybridization locations. Ferreiro et al. (2015) and Salaverria et al. (2014). Potential oncogenes located in the D20S1118 D11S4612 D11S4559 RH27766 SPEC 11q gain/loss Triple Color Probe hybridized MGR are USP2 and PAFAH1B2. The can- 5’ 3’ 5’ 3’ to normal interphase cells as indicated by two green, two orange, and two blue signals per nucleus and didate tumor suppressor genes in the MLR TMPRSS4 FLI1 to metaphase chromosomes of a normal cell. comprise, e.g., FLI1 and ETS1. ~710 kb ~455 kb The 11q-gain/loss pattern in high-grade Cen 11q23.3-q24.3 Tel B-cell lymphoma is significantly more fre- SPEC 11q Probe map (not to scale). quent in lymphoma occurring in the setting of transplantation and immunosuppression than in immunocompetent patients. This suggests that immunosuppression may favor its formation. As identification of patients with the 11q-gain/loss aberration is clinically important but cytogenetically challenging, FISH assay is a useful diagnostic tool to evaluate both post-transplant and immunocompetent Burkitt-like lymphoma tissue section with Burkitt and Burkitt-like lymphoma patients. 11q aberration as indicated by three green signals and one orange signal indicating the References Ferreiro JF, et al. (2015) Haematologica 100: e275-9. gain and loss at 11q, respectively. Salaverria I, et al. (2014) Blood 123: 1187-98. Swerdlow SH, et al. (2016) Blood 127: 2375-90.

Prod. No. Product Label Tests* (Volume) Z-2216-50 ZytoLight SPEC 11q gain/loss Triple Color Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

115 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 115 FE126-1-20 ZytoLight ® SPEC KMT2A Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC KMT2A Dual Color The SPEC KMT2A Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cation involving the 11q23.3 band, two translocations involving the chromosomal probes hybridizing to the 11q23.3 band. orange/green fusion signals are expected region 11q23.3 harboring the KMT2A The green fluorochrome direct labeled representing two normal (non-rearranged) gene. The KMT2A (lysine methyltrans- probe hybridizes proximal to the KMT2A 11q23.3 loci. A signal pattern consisting ferase 2A, a.k.a. MLL) gene encodes a gene, and the orange fluorochrome direct of one orange/green fusion signal, one histone lysine N-methyltransferase and is labeled probe hybridizes distal to the orange signal, and a separate green involved in a variety of cellular processes, KMT2A gene region. This probe is signal indicates one normal 11q23.3 including hematopoiesis, DNA damage approved to be used with a hybridization locus and one 11q23.3 locus affected by response, and cell cycle control. time of 2 hours on cytological specimens. a translocation. Translocations involving the KMT2A gene are identified in 5-6% of all acute myeloid leukemias (AML) and 5-10% of all acute lymphoblastic leukemias (ALL). The frequency of translocations involving the KMT2A gene is significantly higher in infants with AML (50%) as well as with ALL KMT2A (80%). More than 30 fusion partners are Ideogram of chromosome 11 documented for KMT2A, the most common indicating the hybridization locations. translocations are t(4;11) and t(11;19) in D20S1118 D11S1371 ALL, and t(6;11), t(9;11), and t(11;19) in RH120709 D11S2571 5’ 3’ AML patients. SPEC KMT2A Dual Color Break Apart Probe Between 1-15% of cancer patients treated KMT2A hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. with DNA topoisomerase II inhibitor ~710 kb ~595 kb develop therapy-related leukemia (t-AML) Cen 11q23.3 Tel associated with KMT2A translocations. SPEC KMT2A Probe map (not to scale). Generally, the presence of KMT2A rearrangements in patients with acute leukemia indicates a less favorable prognosis. However, recent studies suggest that the specific KMT2A translocation partner may influence response to therapy and overall References prognosis depending on the clinical context. Broeker PL, et al. (1996) Blood 87: 1912-22. De Braekeleer M, et al. (2005) Anticancer Res 25: 1931-44. Hence, detection of KMT2A translocations Ford DJ & Dingwall AK (2015) Cancer Genet 208: 178-91. Gindin T, et al. (2015) Hematol Oncol 33: 239-46. by Fluorescence in situ Hybridization may Keefe JG, et al. (2010) J Mol Diagn 12: 441-52. Langer T, et al. (2003) Genes Chromosomes Cancer 36: 393-401. Bone marrow smear with translocation of be of diagnostic and prognostic relevance. Wechsler DS, et al. (2003) Genes Chromosomes Cancer 36: 26-36. the KMT2A gene as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2193-50 ZytoLight SPEC KMT2A Dual Color Break Apart Probe / 5 (50 μl) Z-2193-200 ZytoLight SPEC KMT2A Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

116 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 116 FE096-1-20 ZytoLight ® SPEC ZNF384 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ZNF384 Dual Color The SPEC ZNF384 Dual Color Break In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Apart Probe is a mixture of two direct la- cation involving the 12p13.31 band, two translocations involving the chromosomal beled probes hybridizing to the 12p13.31 orange/green fusion signals are expected, region 12p13.31 harboring the ZNF384 band. The orange fluorochrome direct representing two normal (non-rearranged) gene. labeled probe hybridizes proximal to the 12p13.31 loci. A signal pattern consisting The ZNF384 (zinc-finger protein 384, ZNF384 gene, the green fluorochrome of one orange/green fusion signal, one a.k.a. CIZ) gene encodes a transcription direct labeled probe hybridizes distal to orange signal, and a separate green sig- factor involved in the regulation of matrix the ZNF384 gene. nal indicates one normal 12p13.31 locus metalloproteinases. and one 12p13.31 locus affected by a ZNF384 Rearrangements of the ZNF384 gene are ZNF384 translocation. recurrent in acute leukemia and are most frequently found in precursor B-cell acute lymphoblastic leukemia (BCP-ALL) in children and young adults with an incidence of about 3-4%. ZNF384-related fusion genes with multiple fusion partners have Ideogram of chromosome 12 been found to define a distinct subgroup indicating the hybridization locations. of pediatric BCP-ALL with a characteristic RH106101 RH39803 immunophenotype. Known translocation RH52677 RH102330 3’ 5’ partners are TCF3 (19p13.3), EWSR1

(22q12.2), TAF15 (17q12), EP300 ZNF384

(22q13.2), ARID1B (6q25.3), ~755 kb ~380 kb Tel 12p13.31 Cen CREBBP (16p13.3), and BMP2K SPEC ZNF384 Dual Color Break Apart Probe (4q21.21) with TCF3 being the most prev- SPEC ZNF384 Probe map (not to scale). hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus and to alent. The breakpoints are located within metaphase chromosomes of a normal cell. the ZNF384 gene. However, the balanced translocations are resulting in fusion genes including the complete protein coding information. Since ZNF384-related fusion genes are difficult to detect by common G-banding, investigation by FISH may be of diagnostic and prognostic relevance.

References Hirabayashi S, et al. (2017) Haematologica 102: 118-29. Krance RA, et al. (1992) Leukemia 6: 251-5. La Starza R, et al. (2005) Leukemia 19: 1696-9. Shago M, et al. (2016) Pediatr Blood Cancer 63: 1915-21.

Bone marrow smear of an ALL case with translocation of the ZNF384 gene as indicated by one orange/green fusion signal, one separate green, and one separate orange signal.

Prod. No. Product Label Tests* (Volume) Z-2275-50 ZytoLight SPEC ZNF384 Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

117 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 117 FE143-1-20 ZytoLight ® SPEC ETV6 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ETV6 Dual Color The SPEC ETV6 Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed for the Probe is a mixture of two direct labeled cation involving the 12p13.2 band, two detection of translocations involving the probes hybridizing to the 12p13.2 band. orange/green fusion signals are expected chromosomal region 12p13.2 harboring The green fluorochrome direct labeled representing two normal (non-rearranged) the ETV6 (ETS variant 6, a.k.a. TEL) gene. probe hybridizes proximal and the orange 12p13.2 loci. A signal pattern consisting ETV6 is a member of the ETS family of fluorochrome direct labeled probe hybrid- of one orange/green fusion signal, one transcription factors. More than 40 translo- izes distal to the ETV6 gene. orange signal, and a separate green cations with ETV6 involvement have been signal indicates one normal 12p13.2 reported in diverse types of hematological ETV6 locus and one 12p13.2 locus affected by and non-hematological malignancies. a translocation. The balanced chromosomal translocation t(12;21)(p13.2;q22.1), which leads to ETV6-RUNX1 fusion, represents the most frequent genetic rearrangement (19-27%) in initial childhood B-cell precursor (BCP) Ideogram of chromosome 12 acute lymphoblastic leukemia (ALL) and indicating the hybridization locations. has been associated with good prognosis. RH39884 RH53036 The ETV6-NTRK3 gene fusion resulting RH77972 BV208481 from the t(12;15)(p13.2;q25) transloca- 5’ 3’ tion was found to be characteristic for ETV6 mammary analogue secretory carcinoma ~415 kb ~580 kb (MASC) of the salivary glands. Since Tel 12p13.2 Cen SPEC ETV6 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated MASC morphologically mimics other neo- SPEC ETV6 Probe map (not to scale). by two orange/green fusion signals per nucleus. plasms, the detection of ETV6 rearrangements may be helpful for the differential diagnosis of MASC. In a subgroup of myeloproliferative disorders, the t(5;12)(q32;p13.2) translocation is a recurrent chromosome abnormality resulting in the fusion of ETV6 to the receptor tyrosine kinase PDGFRB. Patients carrying the t(5;12) translocation can be successfully treated with tyrosine kinase inhibitors. References MASC tissue section of the salivary glands with Bohlander SK (2005) Semin Cancer Biol 15: 162-74. De Braekeleer E, et al. (2012) Leuk Res 36: 945-61. translocation of the ETV6 gene as indicated by Peter A, et al. (2009) Eur J Haematol 83: 420-32. one non-rearranged orange/green fusion signal, Pinto A, et al. (2014) Mod Pathol 27: 30-7. one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2176-50 ZytoLight SPEC ETV6 Dual Color Break Apart Probe / 5 (50 μl) Z-2176-200 ZytoLight SPEC ETV6 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

118 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 118 FE085-1-20 ZytoLight ® SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC ETV6/RUNX1 Dual The SPEC ETV6/RUNX1 Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed for Fusion Probe is a mixture of an orange ange and two green signals are expected. the detection of the specific transloca- fluorochrome direct labeled ETV6 probe A reciprocal translocation involving two tion involving the chromosomal region spanning the known breakpoint region of breakpoints splits the two signals and 12p13.2 harboring the ETV6 (ETS variant the ETV6 gene and a green fluorochrome generates a fusion signal on each of the 6, a.k.a. TEL) gene and the chromosomal direct labeled RUNX1 probe covering the chromosomes involved. The chromosomal region 21q22.12 harboring the RUNX1 known breakpoint region of the RUNX1 regions which are not translocated are (runt related transcription factor 1, a.k.a. gene. indicated by the single orange and green AML1) gene. signal, respectively. The balanced chromosomal translocation ETV6 t(12;21)(p13.2;q22.1), which leads to ETV6/RUNX1 fusion, represents the most frequent genetic rearrangement in initial childhood B-cell precursor (BCP) acute lymphoblastic leukemia (ALL) (19-27%) and has been associated with good prognosis. The ETV6/RUNX1 fusion protein, comprising a putative repressor domain of ETV6, a member of the ETS family of transcription RUNX1

factors, fused to RUNX1, the DNA-binding Ideograms of chromosomes 12 (above) subunit of the RUNX1/CBF beta transcrip- and 21 (below) indicating the hybridization locations. tion factor complex, acts as a trans-domi- SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe RH45210 B402I14/T7 hybridized to normal interphase cells as indicated by nant repressor of RUNX1 regulated target 5’ 3’ two orange and two green signals in each nucleus. genes involved in hematopoiesis. ETV6 Three secondary aberrations in ETV6/ ~870 kb RUNX1 positive ALL have been found to Tel 12p13.2 Cen

negatively influence the clinical course: SPEC ETV6 Probe map (not to scale). deletion of the second non-translocated RH120345 D21S1894 ETV6 allele, gains of the RUNX1 gene, and 3’ 5’ duplication of the derivative chromosome 21. RUNX1 Detection of t(12;21) by Fluorescence in ~550 kb Cen 21q22.12 Tel situ Hybridization enables the simultaneous SPEC RUNX1 Probe map (not to scale). identification of the most common second- References ary changes and thus provides additional Fenrick R, et al. (1999) Mol Cell Biol 19: 6566-74. Martínez-Ramírez A, et al. (2001) Haematologica 86: 1245-53. Bone marrow tissue section with translocation information about the possible outcome of Morrow M, et al. (2007) Oncogene 26: 4404-14. affecting the ETV6/RUNX1 loci as indicated by Peter A, et al. (2009) Eur J Haematol 83: 420-32. one separate orange signal, one separate green the disease in patients with ALL. Shurtleff SA, et al. (1995) Leukemia 9: 1985-9. signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2157-50 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe / 5 (50 μl) Z-2157-200 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

119 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 119 FE072-1-20 ZytoLight ® SPEC KRAS/CEN 12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC KRAS/CEN 12 Dual The SPEC KRAS/CEN 12 Dual Color In a normal interphase nucleus, two or- Color Probe is designed for the detection Probe is a mixture of an orange fluoro- ange and two green signals are expected. of KRAS gene amplifications found e.g. in chrome direct labeled CEN 12 probe Nuclei with amplification of the KRAS lung cancer. specific for the alpha satellite centromeric gene locus 12p12.1 or aneuploidy of The KRAS (KRAS proto-oncogene, GTPase) region of chromosome 12 (D12Z3) and a chromosome 12 will show multiple copies gene located on chromosome 12p12.1 is green fluorochrome direct labeled SPEC of the green signal or large green signal a member of the RAS gene family compris- KRAS probe specific for the KRAS gene at clusters. ing HRAS, KRAS, and NRAS, all of which 12p12.1. encode a 21 kDa protein. The wildtype proteins play a pivotal role in cell prolif- KRAS eration, differentiation, and senescence. CEN 12 (D12Z3) Mutations of KRAS are frequently found in epithelial malignancies and lead to activation of the downstream mitogen-activated protein kinase (MAPK) resulting in unchecked cellular proliferation and tumor Ideogram of chromosome 12 progression. indicating the hybridization locations. Amplifications of KRAS and the im- RH120757 SHGC-130377 plications in tumorigenesis are not as 3’ 5’ well characterized as KRAS mutations. SPEC KRAS/CEN 12 Dual Color Probe hybridized KRAS However, recent studies using different to normal interphase cells as indicated by two ~925 kb orange and two green signals in each nucleus. methods found amplification of KRAS or Tel 12p12.1 Cen copy number gain of the 12p12.1 region SPEC KRAS Probe map (not to scale). including KRAS in various primary tumors, as e.g. in lung, colorectal, pancreatic, and gastric cancers. For non-small cell lung cancer (NSCLC) patients KRAS amplification as assessed by Fluorescence in situ Hybridization (FISH) was detected in about 15% of the tumors. Amplification of KRAS was found to be correlated with poor prognosis and may act synergistically with KRAS mutations to promote tumor progression. Lung cancer tissue section with amplification References of the KRAS gene (green). Little AS, et al. (2011) Sci Signal 4: er2. Mita H, et al. (2009) BMC Cancer 9: 198. Image kindly provided by Prof. Diebold, Lucerne, Switzerland. Sasaki H, et al. (2011) J Thorac Oncol 6: 15-20. Wagner PL, et al. (2011) Lung Cancer 74: 118-23.

Prod. No. Product Label Tests* (Volume) Z-2115-200 ZytoLight SPEC KRAS/CEN 12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

120 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 120 FE056-1-20 ZytoLight ® SPEC ERBB3/CEN 12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ERBB3/CEN 12 The SPEC ERBB3/CEN 12 Dual Color Using the SPEC ERBB3/CEN 12 Dual Dual Color Probe is designed for the de- Probe is a mixture of a green fluoro- Color Probe in a normal interphase nucle- tection of amplifications of the chromosom- chrome direct labeled CEN 12 probe us, two orange and two green signals are al regions harboring the ERBB3 gene. specific for the alpha satellite centromeric expected. In a cell with amplification of The ERBB3 (a.k.a. HER3) gene encodes a region of chromosome 12 (D12Z3) and the ERBB3 gene locus, multiple copies of transmembrane glycoprotein acting as a an orange fluorochrome direct labeled the orange signal or orange signal clusters cellular growth factor receptor. It belongs SPEC ERBB3 probe hybridizing distal and will be observed. to the epidermal growth factor receptor proximal to the human ERBB3 gene in the subgroup of the receptor tyrosine kinase chromosomal region 12q13.2-q13.3. superfamily also including ERBB1 (EGFR), ERBB2, which is known to be affected by gene amplifications in a number of malignant tumors, and ERBB4. ERBB3 Although EGFR and ERBB2 have been shown to represent good predictive markers and appropriate targets for therapeutic approaches, relatively less is known of Ideogram of chromosome 12 comparable significance for ERBB3 and indicating the hybridization locations. ERBB4. However, there is growing evi- BV210665 RH48368 dence that cooperation of all four mem- 5’3’ 5’ 3’ SPEC ERBB3/CEN 12 Dual Color Probe hybridized bers of the ERBB gene family contributes to to normal interphase cells as indicated by two orange CDK2 ERBB3 a more aggressive tumor phenotype and and two green signals in each nucleus. ~680 kb influences therapeutic response. Cen 12q13.2-q13.3 Tel Accordingly, it is assumed that the as- SPEC ERBB3 Probe map (not to scale). sessment of the combined amplification status of ERBB1 to ERBB4 may improve the diagnostic value significantly.

References Alimandi M, et al. (1995) Oncogene 10: 1813-21. Begnami MD, et al. (2011) J Clin Oncol 29: 3030-6. Berghoff AS, et al. (2014) Breast J 23: 637-43. Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Kraus MH, et al. (1989) Proc Natl Acad Sci U S A 86: 9193-7. Lédel F, et al. (2014) Eur J Cancer 50: 656-62. Sassen A, et al. (2008) Breast Cancer Res 10: R2. Sassen A, et al. (2009) Breast Cancer Res 11: R50. Zaczek A, et al. (2005) Histol Histopathol 20: 1005-15. Zimonjic DB, et al. (1995) Oncogene 10: 1235-7.

Prod. No. Product Label Tests* (Volume) Z-2056-200 ZytoLight SPEC ERBB3/CEN 12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

121 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 121 FE008-1-20 ZytoLight ® SPEC DDIT3 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC DDIT3 Dual Color The SPEC DDIT3 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 12q13.3-q14.1 translocations involving the chromosomal probes hybridizing to the 12q13.3-q14.1 band, two orange/green fusion signals region 12q13.3 harboring the DDIT3 band. The orange fluorochrome direct are expected representing two normal (DNA damage inducible transcript 3) labeled probe hybridizes proximal to the (non-rearranged) 12q13.3-q14.1 loci. A gene (a.k.a. CHOP, GADD153). DDIT3 gene and the green fluorochrome signal pattern consisting of one orange/ The DDIT3 gene encodes for a stress-in- direct labeled probe hybridizes distal to green fusion signal, one orange signal, duced dominant-negative inhibitor of that gene. and a separate green signal indicates the transcription factors C/EBP and LAP. one normal 12q13.3-q14.1 locus and DDIT3 is consistently rearranged in myx- one 12q13.3-q14.1 locus affected by a oid liposarcomas (MLS). The most frequent 12q13.3-q14.1 translocation. translocation involving the DDIT3 gene DDIT3 region is t(12;16)(q13.3;p11.2) and occurs in about 90% of patients with MLS. The rearrangement results in a fusion gene comprising the 5´ part of the FUS (fused in Ideogram of chromosome 12 sarcoma) gene, located in 16p11.2, and indicating the hybridization locations. the complete coding region of the DDIT3 gene. The FUS-DDIT3 fusion protein acts RH39898 D9S2056 RH52872 B746H16/T7 as an abnormal transcription factor and 3’5’ development of myxoid liposarcomas is DDIT3 thus regarded as a consequence of dereg- ~480 kb ~750 kb ulated FUS-DDIT3 target genes. Cen 12q13.3-q14.1 Tel Differential diagnosis of liposarcomas and SPEC DDIT3 Dual Color Break Apart Probe SPEC DDIT3 Probe map (not to scale). hybridized to normal interphase cells as indicated accurate classification, the latter being by two orange/green fusion signals per nucleus. especially important with regard to appropriate treatment and prognosis, are often problematic. Therefore, detection of DDIT3 rearrangements via FISH analysis is a valuable tool to confirm the histopathological diagnosis of myxoid liposacrcoma.

References Aman P, et al. (1992) Genes Chromosomes Cancer 5: 278-85. Andersson M, et al. (2010) BMC Cancer 10: 249-58. Germano G, et al. (2010) Cancer Res 70: 2235-44. Meis-Kindblom JM, et al. (2001) Virchows Arch 439: 141-51. Panagopoulos I, et al. (1994) Cancer Res 54: 6500-3. Ron D & Habener JF (1992) Genes Dev 6: 439-53.

Myxoid liposarcoma tissue section with translocation affecting the 12q13.3-q14.1 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2100-50 ZytoLight SPEC DDIT3 Dual Color Break Apart Probe / 5 (50 μl) Z-2100-200 ZytoLight SPEC DDIT3 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

122 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 122 FE033-1-20 ZytoLight ® SPEC CDK4/CEN 12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC CDK4/CEN 12 The SPEC CDK4/CEN 12 Dual Color In a normal interphase nucleus two orange Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- and two green signals are expected. Nu- detection of CDK4 gene amplifications. chrome direct labeled CEN 12 probe clei with amplification of the CDK4 gene The cyclin-dependent kinase 4 (CDK4) specific for the alpha satellite centromeric locus 12q13.3-q14.1, or polysomy of gene is located in the chromosomal region region of chromosome 12 (D12Z3) and a chromosome 12 will show multiple copies 12q14.1, ~10 Mb centromeric to the green fluorochrome direct labeled SPEC of the green signal or large green signal murine double minute (MDM2) gene and CDK4 probe specific for the chromosomal clusters. is frequently coamplified with MDM2 in region 12q13.3-q14.1 harboring the different malignancies. CDK4 gene. In a complex with cyclin D1 (CCND1), the CDK4 encoded serine/threonine kinase phosphorylates the retinoblastoma protein 1 (RB1) which in turn leads to the release of the EF2 transcription factor and subsequently to an upregulation of genes which are required for progression through the S-, G2-, and M-phases of the cell cycle. Ideogram of chromosome 12 Due to amplification of the respective indicating the hybridization locations. chromosomal region, CDK4 is overex- B258J5.GSS/T7 RH79979 pressed in many human tumors such as 3’ 5’ SPEC CDK4/CEN 12 Dual Color Probe hybridized soft tissue sarcomas, osteosarcomas (OS), to normal interphase cells as indicated by two CDK4 orange and two green signals in each nucleus. and gliomas. In glioblastomas, the lack of ~695 kb amplification of several genes like CDK4 Cen 12q13.3-q14.1 Tel was recognized to be associated with a SPEC CDK4 Probe map (not to scale). longer survival time. In OS, coamplification of MDM2 and CDK4, located in two discontinuous regions, occurs frequently in parosteal OS and less often in classical high-grade OS. Although MDM2/CDK4 coamplification is not restricted to atypical lipomatous tumors/well-differentiated liposarcomas (ALT/ WDLPS) and dedifferentiated liposarcomas (DDLPS), its detection is a strong cri- References Binh MB, et al. (2005) Am J Surg Pathol 29: 1340-7. Liposarcoma tissue section, CDK4 signal terion for distinguishing these tumor types Fischer U, et al. (2010) Int J Cancer 126: 2594-602. cluster (green), CEN 12 (orange). Lopes MA, et al. (2001) Oral Oncol 37: 566-71. from other undifferentiated sarcomas and Mejia-Guerrero S, et al. (2010) Genes Chromosomes Cancer 49: 518-25. Sirvent N, et al. (2007) Am J Surg Pathol 31: 1476-89. even from carcinomas and lymphomas. Wunder JS, et al. (1999) Oncogene 18: 783-8.

Prod. No. Product Label Tests* (Volume) Z-2103-50 ZytoLight SPEC CDK4/CEN 12 Dual Color Probe / 5 (50 μl) Z-2103-200 ZytoLight SPEC CDK4/CEN 12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

123 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 123 FE034-1-20 ZytoLight ® SPEC MDM2/CEN 12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC MDM2/CEN 12 The SPEC MDM2/CEN 12 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- ange and two green signals are expected. detection of MDM2 gene amplifications chrome direct labeled CEN 12 probe In a cell with amplification of the MDM2 found in more than 10% of human tumors. specific for the alpha satellite centromeric gene locus, multiple copies of the green The MDM2 (MDM2 proto-oncogene) gene region of chromosome 12 (D12Z3) and a signal or green signal clusters will be is located in the chromosomal region green fluorochrome direct labeled SPEC observed. 12q15 and encodes for an E3 ubiquitin MDM2 probe specific for the MDM2 gene ligase which acts as a major negative at 12q15. regulator of the tumor suppressor p53. Due to amplification of the respective chromosomal region, MDM2 is overexpressed in many human tumors such as soft tissue sarcomas, osteosarcomas, gliomas, NSCLC, gastric and breast carcinomas. Well-differentiated liposarcomas (WDLPS), the most common soft tissue tumors in Ideogram of chromosome 12 adults, are characterized by the amplifica- indicating the hybridization locations. tion of 12q-derived chromosomal material, SHGC-105915 D12S1497 harboring the MDM2 oncogene while 5’ 3’ Normal interphase cells, MDM2 (green), lipomas show balanced translocations in- CEN 12 (orange). MDM2 volving 12q13-15. Accordingly, detection ~495 kb of the 12q14-15 amplification is regard- Cen 12q15 Tel ed as a valuable tool for the differential SPEC MDM2 Probe map (not to scale). diagnosis between WDLPS and lipomas. Furthermore, detection of MDM2 amplification might have prognostic relevance in gastrointestinal stromal tumors (GIST), the most common primary mesenchymal tumor of the gastrointestinal tract.

References Brisson M, et al. (2013) Skeletal Radiol 42: 635-47. Duhamel LA, et al. (2012) Histopathology 60: 357-9. Flanagan AM, et al. (2010) Skeletal Radiol 39: 213-24. Kashima T, et al. (2012) Mod Pathol 25: 1384-96. Kikuchi K, et al. (2013) Sarcoma 2013: 520858. Korcheva VB, et al. (2011) Appl Immunhistochem Mol Morphol 19: 119-25. Liposarcoma tissue section with amplification of Larousserie F, et al. (2013) Eur J Radiol 82: 2149-53. the MDM2 gene (green), CEN 12 (orange). Lokka S, et al. (2014) BMC Clin Pathol 14: 36. Luan SL, et al. (2010) J Pathol 222: 166-79. Momand J, et al. (1992) Cell 69: 1237-45. Oliner JD, et al. (1992) Nature 358: 80-3. Pedeutour F, et al. (1994) Genes Chromosomes Cancer 10: 85-94. Pedeutour F, et al. (2004) Bull Cancer 91: 317-23. Pedeutour F, et al. (2012) Virchows Arch 461: 67-78. Poaty H, et al. (2012) PLoS One 7: e29426. Toledo F & Wahl GM (2006) Nat Rev Cancer 6: 909-23. Tornillo L, et al. (2005) Lab Invest 85: 921-31. Vassilev LT (2007) Trends Mol Med 13: 23-31.

Prod. No. Product Label Tests* (Volume) Z-2013-50 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe / 5 (50 μl) Z-2013-200 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

124 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 124 FE003-1-20 ZytoLight ® SPEC FOXO1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC FOXO1 Dual Color The SPEC FOXO1 Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed for the Probe is a mixture of two direct labeled cation involving the 13q14.11 band, two detection of specific translocations involv- probes hybridizing to the 13q14.11 orange/green fusion signals are expected ing the chromosomal region 13q14.11 band. The orange fluorochrome direct representing two normal (non-rearranged) harboring the FOXO1 (forkhead box O1, labeled probe hybridizes distal, the green 13q14.11 loci. A signal pattern consisting a.k.a. FKHR) gene characteristic for alveo- fluorochrome direct labeled probe hybrid- of one orange/green fusion signal, one lar rhabdomyosarcoma. izes proximal to the breakpoint region of orange signal, and a separate green sig- Among solid tumors of the childhood, the FOXO1 gene. nal indicates one normal 13q14.11 locus rhabdomyosarcoma (RMS) is the most and one 13q14.11 locus affected by a common soft tissue sarcoma. RMS are translocation. classified in two main categories: embry- FOXO1 onal rhabdomyosarcoma (ERMS) and alveolar rhabdomyosarcoma (ARMS). The alveolar histology is associated with a poorer prognosis. ARMS is characterized Ideogram of chromosome 13 by two tumor-specific reciprocal translo- indicating the hybridization locations. cations t(2;13)(q36;q14.1) and t(1;13) RH53132 D13S320 (p36.1;q14.1) detectable in more than RH45178 RH37996 80% of all ARMS. These translocations 3’ 5’

fuse the FOXO1 locus on 13q14.11 FOXO1

to either PAX3 on chromosome 2 or to ~845 kb ~810 kb PAX7 on chromosome 1. The resulting Cen 13q14.11 Tel fusion transcripts encode for the chimeric SPEC FOXO1 Probe map (not to scale). SPEC FOXO1 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated proteins PAX3-FOXO1 and PAX7-FOXO1 by two orange/green fusion signals per nucleus. that combine transcriptional domains from the corresponding wild-type proteins and thereby acquire oncogenic activity. The translocations and their fusion genes represent highly specific genetic markers useful in the diagnosis of ARMS.

References Dal Cin P, et al. (1991) Cancer Genet Cytogenet 55: 191-5. Douglass EC, et al. (1991) Genes Chromosomes Cancer 3: 480-2. Gunawan B, et al. (1999) Pathol Oncol Res 5: 211-3. Seidal T, et al. (1982) Acta Pathol Microbiol Immunol Scand A 90: 345-54. Sorensen PH, et al. (2002) J Clin Oncol 20: 2672-9.

Rhabdomyosarcoma tissue section with translocation affecting the 13q14.11 locus harboring FOXO1 as indicated by one orange/green fusion signal (non-rearranged), one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2139-50 ZytoLight SPEC FOXO1 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

125 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 125 FE059-1-20 ZytoLight ® SPEC FOXO1/PAX3 Dual Color Single Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC FOXO1/PAX3 The SPEC FOXO1/PAX3 Dual Color Sin- In an interphase nucleus lacking the Dual Color Single Fusion Probe is de- gle Fusion Probe is a mixture of two direct t(2;13), two orange and two green signals signed to detect the translocation t(2;13) labeled probes hybridizing to the 2q36.1 are expected. (q36;q14.1) in alveolar rhabdomyosarco- and 13q14.11 band. The green fluoro- In a cell harboring the t(2;13), one mas. chrome direct labeled probe hybridizes orange signal, one green signal, and Among solid tumors of the childhood, distal to the PAX3 gene at 2q36.1, the one orange/green fusion signal will be rhabdomyosarcoma is the most common orange fluorochrome direct labeled probe observed. soft tissue sarcoma. Rhabdomyosarcomas hybridizes proximal to the FOXO1 gene are classified in two main categories: em- at 13q14.11. bryonal and alveolar rhabdomyosarcoma. The alveolar histology is associated with a poorer prognosis. FOXO1 Alveolar rhabdomyosarcoma is characterized by two tumor-specific translocations, i.e., t(2;13)(q36;q14.1) and t(1;13) (p36.1;q14.1) which are detectable in most cases of alveolar rhabdomyosarcomas. The translocations and their fusion genes

represent highly specific genetic markers SPEC FOXO1/PAX3 Dual Color Single Fusion useful in the diagnosis of alveolar rhabdo- Probe hybridized to abnormal nuclei harboring a t(2;13)(q35;q14) as indicated by one orange, one myosarcomas. green, and one orange/green fusion signal. Correlations between the type of translocation and clinical features as e.g. longer disease-free survival have been identified.

References Dal Cin P, et al. (1991) Cancer Genet Cytogenet 55: 191-5. PAX3 Douglass EC, et al. (1991) Genes Chromosomes Cancer 3: 480-2. Gunawan B, et al. (1999) Pathol Oncol Res 5: 211-3. Ideograms of chromosomes 13 (above) and 2 Rekhi B, et al. (2014) Pathol Res Pract 210: 328-33. Seidal T, et al. (1982) Acta Pathol Microbiol Immunol Scand A: 345-54. (below) indicating the hybridization locations.

RH98592 D13S1162 3’ 5’

FOXO1

~315 kb Cen 13q14.11 Tel

SPEC FOXO1 Probe map (not to scale).

RH33630 RH47422 3’ 5’

PAX3

~345 kb Cen 2q36.1 Tel

SPEC PAX3 Probe map (not to scale). Prod. No. Product Label Tests* (Volume) Z-2018-50 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe / 5 (50 μl) Z-2018-200 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

126 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 126 FE007-1-20 ZytoLight ® SPEC FOXO1/PAX3 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC FOXO1/PAX3 The SPEC FOXO1/PAX3 TriCheck™ Probe In an interphase nucleus without PAX3- TriCheck™ Probe is designed to detect trans- is a mixture of three direct labeled probes FOXO1 rearrangement, two green/ locations involving the chromosomal region hybridizing to the 13q14.11 and 2q36.1 orange fusion signals and two blue signals 13q14.11 harboring the FOXO1 (fork- bands. The orange fluorochrome direct la- are expected. head box O1, a.k.a. FKHR) gene and the beled probe hybridizes proximal and the A PAX3-FOXO1 fusion is indicated by one chromosomal region 2q36.1 harboring the green fluorochrome direct labeled probe separate orange signal co-localizing with PAX3 (paired box 3, a.k.a. HUP2) gene. hybridizes distal to the FOXO1 breakpoint one blue signal and one separate green Among solid tumors of the childhood, rhab- region at 13q14.11. The blue fluoro- signal. domyosarcoma is the most common soft chrome direct labeled probe hybridizes A FOXO1 translocation without involve- tissue sarcoma. Rhabdomyosarcomas are distal to the PAX3 gene at 2q36.1. ment of PAX3 is indicated by the split of classified in two main categories: embryo- one green/orange fusion signal without nal and alveolar rhabdomyosarcoma. co-localization of the separated orange Generally, the alveolar histology is associat- FOXO1 signal with one blue signal. ed with a poorer prognosis. Alveolar rhabdomyosarcoma (ARMS) is characterized by two tumor-specific translocations, i.e., t(2;13)(q36;q14.1) and t(1;13)(p36.1;q14.1) which are detectable in most cases of ARMS. The translocations involve the FOXO1 gene and either PAX7 on chromosome 1p36.13 or PAX3 on chromosome 2q36.1. PAX7-FOXO1 is less common but is associated with a better prognosis than PAX3-FOXO1 fusion. The translocations and their fusion genes represent highly specific genetic markers SPEC FOXO1/PAX3 TriCheck™ Probe hybridized to useful in the diagnosis and prognosis of normal interphase cells as indicated by two orange/ green fusion signals and two blue signals per nucleus. ARMS. References PAX3 Dal Cin P, et al. (1991) Cancer Genet Cytogenet 55: 191-5. Douglass EC, et al. (1991) Genes Chromosomes Cancer 3: 480-2. Ideograms of chromosomes 13 (above) and Gunawan B, et al. (1999) Pathol Oncol Res 5: 211-3. 2 (below) indicating the hybridization locations. Jain S, et al. (2010) Int J Clin Exp Pathol 3: 416-28. Seidal T, et al. (1982) Acta Pathol Microbiol Immunol Scand A 90: 345-54.

D13S1162 D13S320 RH33630 BV208633 SHGC-110059 RH37996 3’ 5’ 3’ 5’

PAX3 FOXO1 ~740 kb ~555 kb ~810 kb Cen 2q36.1 Tel Cen 13q14.11 Tel ARMS tissue section with PAX3-FOXO1 fusion SPEC FOXO1 Probe map (not to scale). SPEC PAX3 Probe map (not to scale). as indicated by orange/blue fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2185-50 ZytoLight SPEC FOXO1/PAX3 TriCheck Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

127 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 127 FE114-1-20 ZytoLight ® SPEC FOXO1/PAX7 Dual Color Single Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC FOXO1/PAX7 The SPEC FOXO1/PAX7 Dual Color In an interphase nucleus lacking the Dual Color Single Fusion Probe is de- Single Fusion Probe is a mixture of two t(1;13), two orange and two green signals signed to detect the translocation t(1;13) direct labeled probes hybridizing to the are expected. (p36.1;q14.1) in alveolar rhabdomyosar- 1p36.13 and 13q14.11 band. The green In a cell harboring the t(1;13), one comas. fluorochrome direct labeled probe hybrid- orange signal, one green signal, and Among solid tumors of the childhood, izes distal to the PAX7 gene at 1p36.13, one orange/green fusion signal will be rhabdomyosarcoma is the most common the orange fluorochrome direct labeled observed. soft tissue sarcoma. Rhabdomyosarcomas probe hybridizes proximal to the FOXO1 are classified in two main categories: em- gene at 13q14.11. bryonal and alveolar rhabdomyosarcoma. The alveolar histology is associated with a poorer prognosis. FOXO1 Alveolar rhabdomyosarcoma is characterized by two tumor-specific translocations, i.e., t(2;13)(q36;q14.1) and t(1;13) (p36.1;q14.1) which are detectable in PAX7 most cases of alveolar rhabdomyosarcomas. The translocations and their fusion genes represent highly specific genetic markers SPEC FOXO1/PAX7 Dual Color Single Fusion Probe hybridized to normal interphase cells as indicated useful in the diagnosis of alveolar rhabdo- by two orange and two green signals. myosarcomas. Correlations between the type of translocation and clinical features as e.g. longer disease-free survival have been identified.

References Dal Cin P, et al. (1991) Cancer Genet Cytogenet 55: 191-5. Douglass EC, et al. (1991) Genes Chromosomes Cancer 3: 480-2. Gunawan B, et al. (1999) Pathol Oncol Res 5: 211-3. Ideograms of chromosomes 13 (above) and 1 Rekhi B, et al. (2014) Pathol Res Pract 210: 328-33. (below) indicating the hybridization locations. Seidal T, et al. (1982) Acta Pathol Microbiol Immunol Scand A: 345-54. RH98592 D13S1162 3’ 5’

FOXO1

~315 kb Cen 13q14.11 Tel

SPEC FOXO1 Probe map (not to scale).

SHGC-142982 STSG27587 5’ 3’

PAX7

~815 kb Tel 1p36.13 Cen

SPEC PAX7 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2019-50 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe / 5 (50 μl) Z-2019-200 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

128 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 128 FE131-1-20 ZytoLight ® SPEC RB1/13q12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC RB1/13q12 Dual The SPEC RB1/13q12 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome ange and two green signals are expected. of deletions affecting the RB1 gene. direct labeled SPEC RB1 probe specific for In a cell with deletions affecting the RB1 The RB1 (RB transcriptional corepressor 1, the RB1 gene in the chromosomal region gene locus, one or no copy of the orange a.k.a. pRb) gene is located on 13q14.2 13q14.2 and a green fluorochrome direct signal will be observed. and encodes a protein which acts as a labeled SPEC 13q12 probe specific for tumor suppressor playing a crucial role in the chromosomal region 13q12.11. The cell cycle regulation and genome stability. SPEC 13q12 Probe is designed to hybrid- Deletions of RB1 are frequently found in ize in close proximity of centromere 13 at retinoblastoma. 13q12.11. Since chromosomes 13 and However, either monoallelic or biallelic de- 21 share the same repetitive sequences, letions of RB1 are also common in a wide they cannot be differentiated by probes variety of solid tumors and hematologic detecting centromere specific repeats. malignancies such as multiple myeloma (MM) and chronic lymphocytic leukemia 13q12 (CLL). RB1 While 13q14 deletions exclusive of RB1 confer a more favorable prognosis in CLL SPEC RB1/13q12 Dual Color Probe hybridized patients, 13q14 deletions that encompass to normal interphase cells as indicated by two orange and two green signals in each nucleus. the RB1 locus (present in approx. 20% of Ideogram of chromosome 13 all CLL cases) are associated with short- indicating the hybridization locations. ened survival. SHGC-16661 SHGC-106473 Hence, Fluorescence in situ Hybridization 5’ 3’ is a valuable tool for the detection of RB1 RB1 gene deletions and can be used in com- ~420 kb bination with further biological markers, Cen 13q14.2 Tel

morphology and clinical information for SPEC RB1 Probe map (not to scale). the prediction of disease progression and overall survival. D13S1574 D13S1558

References Dal Bo M, et al. (2011) Genes Chromosomes Cancer 50: 633-43. Dao DD, et al. (1994) Leukemia 8: 1280-4. Di Fiore R, et al. (2013) J Cell Physiol 228: 1676-87. ~690 kb Juge-Morineau N, et al. (1997) Leuk Lymphoma 24: 229-37. Orlandi EM, et al. (2013) Hematol Oncol 31: 136-42. Cen 13q12.11 Tel SPEC RB1/13q12 Dual Color Probe hybridized Ouillette P, et al. (2011) Clin Cancer Res 17: 6778-90. to benign spindle cell lipoma tissue section with SPEC 13q12 Probe map (not to scale). deletion of the RB1 gene as indicated by one orange signal and two green signals in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2165-50 ZytoLight SPEC RB1/13q12 Dual Color Probe / 5 (50 μl) Z-2165-200 ZytoLight SPEC RB1/13q12 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

129 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 129 FE076-1-20 ZytoLight ® SPEC IGH Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC IGH Dual Color The SPEC IGH Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cation involving the 14q32.33 band two translocations involving the chromosomal probes hybridizing to the 14q32.33 orange/green fusion signals are expected region 14q32.33 harboring the IGH band. The orange fluorochrome direct la- representing two normal (non-rearranged) gene. beled probe hybridizes proximal, and the 14q32.33 loci. A signal pattern consisting Rearrangements involving the IGH (im- green fluorochrome direct labeled probe of one orange/green fusion signal, one munoglobulin heavy locus, a.k.a. IGH@) hybridizes distal to the constant regions of orange signal, and a separate green sig- gene are considered to be cytogenetic the IGH locus. nal indicates one normal 14q32.33 locus hallmarks for non-Hodgkin lymphoma and one 14q32.33 locus affected by a (NHL). NHLs represent 50% of all hemato- translocation. logical malignancies. IGH gene rearrangements have been identified in about 50% of NHLs and are associated with specific subtypes of NHLs. Translocation t(11;14)(q13.3;q32.3) can Ideogram of chromosome 14 be found in about in 95% of mantle cell indicating the hybridization locations. lymphoma (MCL), t(14;18)(q32.3;q21.3) H14A434 RH69050 in 80% of follicular lymphoma (FL), t(3;14) H14A1911 RH70844 (q27;q32.3) in diffuse large B-cell lympho- 3’ 5’ 3’ 5’

ma (DLBCL), and t(8;14)(q24.21;q32.3) JAG2 IGH

in Burkitt lymphoma. In all of these trans- ~615 kb ~580 kb locations an oncogene located near the Cen 14q32.33 Tel SPEC IGH Dual Color Break Apart Probe breakpoint of the translocation partner is hybridized to normal interphase cells as indicated SPEC IGH Probe map (not to scale). by two orange/green fusion signals per nucleus. activated by juxtaposing to IGH regulatory sequences. Rearrangements involving 14q32.33 have unique biological characteristics and correlate with clinical, morphological, and immunophenotypic features. Fluorescence in situ Hybridization is a helpful tool for the diagnosis, selecting treatment, and giving prognostic information.

References Bernicot I, et al. (2007) Cytogenet Genome Res 118: 345-52. Hehne S, et al. (2012) Pathol Res Pract 208: 510-7. Lu S, et al. (2004) Cancer Genet and Cytogenet 152: 141-5. Nishida K, et al. (1997) Blood 90: 526-34. Burkitt lymphoma tissue section with translocation Quintero-Rivera F, et al. (2009) Cancer Genet and Cytogenet 190: 33-9. affecting the 14q32.33 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2110-50 ZytoLight SPEC IGH Dual Color Break Apart Probe / 5 (50 μl) Z-2110-200 ZytoLight SPEC IGH Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

130 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 130 FE038-1-20 ZytoLight ® SPEC NUTM1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NUTM1 Dual Color The SPEC NUTM1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 15q14 band, two translocations involving the chromosomal probes hybridizing to the 15q14 band. orange/green fusion signals are expected region 15q14 harboring the NUTM1 (NUT The green fluorochrome direct labeled representing two normal (non-rearranged) midline carcinoma family member 1, a.k.a. probe hybridizes proximal and the orange 15q14 loci. A signal pattern consisting of NUT) gene. fluorochrome direct labeled probe hybrid- one orange/green fusion signal, one or- NUT midline carcinoma (NMC) is a rare izes distal to the NUTM1 gene. ange signal, and a separate green signal and aggressive form of squamous cell indicates one normal 15q14 locus and carcinoma that arises mainly in the head, one 15q14 locus affected by a transloca- neck, or mediastinum. NMC is genetically NUTM1 tion. defined by the presence of chromosomal rearrangements involving the NUTM1 gene. Two-thirds of NMCs have t(15;19) (q14;p13.1) fusing the NUTM1 gene to the Ideogram of chromosome 15 BRD4 gene. Less commonly, NMC harbors indicating the hybridization locations.

a NUTM1-variant fusion gene involving RH46282 SHGC-107223 BRD3 or still-uncharacterized genes. NMCs RH123440 D15S877E 5’ 3’ may be indistinguishable from more common squamous cell carcinomas and are NUTM1 thus an underdiagnosed entity. Therefore, ~590 kb ~455 kb Cen 15q14 Tel the diagnosis of NMC depends on the con- firmation of NUTM1 rearrangement. SPEC NUTM1 Probe map (not to scale). BRD3 and BRD4 belong to the bromo and SPEC NUTM1 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated extra terminal (BET) family of bromodomain by two orange/green fusion signals per nucleus. proteins. BRD-NUTM1 chimeric oncopro- teins repress squamous differentiation, possibly by sequestering histone acetyltransferase activity. Accordingly, histone deacetylase inhibitors or BET inhibitors were shown to reverse the effects of BRD-NUTM1 fusion proteins by inducing terminal differentiation of NMC cells in vitro and in xenograft models. Hence, detection of NUTM1 rearrangements by FISH represents a useful tool in the differential diagnosis of NMC and may be of therapeutic significance. NMC tissue section with translocation of the References NUTM1 gene as indicated by one French CA (2012) Annu Rev Pathol 7: 247-65. non-rearranged orange/green fusion signal, Kubonishi I, et al. (1991) Cancer Res 51: 3327-8. Müller S & Knapp S (2014) Med Chem Commun 5: 288-96. one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2208-200 ZytoLight SPEC NUTM1 Dual Color Break Apart Probe •/• 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

131 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 131 FE120-1-20 ZytoLight ® SPEC PML/RARA Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC PML/RARA Dual The SPEC PML/RARA Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed to de- Fusion Probe is a mixture of an orange ange and two green signals are expected. tect the translocation t(15;17)(q24;q21.2) fluorochrome direct labeled PML probe A reciprocal PML/RARA translocation affecting the PML gene in the chromosom- spanning the known PML breakpoints, and leads to two orange/green fusion signals al region 15q24.1 and the RARA locus in a green fluorochrome direct labeled RARA indicating both rearranged chromosomes. 17q21.2. probe spanning the known breakpoints of Additionally, the non-rearranged chromo- Translocations involving the PML (promye- RARA. This probe is approved to be used somes are indicated by one orange signal locytic leukemia, a.k.a. MYL) gene and with a hybridization time of 2 hours on and a separate green signal, respectively. the RARA (retinoic acid receptor alpha, cytological specimens. a.k.a. RARα) gene are considered to be characteristic for acute promyelocytic leukemia (APL), a subtype of acute myeloid leukemia. Various fusion partners of RARA have PML been identified, however, in 95% of all APL cases, rearrangements involving the PML gene are detectable. This translocation t(15;17)(q24;q21) leads to a gene RARA fusion of the PML and the RARA gene. The fusion is supposed to play a fundamental role in induction, development, and pro- SPEC PML/RARA Dual Color Dual Fusion Probe gression of APL. Ideograms of chromosomes 15 (above) and hybridized to normal interphase cells as indicated by 17 (below) indicating the hybridization locations. two orange and two green signals in each nucleus. Since the PML/RARA fusion accounts for the response of these neoplasms to SHGC-149501 AFMB292XH5 5’ 3’ all-trans retinoic acid (ATRA) therapy and other conventional chemotherapy it is im- PML portant to accurately distinguish between ~790 kb Cen 15q24.1 Tel t(15;17) translocations and translocations involving other partners of RARA. SPEC PML Probe map (not to scale).

References D17S1215 RH25598 Abe S, et al. (2008) Cancer Genet and Cytogenet 184: 44-7. 5’ 3’ Brockmann SR, et al. (2003) Cancer Genet and Cytogenet 145: 144-51. Reiter A, et al. (2004) Acta Hematol 112: 55-67. Sanz MA, et al. (2009) Blood 113: 1875-91. RARA

~865 kb Cen 17q12-q21.2 Tel

SPEC RARA Probe map (not to scale). Bone marrow biopsy section with translocation affecting the PML/RARA loci as indicated by one separate orange signal, one separate green signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2113-50 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe / 5 (50 μl) Z-2113-200 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

132 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 132 FE049-1-20 ZytoLight ® SPEC NTRK3 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC NTRK3 Dual Color The SPEC NTRK3 Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving translocations involving the chromosomal probes hybridizing to the 15q25.3-q26.1 the 15q25.3-q26.1 band, two orange/ region 15q25.3 harboring the NTRK3 (neu- band. The orange fluorochrome direct green fusion signals are expected rotrophic receptor tyrosine kinase 3, a.k.a. labeled probe hybridizes proximal to the representing two normal (non-rear- TRKC) gene. NTRK3 breakpoint region at 15q25.3, the ranged) 15q25.3-q26.1 loci. A signal NTRK3 is a receptor tyrosine kinase (TK) for green fluorochrome direct labeled probe pattern consisting of one orange/green neurotrophin 3 (NT3) and plays a key role in hybridizes distal to the NTRK3 breakpoint fusion signal, one orange signal, and central and peripheral nervous system devel- region at 15q25.3-q26.1. a separate green signal indicates one opment as well as in cell survival. Transloca- normal 15q25.3-q26.1 locus and one tions affecting the NTRK3 gene have been 15q25.3-q26.1 locus affected by a trans- reported in several cancer types, including location. glioblastomas, Philadelphia chromosome-like acute lymphoblastic leukemia, congenital fibrosarcomas, cellular mesoblastic nephro- NTRK3 mas, acute myeloid leukemia, radiation-as- Ideogram of chromosome 15 sociated thyroid cancer, secretory breast indicating the hybridization locations. carcinoma, and mammary analog secretory carcinoma of the salivary gland. The most D15S99 RH122647 SHGC-141093 RH123468 frequent rearrangement involving the NTRK3 3’ 5’ gene is the t(12;15)(p13;q25) which results in a fusion between the 5’ part of the ETV6 NTRK3 gene and the 3’ part of the NTRK3 gene. ~395 kb ~740 kb Cen 15q25.3-q26.1 Tel SPEC NTRK3 Dual Color Break Apart Probe This fusion gene encodes a hybrid protein hybridized to normal interphase cells as indicated comprising the TK domain of NTRK3 and the SPEC NTRK3 Probe map (not to scale). by two orange/green fusion signals per nucleus. dimerization domain of ETV6 which leads to a ligand-independent TK activity. Currently, there are several ongoing clinical trials involving drugs with known inhibitory activity of NTRK-related kinases. Entrectinib and LOXO-101 represent two of these TRK inhibitors which have shown promising activity and good tolerability in patients with References Amatu A, et al. (2016) ESMO Open 1: e000023. advanced solid tumors and NSCLC harbor- Arce C, et al. (2005) World J Surg Oncol 3: 35. Knezevich SR, et al. (1998) Nat Genet 18: 184-7. ing NTRK1, 2, and 3 rearrangements. Leeman-Neill RJ, et al. (2014) Cancer 120: 799-807. Nagasubramanian R, et al. (2016) Pediatr Blood Cancer 63: 1468-70. Hence, detection of NTRK3 translocations by Raez LE & Rolfo C (2016) Lung Cancer Manag 5: 1-4. Secretory breast carcinoma tissue section Roberts KG, et al. (2014) N Engl J Med 371: 1005-15. with translocation affecting the 15q25.3-q26.1 Fluorescence in situ Hybridization (FISH) may Skálová A, et al. (2010) Am J Surg Pathol 34: 599-608. Tognon C, et al. (2002) Cancer Cell 2: 367-76. locus as indicated by one non-rearranged be of diagnostic and therapeutic relevance. Wu G, et al. (2014) Nat Genet 46: 444-50. orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2206-50 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe •/• 5 (50 μl) Z-2206-200 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe •/• 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

133 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 133 FE118-1-20 ZytoLight ® SPEC CREBBP Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CREBBP Dual Color The SPEC CREBBP Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed for the Probe is a mixture of two direct labeled cation involving the 16p13.3 band, two detection of translocations involving the probes hybridizing to the 16p13.3 band. orange/green fusion signals are expected chromosomal region 16p13.3 harboring The green fluorochrome direct labeled representing two normal (non-rearranged) the CREBBP (CREB binding protein, a.k.a. probe hybridizes proximal and the orange 16p13.3 loci. A signal pattern consisting CBP, RTS) gene. The CREBBP protein fluorochrome direct labeled probe hybrid- of one orange/green fusion signal, one regulates transcription by means of histone izes distal to the CREBBP gene. orange signal, and a separate green acetyltransferase activity and by binding signal indicates one normal 16p13.3 CREBBP to several proteins with key cell cycle locus and one 16p13.3 locus affected by functions, such as p53 and NFκB. a translocation. Rearrangements of the CREBBP gene have been observed in several hematologic malignancies. Three different fusion partners have been described so far. Ideogram of chromosome 16 indicating the hybridization locations. KMT2A (a.k.a. MLL) is fused to CREBBP in therapy-related acute myeloid (AML) or RH68351 RH15630 RH99617 RH91706 lymphoid leukemia (ALL) and myelo- 3’ 5’ dysplastic syndrome (MDS) with t(11;16) (q23.3;p13.3). The translocation t(10;16) CREBBP (q22.2;p13.3) was reported in some AML ~475 kb ~545 kb Tel 16p13.3 Cen cases and fuses KAT6B (a.k.a. MORF) to CREBBP. CREBBP is also rearranged SPEC CREBBP Probe map (not to scale).

with KAT6A (a.k.a. MOZ) in de novo SPEC CREBBP Dual Color Break Apart Probe and therapy-related AML with t(8;16) hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. (p11.2;p13.3) after treatment with topoisomerase II inhibitors. This rearrangement is associated with an infrequent but well-defined type of AML that has characteristic morphocytochemical features. The prognosis is usually extremely poor, with a median survival of two months. The KAT6A/CREBBP AML tends to develop within two years of adjuvant chemotherapy, especially in former breast cancer patients. Thus, FISH analysis for the detection of CREBBP translocation may serve as a di- References Borrow J, et al. (1996) Nat Genet 14: 33-41. agnostic tool to identify cases with hema- Camós M, et al. (2006) Cancer Res 66: 6947-54. Gupta A, et al. (2014) Case Rep Oncol Med 2014: 361748. tologic malignancies with an aggressive Rozman M, et al. (2004) Genes Chromosomes Cancer 40: 140-5. Taki T, et al. (1997) Blood 89: 3945-50. presentation. Vizmanos JL, et al. (2003) Genes Chromosomes Cancer 36: 402-5.

Prod. No. Product Label Tests* (Volume) Z-2267-50 ZytoLight SPEC CREBBP Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

134 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 134 FE137-1-20 ZytoLight ® SPEC FUS Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC FUS Dual Color The SPEC FUS Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cation involving the 16p11.2 band, two translocations involving the chromosomal probes hybridizing to the 16p11.2 band. orange/green fusion signals are expected region 16p11.2 harboring the FUS (FUS The orange fluorochrome direct labeled representing two normal (non-rearranged) RNA binding protein) gene (a.k.a. TLS, probe hybridizes proximal to the FUS 16p11.2 loci. A signal pattern consisting FUS/TLS, hnRNP P2). gene, the green fluorochrome direct labe- of one orange/green fusion signal, one The FUS gene encodes an RNA-binding led probe hybridizes distal to that gene. orange signal, and a separate green protein, the C-terminal end of which is signal indicates one normal 16p11.2 involved in protein and RNA binding and locus and one 16p11.2 locus affected by which appears to be involved in transcrip- FUS a 16p11.2 translocation. tional activation with its N-terminal end. It shares distinct characteristics with EWSR1 and TAF15 which together with FUS are Ideogram of chromosome 16 frequently referred to as the FET family of indicating the hybridization locations. proteins. FUS gene rearrangements have been RH28513 SHGC-106097 RH33714 CHLC.GCT13F06 shown to be involved in both solid tumors 5’ 3’ and leukemias fusing the N-terminal end FUS of FUS to various fusion partners. The most ~445 kb ~715 kb frequent translocation involving the FUS Tel 16p11.2 Cen gene region is t(12;16)(q13.3;p11.2). SPEC FUS Probe map (not to scale). Occurring in over 90% of myxoid liposarcomas, the FUS-DDIT3 fusion protein is SPEC FUS Dual Color Break Apart Probe hybridized regarded as being consequential for the to normal interphase cells as indicated by two orange/green fusion signals per nucleus. development of myxoid liposarcomas by acting as an abnormal transcription factor and thus deregulating FUS-DDIT3 target genes. Differential diagnosis of liposarcomas and accurate classification, the latter being especially important with regard to appropriate treatment and prognosis, are often problematic. Therefore, detection of FUS rearrangements via in situ Hybridiza- tion analysis is a valuable tool to confirm the histopathological diagnosis of myxoid liposarcoma. Myxoid liposarcoma tissue section with translocation References affecting the 16p11.2 locus as indicated by one Andersson M, et al. (2010) BMC Cancer 10: 249-58. non-rearranged orange/green fusion signal, one Antonescu C, et al. (2000) J Mol Diagn 2: 132-8. orange signal, and one separate green signal. Germano G, et al. (2010) Cancer Res 70: 2235-44. Kuroda M, et al. (1995) Am J Pathol 147: 1221-7. Meis-Kindblom JM, et al. (2001) Virchows Arch 439: 141-51. Panagopoulos I, et al. (1994) Cancer Res 54: 6500-3. Panagopoulos I, et al. (1997) Oncogene 15: 1357-62.

Prod. No. Product Label Tests* (Volume) Z-2130-50 ZytoLight SPEC FUS Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

135 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 135 FE053-1-20 ZytoLight ® SPEC CBFB Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CBFB Dual Color The SPEC CBFB Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking a translocation involving the rearrangements involving the chromosom- probes hybridizing to the 16q22.1 band. 16q22.1 band, two orange/green fusion al region 16q22.1 harboring the CBFB The orange fluorochrome direct labeled signals are expected representing two (core-binding factor subunit beta, a.k.a. probe hybridizes proximal, the green fluo- normal (non-rearranged) 16q22.1 loci. A PEBP2B) gene. CBFB encodes the beta rochrome direct labeled probe hybridizes signal pattern consisting of one orange/ subunit of the CBFA/CBFB transcription distal to the CBFB gene breakpoint region green fusion signal, one orange signal, factor complex involved in myeloid differ- at 16q22.1. This probe is approved to be and a separate green signal indicates one entiation. The chromosomal aberrations used with a hybridization time of 2 hours normal 16q22.1 locus and one 16q22.1 inv(16)(p13.1q22.1) and the related on cytological specimens. locus affected by a translocation. In case translocation t(16;16)(p13.1;q22.1), of a deletion distal to the CBFB breakpoint which have been detected in about 10% region a single orange signal can be of patients with AML (acute myeloblastic expected. leukemia), lead to the fusion of the CBFB gene with the MYH11 (myosin heavy CBFB chain 11) gene on 16p13.1. The resulting CBFB-MYH11 fusion gene is involved in a Ideogram of chromosome 16 indicating the hybridization locations. leukemic transformation. The 5’ segment of the MYH11 gene is known to be delet- D16S2935E RH70212 RH21996 RH54698 ed occasionally as a result of the inversion 5’ 3’ event. AML patients with these genetic CBFB rearrangements have a favorable prog- ~220 kb ~445 kb nosis. Inv(16) may sometimes be difficult Cen 16q22.1 Tel to identify using conventional cytogenetic SPEC CBFB Probe map (not to scale). analysis. Accordingly, Fluorescence in

situ Hybridization proved to be a reliable SPEC CBFB Dual Color Break Apart Probe method overcoming this problem and hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. might consequently be a helpful tool to predict the prognosis of AML patients.

References Aventín A, et al. (2002) Cancer Genet Cytogenet 134: 142-4. Dawson AJ, et al. (2011) Cancer Genet 204: 344-7. Dierlamm J, et al. (1998) Genes Chromosomes Cancer 22: 87-94. Krauter J, et al. (2001) Genes Chromosomes Cancer 30: 342-8. Le Beau MM, et al. (1983) N Engl J Med 309: 630-6. Li MM, et al. (2013) Curr Genet Med Rep 1: 99-112.

Prod. No. Product Label Tests* (Volume) Z-2207-50 ZytoLight SPEC CBFB Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

136 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 136 FE123-1-20 ZytoLight ® SPEC MAF/IGH Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC MAF/IGH Dual The SPEC MAF/IGH Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed to Fusion Probe is a mixture of an orange ange and two green signals are expected. detect the translocations affecting the MAF fluorochrome direct labeled MAF probe A reciprocal translocation involving two gene in the chromosomal region 16q23.2 spanning the known MAF breakpoints, breakpoints splits the two signals and and the IGH locus in 14q32.33. The trans- and a green fluorochrome direct labeled generates a fusion signal on each of the location t(14;16)(q32.3;q23) is frequently IGH probe spanning the known break- chromosomes involved. The chromosomal found in multiple myeloma (MM). points of IGH. regions which are not translocated are MM is a malignant post-germinal center indicated by the single orange and green tumor of somatically-mutated, iso- signal, respectively. type-switched plasma cells that accumulate in the bone marrow. It is often preceded by a premalignant state known as mono- MAF clonal gammopathy of undetermined significance (MGUS). Five recurrent primary translocations involving the immunoglobulin heavy locus (IGH) have been identified in 40% of MGUS and MM tumors. They include t(11;14)(q13.3;q32.3), t(6;14) (p21.1;q32.3), t(4;14)(p16.3;q32.3), IGH t(14;16)(q32.3;q23), and t(14;20) Ideograms of chromosome 16 (above) and 14 (q32.3;q12), which involve the genes (below) indicating the hybridization locations.

CCND1, CCND3, FGFR3 and NSD2, SPEC MAF/IGH Dual Color Dual Fusion Probe SHGC-148602 CHLC.GGAT12D04 MAF, and MAFB, respectively. All of these 5’ 3’ 3’ 5’ hybridized to normal interphase cells as indicated by two orange and two green signals in each nucleus. translocations lead to the dysregulation WWOX MAF and overexpression of the target genes ~1.6 Mb as a consequence of their juxtaposition to Cen 16q23.1-q23.2 Tel regulatory sequences of the IGH locus. SPEC MAF Probe map (not to scale). t(14;16) occurs in approximately 5% of MM patients and is associated with a BV448316 RH119884 3’ 5’ 3’ 5’ more aggressive clinical outcome. The 16q23 breakpoints have been found to be JAG2 IGH scattered 550-1280 kb centromerically to ~ 1.5 Mb Cen 14q32.33 Tel the MAF gene within the WWOX gene. Hence, detection of t(14;16) by FISH rep- SPEC IGH Probe map (not to scale). resents a useful prognostic tool and may aid in therapeutic decision making in MM. Bone marrow CD138+ cells with translocation References Chesi M, et al. (1998) Blood 91: 4457-63. affecting the MAF/IGH loci as indicated by two Fabris S, et al. (2005) Genes Chromosomes Cancer 42: 117-27. orange/green fusion signals, a single orange, and Fonseca R, et al. (2009) Leukemia 23: 2210-21. a separate green signal in each nucleus. Gabrea A, et al. (2006) DNA Repair (Amst) 5: 1225-33. Kindly provided by Prof. Dr. Oskar A. Haas, Vienna, Austria.

Prod. No. Product Label Tests* (Volume) Z-2270-50 ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

137 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 137 FE139-1-20 ZytoLight ® SPEC MAFB/IGH Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC MAFB/IGH Dual The SPEC MAFB/IGH Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed to de- Fusion Probe is a mixture of an orange ange and two green signals are expected. tect the translocations affecting the MAFB fluorochrome direct labeled MAFB probe A reciprocal translocation involving two gene in the chromosomal region 20q12 spanning MAFB and proximal regions breakpoints splits the two signals and and the IGH locus in 14q32.33. The trans- known for variable breakpoints, and a generates a fusion signal on each of the location t(14;20)(q32.3;q12) is frequently green fluorochrome direct labeled IGH chromosomes involved. The chromosomal found in multiple myeloma (MM). probe spanning the known breakpoints of regions which are not translocated are MM is a low proliferative, malignant the IGH locus. indicated by the single orange and green post-germinal center tumor of somatically signal, respectively. mutated, isotype-switched plasma cells that accumulate in the bone marrow. It is often preceded by a premalignant MAFB state known as monoclonal gammopathy of undetermined significance (MGUS). Five recurrent primary translocations involving the immunoglobulin heavy locus (IGH) have been identified in 40% of MGUS and MM tumors. They include t(11;14)(q13.3;q32.3), t(6;14) IGH (p21.1;q32.3), t(4;14)(p16.3;q32.3), Ideograms of chromosome 20 (above) and 14 t(14;16)(q32.3;q23), and t(14;20) (below) indicating the hybridization locations. (q32.3;q12), which involve the genes STSG25129 STSG25332 SPEC MAFB/IGH Dual Color Dual Fusion Probe CCND1, CCND3, FGFR3 and NSD2, 3’5’ hybridized to normal interphase cells as indicated by two orange and two green signals in each nucleus MAF, and MAFB, respectively. All of these and to metaphase chromosomes of a normal cell. MAFB translocations lead to the deregulation ~1.6 Mb and overexpression of the target genes Cen 20q12 Tel

as a consequence of their juxtaposition to SPEC MAFB Probe map (not to scale). regulatory sequences of the IGH locus. The t(14;20) occurs in approximately BV448316 RH119884 3’ 5’ 3’ 5’ 1-2% of MM patients and is associated with an adverse prognosis. Thus, currently, JAG2 IGH detection of t(14;20) by FISH is a reliable ~ 1.5 Mb Cen 14q32.33 Tel prognostic tool and may sustain therapeutic decision making in MM. SPEC IGH Probe map (not to scale).

References Boersma-Vreugdenhil GR, et al. (2004) Br J Haematol 126: 355-63. Chesi M, et al. (1998) Blood 92: 4457-63. Fabris S, et al. (2005) Genes Chromosomes Cancer 42: 117-27. Fonseca R, et al. (2009) Leukemia 23: 2210-21. Bone marrow CD138+ cells with translocation Gabrea A, et al. (2006) DNA Repair (Amst) 5: 1225-33. affecting the MAFB/IGH loci as indicated by two Hanamura I, et al. (2001) Jpn N Cancer Res 92: 638-44. orange/green fusion signals, a single orange, and a separate green signal in each nucleus.

Kindly provided by Prof. Dr. Oskar A. Haas, Vienna, Austria.

Prod. No. Product Label Tests* (Volume) Z-2271-50 ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

138 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 138 FE140-1-20 ZytoLight ® SPEC TP53/17q22 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC TP53/17q22 Dual The SPEC TP53/17q22 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed for the detection is a mixture of an orange fluorochrome di- ange and two green signals are expected. of TP53 deletions as well as for the deter- rect labeled SPEC TP53 probe hybridizing In a cell with deletion of the TP53 gene mination of copy number changes of the to the TP53 gene in the chromosomal re- locus, one orange signal and two green chromosomal region 17q22, harboring gion 17p13.1 and a green fluorochrome signals can be detected. A gain of 17q the MPO (myeloperoxidase) gene. TP53 direct labeled SPEC 17q22 probe specific involving the 17q22 region will result loss in combination with signal gain of for the chromosomal region 17q22 har- in three or more green signals and two the 17q22 chromosomal region serves as boring the MPO gene. orange signals. Isochromosome 17q is a marker for the detection of isochromo- indicated by three green signals and one somes often found in hematologic malig- TP53 orange signal. nancies as well as in neuroblastoma. The TP53 gene (tumor protein p53, a.k.a. 17q22 p53, BCC7, LFS1, TRP53) is located in the chromosomal region 17p13.1 and en- Ideogram of chromosome 17 codes a 53 kDa transcription factor. TP53 indicating the hybridization locations. gene deletions have been detected in RH76489 SGC30770 patients with chronic lymphocytic leukemia 3’ 5’ (CLL), multiple myeloma (MM), and acute TP53 myeloid leukemia (AML). In CLL patients, ~165 kb allelic loss of the short arm of chromosome Tel 17p13.1 Cen 17 is associated with treatment failure SPEC TP53 Probe map (not to scale). with alkylating agents and short survival times. RH46553 40H14T7 3’ 5’ SPEC TP53/17q22 Dual Color Probe hybridized Isochromosome 17q is a frequent cytoge- to bone marrow tissue section with deletion of the TP53 gene as indicated by one green signal netic abnormality seen in hematologic MPO and two orange signals in each nucleus. malignancies including blast phase of ~470 kb chronic myelogenous leukemia (CML), Cen 17q22 Tel AML, Hodgkin and non-Hodgkin lym- SPEC 17q22 Probe map (not to scale). phomas. In neuroblastoma, gain of the 17q21-qter is associated with stronger tumor progression. Thus, the combined detection of both targets by Fluorescence in situ Hybridiza- tion allows for a sensitive determination of isochromosomes and may be a helpful tool for diagnosis and selecting treatment.

References Becher R, et al. (1990) Blood 8: 1679-83. Bown N, et al. (1999) N Engl J Med 340: 1954-61. Fioretos T, et al. (1999) Blood 94: 225-32. SPEC TP53/17q22 Dual Color Probe Pettitt AR, et al. (2001) Blood 98: 814-22. hybridized to a bone marrow smear with Ripollés L, et al. (2006) Cancer Genet Cytogenet 171: 57-64. isochromosome 17q as indicated by three Shanafelt TD, et al. (2006) Ann Intern Med 145: 435-47. green signals and one orange signal.

Prod. No. Product Label Tests* (Volume) Z-2198-50 ZytoLight SPEC TP53/17q22 Dual Color Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

139 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 139 FE112-1-20 ZytoLight ® SPEC TP53/CEN 17 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC TP53/CEN 17 Dual The SPEC TP53/CEN 17 Dual Color In a normal interphase nucleus, two or- Color Probe is designed for the detection Probe is a mixture of a green fluoro- ange and two green signals are expected. of TP53 gene deletions observed e.g. in chrome direct labeled CEN 17 probe In a cell with deletions affecting the TP53 chronic lymphocytic leukemia (CLL). specific for the alpha satellite centromeric gene locus, one or no copy of the orange The TP53 gene (tumor protein p53, a.k.a. region of chromosome 17 (D17Z1) and signal will be observed. p53, BCC7, LFS1, TRP53) is located in an orange fluorochrome direct labeled the chromosomal region 17p13.1 and SPEC TP53 probe specific for the TP53 encodes a 53 kDa transcription factor gene at 17p13.1. which regulates cell proliferation, differen- TP53 tiation, and apoptosis and which functions as a tumor suppressor by activating CEN 17 (D17Z1) the expression of genes that inhibit cell growth. Deletions affecting the short arm of chromosome 17 (17p), the site of the Ideogram of chromosome 17 TP53 gene, are often accompanied by indicating the hybridization locations. mutations in the remaining allele, and thus RH76489 SGC30770 result in the loss of TP53 tumor suppressor 3’ 5’ activity. SPEC TP53/CEN 17 Dual Color Probe hybridized TP53 TP53 gene deletions have been detect- to normal interphase cells as indicated by two ~165 kb orange and two green signals in each nucleus. ed in patients with chronic lymphocytic Tel 17p13.1 Cen leukemia (CLL), multiple myeloma (MM), SPEC TP53 Probe map (not to scale). acute myeloid leukemia (AML), and are also very frequent in primary solid tumors of different histological origin. The presence of TP53 deletion has been shown to correlate with more aggressive disease, shortened survival, and poor response to standard treatment. CLL patients with deletion of 17p are more likely to respond to treatment with the monoclonal anti-CD52 antibody alemtuzumab than to conven-

tional chemotherapy. FISH is an effective SPEC TP53/CEN 17 Dual Color Probe hybridized method to screen for deletions affecting to bone marrow tissue section with deletion of the TP53 gene as indicated by one orange signal the TP53 gene locus in order to identify References and two green signals in each nucleus. Amiel A, et al. (1997) Cancer Genet Cytogenet 97: 97-100. patients who are candidates for alterna- Chang H, et al. (2005) Blood 105: 358-60. Chang H, et al. (2010) Am J Clin Pathol 133: 70-4. tive treatment and to avoid administration Herrera JC, et al. (2010) Biomedica 30: 390-400. Lozanski G, et al. (2004) Blood 103: 3278-81. of otherwise ineffective therapy. Tavor S, et al. (2011) Leuk Lymphoma 52: 642-7.

Prod. No. Product Label Tests* (Volume) Z-2153-50 ZytoLight SPEC TP53/CEN 17 Dual Color Probe / 5 (50 μl) Z-2153-200 ZytoLight SPEC TP53/CEN 17 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

140 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 140 FE070-1-20 ZytoLight ® SPEC USP6 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC USP6 Dual Color The SPEC USP6 Dual Color Break Apart In an interphase nucleus lacking a translo- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cation involving the 17p13.2 band, two translocations involving the chromosomal probes hybridizing to the 17p13.2 band. orange/green fusion signals are expected region 17p13.2 harboring the USP6 The orange fluorochrome direct labeled representing two normal (non-rearranged) (Ubiquitin-specific peptidase 6, a.k.a. probe hybridizes proximal to the USP6 17p13.2 loci. A signal pattern consisting TRE2 or TRE17) gene. gene and the green fluorochrome direct la- of one orange/green fusion signal, one Translocations affecting USP6 have been beled probe hybridizes distal to that gene. orange signal, and a separate green initially found in primary aneurysmal bone signal indicates one normal 17p13.2 USP6 cysts (ABC), a benign, but locally aggres- locus and one 17p13.2 locus affected by sive bone lesion that occurs predominantly a translocation. during the first two decades of life. USP6 rearrangements are restricted to spindle cells in primary ABC, indistinguishable Ideogram of chromosome 17 from surrounding normal spindle cells. indicating the hybridization locations. The resulting fusion genes detected are RH41805 RH54973 formed by juxtaposition of the USP6 cod- RH41994 RH36080 ing sequences to the highly active promot- 5’ 3’

er sequences of several partner genes, as USP6

e.g. CDH11, COL1A1, OMD, TRAP150, ~530 kb ~275 kb and ZNF9, leading to the transcriptional Tel 17p13.2 Cen upregulation of USP6. No true fusion SPEC USP6 Probe map (not to scale). genes are formed. More recently, nodular fasciitis (NF), an- SPEC USP6 Break Apart Probe hybridized to aneurysmal bone cyst tissue section with polysomy other mesenchymal lesion, has been tested of chromosome 17 but without translocation affecting positive for USP6 rearrangements. NF is a the 17p13.2 locus as indicated by multiple orange/ green fusion signals per nucleus. subcutaneous pseudosarcomatous myofibroblastic proliferation of unknown pathogenesis that regresses spontaneously when not surgically resected. The translocation results in the fusion of the promoter region of MYH9 located on 22q12.3 to the entire coding sequence of USP6 and subsequently in upregulated USP6 expression. For both lesions it is assumed that the detection of USP6 rearrangements by Fluorescence in situ Hybridization might represent a valuable diagnostic tool.

References Erickson-Johnson MR, et al. (2011) Lab Invest 91: 1427-33. Aneurysmal bone cyst tissue section with translocation Nakamura T, et al. (1988) Oncogene Res 2: 357-70. affecting the 17p13.2 locus as indicated by one Oliveira AM, et al. (2004) Cancer Res 64: 1920-3. Oliveira AM, et al. (2005) Oncogene 24: 3419-26. orange/green fusion (non-rearranged) signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2151-50 ZytoLight SPEC USP6 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

141 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 141 FE071-1-20 ZytoLight ® SPEC YWHAE Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC YWHAE Dual Color The SPEC YWHAE Dual Color Break In an interphase nucleus of a normal Break Apart Probe is designed to detect Apart Probe is a mixture of two direct cell lacking a translocation involving the rearrangements involving the chromosom- labeled probes hybridizing to the 17p13.3 band, two orange/green fusion al region 17p13.3 harboring the YWHAE 17p13.3 band. The orange fluorochrome signals are expected representing two (tyrosine 3-monooxygenase/tryptophan direct labeled probe hybridizes distal to normal (non-rearranged) 17p13.3 loci. 5-monooxygenase activation protein, the YWHAE gene breakpoint region at A signal pattern consisting of one orange/ epsilon a.k.a. 14-3-3 epsilon) gene. 17p13.3, the green fluorochrome direct green fusion signal, one orange signal, YWHAE encodes a protein of the 14-3-3 labeled probe hybridizes proximal to and a separate green signal indicates one family which is involved in regulation of the YWHAE gene breakpoint region at normal 17p13.3 locus and one 17p13.3 cellular proliferation, metabolism, and 17p13.3. locus affected by a translocation. differentiation. However, altered expression of 14-3-3 family proteins is associat- YWHAE ed with development and progression of cancer. The fusion between YWHAE and one of the FAM22 family members (FAM22A or Ideogram of chromosome 17 FAM22B) caused by a t(10;17)(q22;p13) indicating the hybridization locations. has been identified in the clinically ag- RH113855 CRK gressive, high-grade endometrial stromal SHGC-130461 HUMUT6768 sarcoma (ESS) as well as in clear cell 3’ 5’

sarcoma of the kidney (CCSK). YWHAE

In contrast to the classic low-grade form ~335 kb ~375 kb of ESS harboring JAZF1 gene fusions, Tel 17p13.3 Cen Endometrial stromal sarcoma tissue section with translocation affecting the YWHAE gene as indicated YWHAE-FAM22 ESS display high-grade SPEC YWHAE Probe map (not to scale). by one non-rearranged orange/green fusion signal, histologic features and an aggressive one orange, and one separate green signal. clinical course. Moreover, due to the lack of estrogen and progesterone receptor expression in YWHAE-FAM22 ESS, the hormonal therapy used to treat low-grade ESS is likely to be ineffective. Consequent- ly, differentiation between YWHAE-FAM22 and JAZF1 ESS by FISH is clinically relevant to support the diagnosis and may aid in therapeutic decision making. References Isphording A, et al. (2013) Hum Pathol 44: 837-43. Lee CH, et al. (2012) Proc Natl Acad Sci U S A 109: 929-34. O‘Meara E, et al. (2012) J Pathol 227: 72-80. Stewart JC, et al. (2014) Histopathology 65: 473-82.

Prod. No. Product Label Tests* (Volume) Z-2175-50 ZytoLight SPEC YWHAE Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

142 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 142 FE093-1-20 ZytoLight ® SPEC ERBB2/CEN 17 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ERBB2/CEN 17 The SPEC ERBB2/CEN 17 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of an orange fluoro- ange and two green signals are expected. detection of ERBB2 gene amplification chrome direct labeled CEN 17 probe In a cell with amplification of the ERBB2 frequently observed in solid malignant specific for the alpha satellite centro- gene locus, multiple copies of the green neoplasms e.g. breast cancer samples. meric region of chromosome 17 (D17Z1) signal or green signal clusters will be The ERBB2 gene (a.k.a. HER2 and NEU) is and a green fluorochrome direct labeled observed. located in the chromosomal region 17q12 SPEC ERBB2 probe specific for the chro- and encodes a 185-190 kDa transmem- mosomal region 17q12-q21.1 harboring brane glycoprotein, p185, acting as a the ERBB2 gene. cellular growth factor receptor. The p185 protein belongs to the EGFR (epidermal CEN 17 (D17Z1) growth factor receptor) subgroup of the ERBB2 RTK (receptor tyrosine kinase) superfamily also including EGFR (ERBB1, HER1),

ERBB3 (HER3), and ERBB4 (HER4). Ideogram of chromosome 17 Amplification of the proto-oncogene indicating the hybridization locations. ERBB2, observed in approximately 20% of HUMUT6435 D17S1179 all breast cancer samples, has been cor- 5’ 3’ related with a poor prognosis of the dis- ERBB2 Normal interphase cells, ease. Similar results have been obtained ~610 kb ERBB2 (green), CEN 17 (orange). for a variety of other malignant neoplasms Cen 17q12-q21.1 Tel

e.g. ovarian cancer, stomach cancer, and SPEC ERBB2 Probe map (not to scale). carcinomas of the salivary gland.

References Baselga J, et al. (1999) Semin Oncol 26: 78-83. Brockhoff G, et al. (2016) Histopathology 69: 635-46. Brunello E, et al. (2012) Histopathology 60: 482-8. Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Coussens L, et al. (1985) Science 230: 1132-9. Ettl T, et al. (2012) Br J Cancer 106: 719-26. Hwang CC, et al. (2011) Histopathology 59: 984-92. Hynes NE & Stern DF (1994) Biochim Biophys Acta 1198: 165-84. Moelans CB, et al. (2011) Crit Rev Oncol Hematol 80: 380-92. Park JB, et al. (1989) Cancer Res 49: 6605-9. Popescu NC, et al. (1989) Genomics 4: 362-6. Sassen A, et al. (2008) Breast Cancer Res 10: R2. Slamon DJ, et al. (1987) Science 235: 177-82. Voutsas IF, et al. (2013) Int J Radiat Biol 89: 319-25. Wolff AC, et al. (2018) J Clin Oncol 14: 437-41.

Breast carcinoma tissue section, ERBB2 gene cluster (green), CEN 17 (orange).

Prod. No. Product Label Tests* (Volume) Z-2015-50 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe / 5 (50 μl) Z-2015-200 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe / 20 (200 μl) Z-2020-5 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit / 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; Probe, 0.05 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2020-20 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit / 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; Probe, 0.2 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

143 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 143 FE006-1-20 ZytoLight ® CEN 17/SPEC ERBB2 Dual Color Probe

Background Probe Description Results The ZytoLight ® CEN 17/SPEC ERBB2 The CEN 17/SPEC ERBB2 Dual Color In a normal interphase nucleus, two orange Dual Color Probe is designed for the Probe is a mixture of a green fluorochrome and two green signals are expected. In a detection of ERBB2 gene amplification direct labeled CEN 17 probe specific for cell with amplification of the ERBB2 gene frequently observed in solid malignant the alpha satellite centromeric region of locus, multiple copies of the orange signal neoplasms e.g. breast cancer samples. chromosome 17 (D17Z1) and an orange or orange signal clusters will be observed. The ERBB2 gene (a.k.a. HER2 and NEU) is fluorochrome direct labeled SPEC ERBB2 located in the chromosomal region 17q12 probe specific for the chromosomal region and encodes a 185-190 kDa transmem- 17q12-q21.1 harboring the ERBB2 gene. brane glycoprotein, p185, acting as a cellular growth factor receptor. The p185 protein belongs to the EGFR (epidermal CEN 17 (D17Z1) ERBB2 growth factor receptor) subgroup of the RTK (receptor tyrosine kinase) superfamily also including EGFR (ERBB1, HER1), Ideogram of chromosome 17 ERBB3 (HER3), and ERBB4 (HER4). indicating the hybridization locations. Amplification of the proto-oncogene HUMUT6435 D17S1179 ERBB2, observed in approximately 20% 5’ 3’ of all breast cancer samples, has been Normal interphase cells, correlated with a poor prognosis of the ERBB2 ERBB2 (orange), CEN 17 (green). disease. ~610 kb Cen 17q12-q21.1 Tel Similar results have been obtained for SPEC ERBB2 Probe map (not to scale). a variety of other malignant neoplasms e.g. ovarian cancer, stomach cancer, and carcinomas of the salivary gland.

Prod. No. Product Label Tests* (Volume) Z-2077-50 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe / 5 (50 μl) Z-2077-200 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

144 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 144 FE099-1-20 ZytoLight ® SPEC ERBB2/D17S122 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC ERBB2/D17S122 The SPEC ERBB2/D17S122 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of a green fluorochrome ange and two green signals are expected. detection of ERBB2 gene amplification direct labeled SPEC ERBB2 probe specific In a cell with amplification of the ERBB2 frequently observed in solid malignant neo- for the chromosomal region 17q12-q21.1 gene locus, multiple copies of the green plasms e.g. breast cancer samples. harboring the ERBB2 gene and an signal or green signal clusters will be The ERBB2 gene (a.k.a. HER2 and NEU) is orange fluorochrome direct labeled SPEC observed. located in the chromosomal region 17q12 D17S122 probe specific for the chromo- and encodes a 185-190 kDa transmem- somal region 17p12. The SPEC D17S122 brane glycoprotein, p185, acting as a probe is designed to be used for chromo- cellular growth factor receptor. The p185 some 17 copy number detection. protein belongs to the EGFR (epidermal growth factor receptor) subgroup of the D17S122 RTK (receptor tyrosine kinase) superfamily ERBB2 also including EGFR (ERBB1, HER1), ERBB3 (HER3), and ERBB4 (HER4). Amplification

of the proto-oncogene ERBB2, observed Ideogram of chromosome 17 in approximately 20% of all breast cancer indicating the hybridization locations. samples, has been correlated with a poor HUMUT6435 D17S1179 prognosis of the disease. 5’ 3’ Normal interphase cells, ERBB2 (green), Similar results have been obtained for a D17S122 (orange). ERBB2 variety of other malignant neoplasms e.g. ~610 kb ovarian cancer, stomach cancer, and carci- Cen 17q12-q21.1 Tel

nomas of the salivary gland. SPEC ERBB2 Probe map (not to scale). Fluorescence in situ Hybridization targeting the alpha satellite centromeric regions of D17S1296 D17S122 SHGC-142057 chromosome 17 may be misleading in some cases due to possible gains or losses of this region. For these cases, reflex testing ~480 kb Tel 17p12 Cen is recommended using the SPEC ERBB2/ SPEC D17S122 Probe map (not to scale). D17S122 Dual Color Probe. References Baselga J, et al. (1999) Semin Oncol 26: 78-83. Coussens L, et al. (1985) Science 230: 1132-9. Hwang CC, et al. (2011) Histopathology 59: 984-92. Hynes NE & Stern DF (1994) Biochim Biophys Acta 1198: 165-84. Breast carcinoma tissue section, ERBB2 Moelans CB, et al. (2011) Crit Rev Oncol Hematol 80: 380-92. gene cluster (green), D17S122 (orange). Park JB, et al. (1989) Cancer Res 49: 6605-9. Popescu NC, et al. (1989) Genomics 4: 362-6. Slamon DJ, et al. (1987) Science 235: 177-82. Voutsas IF, et al. (2013) Int J Radiat Biol 89: 319-25. Wolff AC, et al. (2018) J Clin Oncol 14: 437-41.

Prod. No. Product Label Tests* (Volume) Z-2190-50 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe / 5 (50 μl) Z-2190-200 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

145 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 145 FE091-1-20 ZytoLight ® SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe

Background Probe Description Results The ZytoLight ® SPEC ERBB2/TOP2A/ The SPEC ERBB2/TOP2A/CEN 17 Triple In a normal interphase nucleus, two green, CEN 17 Triple Color Probe is designed for Color Probe is a mixture of a green two orange, and two blue signals are ex- the simultaneous detection of ERBB2 and fluorochrome direct labeled SPEC ERBB2 pected. In a cell with amplification of the TOP2A gene status. probe specific for the chromosomal region ERBB2 gene locus, multiple copies of the The ERBB2 gene (a.k.a. HER2 and NEU) is 17q12-q21.1 harboring the ERBB2 gene, green signal or large green signal clusters located in the chromosomal region 17q12 an orange fluorochrome direct labeled will be observed. Amplification of TOP2A and encodes a 185 kDa transmembrane SPEC TOP2A probe specific for the will result in multiple copies of the orange glycoprotein. The TOP2A (DNA topo- TOP2A gene at 17q21.2, and a blue flu- signal or large orange signal clusters. isomerase II alpha) gene is located in orochrome direct labeled CEN 17 probe Deletion of the TOP2A gene results in a the chromosomal region 17q21.2 and specific for the alpha satellite centromeric reduced number of orange signals. encodes a 170 kDa DNA topoisomerase. region of chromosome 17 (D17Z1). The TOP2A gene is frequently either co-amplified or deleted in ERBB2 positive breast cancer cases. TOP2A functions as the target for several anticancer agents, ERBB2 e.g. anthracyclines. Recent data suggests that amplification and deletion of the Ideogram of chromosome 17 TOP2A gene locus may account for indicating the hybridization locations. relative chemosensitivity or resistance to D17S1179 D17S1436E TOP2A inhibitor therapy in ERBB2 positive HUMUT6432 RH25598 breast cancer. Thus, determination of the 5’ 3’ 3’ 5’

ERBB2 and TOP2A status may help to pre- ERBB2 TOP2A

dict benefit from adjuvant anthracyclines ~610 kb ~495 kb SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe in breast cancer treatment. Cen 17q12-q21.2 Tel hybridized to normal interphase cells as indicated by two green, two orange, and References SPEC ERBB2/TOP2A Probe map (not to scale). two blue signals per nucleus. Arriola E, et al. (2007) Breast Cancer Res Treat 106: 181-9. Brunello E, et al. (2012) Histopathology 60: 482–8. Coussens L, et al. (1985) Science 230: 1132-9. Fountzilas G, et al. (2012) J Transl Med 10: 212. Fountzilas G, et al. (2012) PLoS One 7: e37946. Fountzilas G, et al. (2013) BMC Cancer 13: 163. Järvinen TA & Liu ET (2006) Curr Cancer Drug Targets 6: 579-602. Popescu NC, et al. (1989) Genomics 4: 362-6. Pritchard KI, et al. (2008) J Clin Oncol 26: 736-44. Razis E, et al. (2011) Breast Cancer Res Treat 128: 447-56. Tsai-Pflugfelder M, et al. (1988) Proc Natl Acad Sci U S A 85: 7177-81.

Breast cancer tissue section with two copies of chromosome 17 (blue) and TOP2A (orange) and ERBB2 gene clusters (green) in each nucleus.

Prod. No. Product Label Tests* (Volume)

Z-2093-50 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe •/•/• 5 (50 μl)

Z-2093-200 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe •/•/• 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

146 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 146 FE016-1-20 ZytoLight ® SPEC COL1A1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC COL1A1 Dual The SPEC COL1A1 Dual Color Break In a normal interphase nucleus lacking Color Break Apart Probe is designed for Apart Probe is a mixture of two direct la- a translocation involving the 17q21.33 the detection of the specific transloca- beled probes hybridizing to the 17q21.33 band, two orange/green fusion signals tions involving the chromosomal region band. The orange fluorochrome direct are expected representing two normal 17q21.33 harboring the COL1A1 (a.k.a. labeled probe hybridizes distal, and the (non-rearranged) 17q21.33 loci. A signal OI4) gene. green fluorochrome direct labeled probe pattern consisting of one orange/green fu- Reciprocal translocations involving hybridizes proximal to the COL1A1 gene. sion signal, one orange signal, and a sep- t(17;22)(q21.3;q13.1) are characteristic arate green signal indicates one normal for dermatofibrosarcoma protuberans 17q21.33 locus and one 17q21.33 locus (DFSP). DFSP is a highly recurrent, infiltra- affected by a 17q21.33 translocation. tive skin tumor of intermediate malignancy. COL1A1 The rearrangements are cytogenetically characterized by the presence of super- Ideograms of chromosome 17 numerary ring chromosomes containing indicating the hybridization locations. low-level amplified sequences from chro- D17S588 D11S2843 mosomes 17q21-qter and 22q10-q13.1, RH55170 RH55094 or unbalanced derivatives of the t(17;22) 3’ 5’

(q21.3;q13.1) translocation. COL1A1

The rearrangement frequently results in ~555 kb ~395 kb formation of a COL1A1-PDGFB fusion Cen 17q21.33 Tel

protein which is post-transcriptionally SPEC COL1A1 Probe map (not to scale). processed to a functional platelet-derived growth factor beta chain (PDGFB) protein, DFSP tissue section with translocation affecting and results in PDFGB-mediated autocrine the 17q21.33 locus as indicated by one non- rearranged orange/green fusion signal, one and/or paracrine activation of the orange signal, and one separate green signal. plateled-derived growth factor receptor-β (PDGFRB). The accurate diagnosis of DFSP is important because of the intermediate malignant nature of the DFSP and can be facilitated by Fluorescence in situ Hybridization (FISH) analyses.

References Labropoulos SV & Razis ED (2007) Biologics 4: 347-53. Patel KU, et al. (2008) Human Pathol 39: 184-93. Shimizu A, et al. (1999) Cancer Res 59: 3719-23. Simon MP, et al. (1997) Nat Genet 15: 95-8.

DFSP tissue section with amplification of the 17q21-qter and 22q10–q13.1 sequences probably due to a COL1A1-PDGFB fusion product on the ring chromosome.

Image kindly provided by Dr. Schildhaus, Essen, Germany.

Prod. No. Product Label Tests* (Volume) Z-2121-200 ZytoLight SPEC COL1A1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

147 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 147 FE043-1-20 ZytoLight ® SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC COL1A1/PDGFB The SPEC COL1A1/PDGFB Dual Color In a normal interphase nucleus, two or- Dual Color Dual Fusion Probe is designed Dual Fusion Probe is a mixture of an ange and two green signals are expected. for the detection of the specific transloca- orange fluorochrome direct labeled A reciprocal translocation involving two tions involving the chromosomal region COL1A1 probe covering the breakpoint breakpoints splits the two signals and 17q21.33 harboring the COL1A1 (a.k.a. region of the COL1A1 gene and a green generates a fusion signal on each of the OI4) gene, and the chromosomal region fluorochrome direct labeled PDGFB probe chromosomes involved. The chromosomal 22q13.1, harboring the PDGFB (a.k.a covering the breakpoint region of the regions which are not translocated are PDGF2, SIS) gene. PDGFB gene. indicated by the single orange and green The reciprocal translocations involving signal, respectively. t(17;22)(q21.3;q13.1) are characteristic for dermatofibrosarcoma protuberans (DFSP) patients. DFSP is a highly recur- COL1A1 rent, infiltrative skin tumor of intermediate malignancy. The rearrangements are cytogenetically characterized by the presence of super- numerary ring chromosomes containing PDGFB low-level amplified sequences from chro- Ideograms of chromosomes 17 (above) and mosomes 17q21-qter and 22q10-q13.1, 22 (below) indicating the hybridization locations. or unbalanced derivatives of the t(17;22) SHGC-2255 RH55094 (q21.3;q13.1) translocation. 3’ 5’ The rearrangement results in a COL1A1- SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe PDGFB fusion protein which is post-tran- COL1A1 hybridized to normal interphase cells as indicated by two orange and two green signals in each nucleus. scriptionally processed to a functional ~925 kb Cen 17q21.33 Tel platelet-derived growth factor beta chain SPEC COL1A1 Probe map (not to scale). (PDGFB) protein, and results in PDFGB- mediated autocrine and/or paracrine STDJ358M15T7 D22S575 3’ 5’ activation of the plateled-derived growth factor receptor-β (PDGFRB). PDGFB The accurate diagnosis of DFSP is impor- ~1.3 Mb tant because of the intermediate malignant Cen 22q13.1 Tel nature of the DFSP and can be facilitated SPEC PDGFB Probe map (not to scale). by Fluorescence in situ Hybridization (FISH) analyses.

References Labropoulos SV & Razis ED (2007) Biologics 4: 347-53. Patel KU, et al. (2008) Human Pathol 39: 184-93. Shimizu A, et al. (1999) Cancer Res 59: 3719-23. DFSP tissue section with translocation affecting Simon MP, et al. (1997) Nat Genet 15: 95-8. the COL1A1/PDGFB loci as indicated by one Walluks K, et al. (2013) Pathol Res Pract 209: 30-5. separate orange signal, one separate green signal, and two orange/green fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2116-50 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe / 5 (50 μl) Z-2116-200 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

148 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 148 FE050-1-20 ZytoLight ® SPEC SS18 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC SS18 Dual Color The SPEC SS18 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 18q11.2 band two translocations involving the chromosomal probes hybridizing to the 18q11.2 band. orange/green fusion signals are expected region 18q11.2 harboring the SS18 The orange fluorochrome direct labeled representing two normal (non-rearranged) (SS18, nBAF chromatin remodeling com- probe hybridizes distal to the SS18 gene, 18q11.2 loci. A signal pattern consisting plex subunit, a.k.a. SYT) gene. the green fluorochrome direct labeled of one orange/green fusion signal, one Translocations involving the region probe hybridizes proximal to that gene. orange signal, and a separate green sig- 18q11.2 are found in over 90% of nal indicates one normal 18q11.2 locus synovial sarcoma. Among soft tissue and one 18q11.2 locus affected by an sarcomas, synovial sarcoma is one of SS18 18q11.2 translocation. the most common and classically occurs in the extremities of young adults with greater prevalence in males even though, Ideogram of chromosome 18 indicating the hybridization locations. the occurrence of synovial sarcoma has also been described in a wide variety of SHGC-140723 RH112040 RH119469 RH18471 anatomical locations and in all ages. 3’ 5’ The most frequent translocation involving the SS18 gene region is t(X;18) SS18 (p11.23;q11.2) juxtaposing the SS18 ~355 kb ~365 kb Cen 18q11.2 Tel gene in 18q11.2 either next to the SSX1 SPEC SS18 Probe map (not to scale). (synovial sarcoma, translocated to X chromosome) or the SSX2 gene, or very rarely

to the SSX4 locus located in Xp11.23. SPEC SS18 Dual Color Break Apart Probe hybridized Complex translocations involving other to normal interphase cells as indicated by two orange/green fusion signals per nucleus. chromosomes are observed in less than 10% of synovial sarcomas. In combination with histopathological diagnosis, detection of SS18 rearrangements via FISH analysis is a valuable tool to confirm the diagnosis of synovial sarcoma.

References Amary MF, et al. (2007) Mod Pathol 20: 482-96. Clark J, et al. (1994) Nat Genet 7: 502-8. Ilmiawan MI, et al. (2012) Med J Indones 21: 196-202. Kawai A, et al. (1998) N Engl J Med 338: 153-60. Surace C, et al. (2004) Lab Invest 84: 1185-92. Torres L, et al. (2008) Cancer Genet Cytogenet 187: 45-9.

Synovial sarcoma tissue section with translocation affecting the 18q11.2 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2097-50 ZytoLight SPEC SS18 Dual Color Break Apart Probe / 5 (50 μl) Z-2097-200 ZytoLight SPEC SS18 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

149 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 149 FE032-1-20 ZytoLight ® SPEC SS18/SSX1 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC SS18/SSX1 The SPEC SS18/SSX1 TriCheck™ Probe is In an interphase nucleus of a normal TriCheck™ Probe is designed to detect trans- a mixture of three direct labeled probes female cell without SS18-SSX1 rearrange- locations involving the chromosomal region hybridizing to the 18q11.2 and Xp11.23 ment, two green/orange fusion signals 18q11.2 harboring the SS18 (SS18, nBAF bands. The green fluorochrome direct la- and two blue signals are expected. chromatin remodeling complex subunit, beled probe hybridizes proximal and the In an interphase nucleus of a normal male a.k.a. SYT) gene and the chromosomal orange fluorochrome direct labeled probe cell without SS18-SSX1 rearrangement, region Xp11.23 harboring the SSX1 (SSX hybridizes distal to the SS18 breakpoint two green/orange fusion signals and one family member 1) gene. region at 18q11.2. The blue fluorochrome blue signal are expected. Synovial sarcoma is characterized by the direct labeled probe hybridizes proximal An SS18-SSX1 or an SS18-SSX4 fusion is t(X;18) found in more than 95% of these to the SSX1 gene at Xp11.23. indicated by one separate orange signal tumors and juxtaposing the SS18 gene in co-localizing with one blue signal and one 18q11.2 either next to the SSX1 or the separate green signal. SSX2 gene, or very rarely to the SSX4 SS18 An SS18-SSX2 fusion is indicated by locus. one separate green signal, one separate Synovial sarcoma is one of the most com- orange signal, and a blue signal in close mon soft tissue tumors that typically occurs proximity of the separated green signal. in the extremities of young adults with An SS18 translocation without involvement greater prevalence in males, even though, of SSX1, SSX2, or SSX4 is indicated by the occurrence of synovial sarcoma has SSX1 the split of one green/orange fusion signal also been described in a wide variety of without co-localization of the separated or- anatomical locations and in all ages. ange or green signal with one blue signal. In combination with histopathological diagnosis, detection of SS18 rearrangements via FISH is a valuable tool to confirm the diagnosis of synovial sarcoma. Moreover, Ideograms of chromosomes 18 (above) and patients with SS18-SSX1 fusions were X (below) indicating the hybridization locations.

shown to have a higher risk of developing SHGC-140723 RH112040 metastases compared to those with SS18- RH119469 RH18471 3’ 5’ SSX2 fusions. Hence, detection of the SS18 fusion gene variant by FISH may also be of SS18 prognostic significance. ~355 kb ~365 kb Male synovial sarcoma tissue section with Cen 18q11.2 Tel SS18-SSX1 or SS18-SSX4 fusion as indicated References by orange/blue fusion signals. Amary MF, et al. (2007) Mod Pathol 20: 482-96. Clark J, et al. (1994) Nat Genet 7: 502-8. SPEC SS18 Probe map (not to scale). Kawai A, et al. (1998) N Engl J Med 338: 153-60. Panagopoulos I, et al. (2001) Genes Chromosomes Cancer 31: 362-72. Surace C, et al. (2004) Lab Invest 84: 1185-92. RH66279 RH18187 Torres L, et al. (2008) Cancer Genet Cytogenet 187: 45-9. 5’ 3’ 5’ 3’ 5’ 3’

SSX1 SSX4 SSX2

~525 kb Tel Xp11.23 Cen

SPEC SSX1 Probe map (not to scale).

Female synovial sarcoma tissue section with SS18- SSX2 fusion as indicated by green/blue fusion signals.

Prod. No. Product Label Tests* (Volume) Z-2184-50 ZytoLight SPEC SS18/SSX1 TriCheck Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

150 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 150 FE116-1-20 ZytoLight ® SPEC BCL2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC BCL2 Dual Color The SPEC BCL2 Dual Color Break In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Apart Probe is a mixture of two di- location involving the 18q21.33-q22.1 translocations involving the chromosomal re- rect labeled probes hybridizing to the band, two orange/green fusion signals gion 18q21.33 harboring the BCL2 gene. 18q21.33-q22.1 band. The green are expected representing two normal The BCL2 (BCL2 apoptosis regulator, a.k.a. fluorochrome direct labeled probe hybrid- (non-rearranged) 18q21.33-q22.1 loci. PPP1R50) gene encodes a mitochondrial izes proximal to the BCL2 gene, and the A signal pattern consisting of one orange/ membrane protein that regulates apoptosis orange fluorochrome direct labeled probe green fusion signal, one orange signal, and is expressed in B-cells. hybridizes distal to the BCL2 locus. and a separate green signal indicates Translocations involving the BCL2 gene are one normal 18q21.33-q22.1 locus and commonly identified in B-cell lymphomas. one 18q21.33-q22.1 locus affected by a In particular, the translocation t(14;18) translocation. (q32.3;q21.3) has been identified in about BCL2 80% of follicular lymphoma (FL), in 20% to 30% of diffuse large B-cell lymphoma (DLB- Ideogram of chromosome 18 indicating CL), and rarely in B-cell chronic lymphocytic the hybridization locations. leukemia (B-CLL). RH55422 RH44852 SHGC-147772 REN12166 In FL this translocation is considered to be a 3’ 5’ cytogenetic hallmark. As a result of this rearrangement, the BCL2 gene is juxtaposed BCL2 to IGH (Immunglobulin heavy locus) at ~545 kb ~665 kb Cen 18q21.33-q22.1 Tel 14q32.33 which leads to overexpression of the anti-apoptotic protein BCL2, and finally SPEC BCL2 Probe map (not to scale). to progression to lymphoma. SPEC BCL2 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated Alternative BCL2 translocations to immuno- by two orange/green fusion signals per nucleus. globulin light chain genes as well as non-IG translocation events have been reported. In DLBCL, BCL2 gene overexpression has been implicated in conferring resistance to chemotherapy and has been associated References Da Cunha Santos G, et al. (2011) Cancer Cytopathol 119: 254-62. with poor prognosis. Dyer MJ, et al. (1994) Blood 83: 3682-8. Gu K, et al. (2008) Arch Pathol Lab Med 132: 1355-61. Hence, detection of BCL2 translocations by Hockenbery D, et al. (1990) Nature 348: 334-6. Impera L, et al. (2008) Oncogene 27: 6187-90. Fluorescence in situ Hybridization (FISH) López-Guillermo A, et al. (1999) Blood 93: 3081-7. Nelson BP, et al. (2007) Am J Clin Pathol 128: 323-32. may be of diagnostic and prognostic Tibiletti MG, et al. (2009) Hum Pathol 40: 645-52. Tomita N, et al. (2009) Haematologica 94: 935-43. relevance. Weinberg OK, et al. (2007) J Mol Diagn 9: 530-7.

Neck lymph node tissue section with translocation of the BCL2 gene as indicated by two non-rearranged orange/green fusion signals, one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2192-50 ZytoLight SPEC BCL2 Dual Color Break Apart Probe / 5 (50 μl) Z-2192-200 ZytoLight SPEC BCL2 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

151 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 151 FE092-1-20 ZytoLight ® SPEC BCL2/CEN 18 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC BCL2/CEN 18 Dual The SPEC BCL2/CEN 18 Dual Color In a normal interphase nucleus, two or- Color Probe is designed for the detection Probe is a mixture of a green fluoro- ange and two green signals are expected. of amplifications of the chromosomal chrome direct labeled SPEC BCL2 probe In a cell with amplification of the BCL2 region harboring the BCL2 gene. hybridizing to the human BCL2 gene in gene locus, multiple copies of the green The BCL2 (BCL2 apoptosis regulator, the chromosomal region 18q21.33 and signal or green signal clusters will be a.k.a. PPP1R50) gene is located on an orange fluorochrome direct labeled observed. chromosome 18q21.33 and encodes an CEN 18 probe specific for the alpha satel- antiapoptosis factor involved in normal lite centromeric region of chromosome 18 B-cell development and differentiation. (D18Z1). The expression of BCL2 usually decreas- es upon B-cell differentiation. However, CEN 18 (D18Z1) increased BCL2 expression has been detected in lymphomas harboring the BCL2 translocation t(14;18)(q32.3;q21.3).

Moreover, overexpression of BCL2 can Ideogram of chromosome 18 indicating also be caused by amplification of the the hybridization locations. BCL2 gene as detected in diffuse large BV677679 RH110938 3’ 5’ B-cell lymphoma (DLBCL) and mantle cell

lymphoma (MCL). BCL2 SPEC BCL2/CEN 18 Dual Color Probe hybridized DLBCL is the most common type of ~505 kb to normal interphase cells as indicated by two orange and two green signals in each nucleus. non-Hodgkin lymphoma characterized by Cen 18q21.33 Tel an aggressive clinical course. On the ba- SPEC BCL2 Probe map (not to scale). sis of their gene expression profiles, ABC (activated B-cell-like) and GCB (germinal center B-cell-like) were identified as two molecular subtypes of DLBCL. BCL2 was found to be frequently amplified in the ABC subgroup of DLBCL but rarely in the GCB subgroup. BCL2 overexpression as a result of 18q21 amplification is associated with poor survival in the ABC subgroup. Hence, the identification of BCL2 gene copy number changes by Fluorescence in situ Hybridization may be of prognostic significance in non-Hodgkin lymphomas.

References Alizadeh AA, et al. (2000) Nature 403: 503-11. Beà S, et al. (2009) Blood 113: 3059-69. Iqbal J, et al. (2006) J Clin Oncol 24: 961-8. Monni O, et al. (1999) Leuk Lymphoma 34: 45-52.

Prod. No. Product Label Tests* (Volume) Z-2174-50 ZytoLight SPEC BCL2/CEN 18 Dual Color Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

152 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 152 FE081-1-20 ZytoLight ® SPEC BCL2/IGH Dual Color Dual Fusion Probe

Background Probe Description Results The ZytoLight ® SPEC BCL2/IGH Dual The SPEC BCL2/IGH Dual Color Dual In a normal interphase nucleus, two or- Color Dual Fusion Probe is designed Fusion Probe is a mixture of an orange ange and two green signals are expected. to detect the translocation t(14;18) fluorochrome direct labeled BCL2 probe A reciprocal translocation involving two (q32.3;q21.3) affecting the BCL2 gene in spanning the known BCL2 breakpoints, breakpoints splits the two signals and the chromosomal region 18q21.33 and and a green fluorochrome direct labeled generates a fusion signal on each of the the IGH locus in 14q32.33. IGH probe spanning the known break- chromosomes involved. The chromosomal Translocations involving the BCL2 (BCL2 points of IGH. regions which are not translocated are apoptosis regulator) gene and the IGH indicated by the single orange and green (immunoglobulin heavy locus, a.k.a. signal, respectively. IGH@) gene are considered to be cytogenetic hallmarks for follicular lymphoma (FL). FL represents one of the most common non-Hodgkin lymphoma (NHL). About 75% of breakpoints on chromosome 18 are clustered in the major breakpoint region (MBR) and the minor cluster region (mcr), whereas the remaining breakpoints are scattered between these clusters, or at the 5’ side (variant cluster Ideograms of chromosomes 18 (above) and region or vcr) of the BCL2 gene. 14 (below) indicating the hybridization locations. The translocation t(14;18)(q32.3;q21.3) D18S507 REN11536 has been identified in about 80% of FLs 3’ 5’ SPEC BCL2/IGH Dual Color Dual Fusion Probe but is also observed in 20% to 30% of hybridized to normal interphase cells as indicated by mcr MBR vcr two orange and two green signals in each nucleus. patients with diffuse large B-cell lymphoma BCL2

(DLBCL). The rearrangement results in jux- ~1.3 Mb taposition of the BCL2 gene at 18q21.33 Cen 18q21.33 Tel next to the IGH (immunoglobulin heavy SPEC BCL2 Probe map (not to scale). chain) locus at 14q32.33 and leads to BV448316 RH119884 overexpression of the anti-apoptotic pro- 3’ 5’ 3’ 5’ tein BCL2. This represents most likely the JAG2 IGH initial step of malignant transformation, ~1.5 Mb leading to suppression of apoptosis and Cen 14q32.33 Tel progression to lymphoma. SPEC IGH Probe map (not to scale). Detection of t(14;18) by Fluorescence in situ Hybridization (FISH) can be used to confirm the diagnosis of FL if histology is References Baró C, et al. (2011) Leuk Res 35: 256-9. Bone marrow biopsy section with translocation inconclusive. Additionally, this method can Da Cunha Santos G, et al. (2011) Cancer Cytopathol 119: 254-62. affecting the BCL2/IGH loci as indicated by Einerson RR, et al. (2005) Am J Clin Pathol 124: 421-9. one separate orange signal, one separate green be used to monitor the response to therapy Gu K, et al. (2008) Arch Pathol Lab Med 132: 1355-61. signal, and two orange/green fusion signals. Nguyen-Khac F, et al. (2011) Am J Blood Res 1: 13-21. and detect recurrent disease. Weinberg OK, et al. (2007) J Mol Diagn 9: 530-7.

Prod. No. Product Label Tests* (Volume) Z-2114-50 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe / 5 (50 μl) Z-2114-200 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

153 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 153 FE042-1-20 ZytoLight ® SPEC MALT1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC MALT1 Dual Color The SPEC MALT1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 18q21.31-q21.32 translocations involving the chromosomal probes hybridizing to the 18q21.31- band, two orange/green fusion signals region 18q21.32 harboring the MALT1 q21.32 band. The green fluorochrome are expected representing two normal gene. The MALT1 (MALT1 paracaspase, direct labeled probe hybridizes proximal (non-rearranged) 18q21.31-q21.32 loci. a.k.a. MLT) gene encodes a human para- to the MALT1 gene at 18q21.31-q21.32, A signal pattern consisting of one orange/ caspase and is often rearranged in MALT and the orange fluorochrome direct labe- green fusion signal, one orange signal, lymphomas accounting for 5-10% of all led probe hybridizes distal to the MALT1 and a separate green signal indicates one B-cell non-Hodgkin lymphomas (NHL). The gene region at 18q21.32. normal 18q21.31-q21.32 locus and one most common translocations affecting the 18q21.31-q21.32 locus affected by a MALT1 gene are t(11;18)(q22.2;q21.3) translocation. and t(14;18)(q32.3;q21.3) occurring in 50% and 15-20% of MALT lymphomas, re- MALT1 spectively. These translocations lead to the expression of BIRC3-MALT1 (a.k.a. API2- Ideogram of chromosome 18 indicating the hybridization locations. MALT1) and IGH-MALT1 fusion proteins, resulting in constitutive activation of the D18S881 D18S1117 SHGC-154558 RH39250 NFκB signaling pathway which controls 5’ 3’ the expression of numerous anti-apoptotic and proliferation-promoting genes. MALT1 The translocation t(11;18)(q22.2;q21.3) ~640 kb ~430 kb Cen 18q21.31-q21.32 Tel is mainly found in pulmonary and SPEC MALT1 Probe map (not to scale). gastric lymphomas, whereas t(14;18) (q32.3;q21.3) occurs more frequently in SPEC MALT1 Dual Color Break Apart Probe hybridized to normal interphase cells as indicated by two non-gastrointestinal MALT lymphomas, orange/green fusion signals per nucleus. e.g., of the skin and salivary glands. The presence of a t(11;18)(q22.2;q21.3) correlates with unresponsiveness to erad- ication of Helicobacter pylori in gastric MALT lymphomas. Hence, detection of MALT1 translocations by Fluorescence in situ Hybridization (FISH) may be a supportive tool to identify patients eligible for anti H. pylori therapy.

References Afonina IS, et al. (2015) FEBS J 282: 3286-97. Beans M, et al. (2014) PloS One 9: e103774. Dierlamm J, et al. (1999) Blood 93: 3601-9. Levine EG, et al. (1989) Blood 74: 1796-800. Lucas PC, et al. (2001) J Biol Chem 276: 19012-9. Martinelli G, et al. (2005) J Clin Oncol 23: 1979-83. Pereira MI & Medeiros JA (2014) World J Gastroenterol 20: 684-98. Lymphoma tissue section with translocation Troppan K, et al. (2015) Gastroenterol Res Pract 2015: 102656. of the MALT1 gene as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2196-50 ZytoLight SPEC MALT1 Dual Color Break Apart Probe / 5 (50 μl) Z-2196-200 ZytoLight SPEC MALT1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

154 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 154 FE097-1-20 ZytoLight ® SPEC CIC Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CIC Dual Color The SPEC CIC Dual Color Break Apart In an interphase nucleus lacking a Break Apart Probe is designed to detect Probe is a mixture of two direct labeled translocation involving the 19q13.2 translocations involving the chromosomal probes hybridizing to the 19q13.2 band. band, two orange/green fusion signals region 19q13.2 harboring the CIC The orange fluorochrome direct labeled are expected representing two normal (capicua transcriptional repressor, a.k.a. probe hybridizes proximal and the green (non-rearranged) 19q13.2 loci. A signal KIAA0306) gene. The CIC gene encodes fluorochrome direct labeled probe hy- pattern consisting of one orange/green a transcriptional repressor that inhibits the bridizes distal to the CIC gene. fusion signal, one orange signal, and a expression of the PEA3 (polyoma enhancer separate green signal indicates one activator 3) gene family, including ETV1, normal 19q13.2 locus and one 19q13.2 ETV4, and ETV5 and regulates receptor locus affected by a translocation. tyrosine kinase signaling pathways. CIC Rearrangements involving the CIC gene Ideogram of chromosome 19 are frequently found in EWSR1-negative indicating the hybridization locations. small blue round cell tumors (SBRCT) which arise in soft tissues of children and young D19S688E RH39155 WI-5683 RH56006 adults and have been described as aggres- 5’3’ sive tumors with an inferior overall survival CIC compared to Ewing sarcoma (EWS). ~770 kb ~540 kb The CIC-DUX4 (double homeobox 4) gene Cen 19q13.2 Tel

fusion is the most frequent genetic event SPEC CIC Probe map (not to scale). in EWSR1-negative SBRCT resulting from either a t(4;19)(q35;q13.2) or a t(10;19)

(q26.3;q13.2). SPEC CIC Dual Color Break Apart Probe hybridized CIC rearrangements have also been to normal interphase cells as indicated by two orange/green fusion signals per nucleus. found in other tumor entities such as lung cancer and medulloblastomas and other fusion partners besides DUX4 are known (FOXO4, LEUTX). As CIC-DUX4-positive patients do not respond well to the common forms of EWS treatment, and show a poor overall survival, novel therapy approaches are needed to treat this type of aggressive tumors. Hence, the detection of CIC translocations by Fluorescence in situ Hybridization (FISH) may be of prognostic and therapeutic relevance. Undifferentiated round cell sarcoma ‚Ewing-like‘ tissue References section with translocation of the 19q13.2 locus as Antonescu R, et al. (2017) Am J Surg Pathol 7: 941-9. Choi K, et al. (2013) Am J Surg Pathol 9: 1379-86. indicated by one non-rearranged orange/green fusion Graham C, et al. (2012) Hum Pathol 2: 180-9. signal, one orange and one separate green signal. Italiano A, et al. (2012) Genes Chromosomes Cancer 3: 207-18. Kawamura-Saito M, et al. (2006) Hum Mol Genet 13: 2125-37. Lee CJ, et al. (2005) J Neurooncol 2: 101-8. Richkind KE, et al. (1996) Cancer Genet Cytogenet 1: 71-4.

Prod. No. Product Label Tests* (Volume) Z-2285-50 ZytoLight SPEC CIC Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

155 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 155 FE154-1-20 ZytoLight ® SPEC C19MC/19p13 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC C19MC/19p13 The SPEC C19MC/19p13 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of a green fluoro- ange and two green signals are expected. detection of amplifications of the C19MC chrome direct labeled SPEC C19MC In a cell with amplification of the C19MC microRNA (miRNA) cluster region located probe hybridizing to the C19MC region region, multiple copies of the green signal at 19q13.42. in the chromosomal region 19q13.42 and or green signal clusters will be observed. The C19MC miRNA cluster region an orange fluorochrome direct labeled comprises 46 micro RNA genes which SPEC 19p13 probe specific for 19p13.3. encode for large polycistronic microRNAs. Since chromosomes 1, 5, and 19 share C19MC amplifications have been report- the same repetitive sequences, probes spe- ed in tumor entities, such as breast cancer cific for 19p13.3 are commonly used for and parathyroid carcinomas. According to chromosome 19 copy number detection. the 2016 WHO classification of tumors of the central nervous system, amplification 19p13 of this region defines a new entity of em- bryonal tumors with multilayered rosettes C19MC (ETMR), C19MC-altered. Amplification miRNA cluster Ideogram of chromosome 19 of C19MC leads to an overexpression of indicating the hybridization locations. miRNAs in this cluster region that drive cell RH38185 STSG605919 SPEC C19MC/19p13 Dual Color Probe proliferation, promote cell survival, and hybridized to normal interphase cells as indicated increase carcinogenicity of cells providing by two orange and two green signals per nucleus C19MC miRNA cluster and to metaphase chromosomes of a normal cell. functional evidence that C19MC miRNAs ~615 kb act as oncogenes. Cen 19q13.42 Tel ETMR with C19MC miRNA cluster ampli- SPEC C19MC Probe map (not to scale). fication are rare brain tumors which occur primarily in infants and young children RH55727 RH120143 showing a poor overall survival. Hence, detection of C19MC miRNA cluster amplification by Fluorescencein situ ~485 kb Hybridization may be of diagnostic and Tel 19p13.3 Cen prognostic relevance. SPEC 19p13 Probe map (not to scale).

References Chhabda S, et al. (2016) Quant Imaging Med Surg 6: 486-9. Li M, et al. (2009) Cancer Cell 16: 533-46. Louis DN, et al. (2016) Acta Neuropathol 131: 803-20. Spence T, et al. (2014) Neuro Oncol 16: 62-71. Spence T, et al. (2014) Acta Neuropathol 128: 291-303. Primitive neuroectodermal tumor tissue section Vaira V, et al. (2012) J Mol Endocrinol 49: 115-24. with amplification of the C19MC miRNA cluster as indicated by multiple green signals.

Specimen kindly provided by Hannu Haapasalo, MD, PhD, Fimlab Laboratories, Finland.

Prod. No. Product Label Tests* (Volume) Z-2274-50 ZytoLight SPEC C19MC/19p13 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

156 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 156 FE147-1-20 ZytoLight ® SPEC BCL2L1/CEN 20 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC BCL2L1/CEN 20 The SPEC BCL2L1/CEN 20 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the de- Probe is a mixture of an orange fluoro- ange and two green signals are expected. tection of BCL2L1 gene amplifications. chrome direct labeled SPEC BCL2L1 probe In a cell with amplification of the BCL2L1 The BCL2L1 (BCL2-like 1, a.k.a. BCLX) hybridizing to the BCL2L1 gene in the gene locus, multiple copies of the orange gene is located in the chromosomal region chromosomal region 20q11.21 and a signal or orange signal clusters will be 20q11.21 and encodes for an anti-ap- green fluorochrome direct labeled CEN observed. optotic protein that belongs to the BCL2 20 probe specific for the alpha satellite family. These genes are involved in a wide centromeric region of chromosome 20 variety of cellular activities including lym- (D20Z2). phocyte development and hematopoiesis. BCL2L1 amplifications have been reported CEN 20 (D20Z2) in several human cancers including lung, BCL2L1 ovarian breast, melanoma, and hematologic malignancies. Ideogram of chromosome 20 Overexpression of BCL2L1 reduces indicating the hybridization locations. MYC-induced apoptosis in immortalized D20S484 D20S425 bronchial epithelial cells. 3’ 5’ Furthermore, BCL2L1 amplifications BCL2L1 are found in many tumor cell lines with SPEC BCL2L1/CEN 20 Dual Color Probe hybridized ~485 kb resistance to chemotherapeutic agents. to normal interphase cells as indicated by two Cen 20q11.21 Tel orange and two green signals in each nucleus. Targeting the BCL2 family proteins with SPEC BCL2L1 Probe map (not to scale). small non-peptidic compounds, so called BH3-mimetics, is currently investigated in clinical trials. Hence, the identification of BCL2L1 amplifications by Fluorescencein situ Hybridiza- tion and the inhibition of BCL2L1 signaling may be of therapeutic significance in various types of tumors.

References Beroukhim R, et al. (2010) Nature 463: 899-905. Booher RN, et al. (2014) PloS One 9: e108371. Sochalska M, et al. (2015) FEBS J 282: 834-49. Yasui K, et al. (2004) Cancer Res 64: 1403-10.

SK-LU-1 cell line with interphase cells showing amplification of the BCL2L1 gene locus as indicated by orange signal clusters in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2171-200 ZytoLight SPEC BCL2L1/CEN 20 Dual Color Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

157 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 157 FE106-1-20 ZytoLight ® SPEC PTPRT/20q11 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC PTPRT/20q11 Dual The SPEC PTPRT/20q11 Dual Color Probe In a normal interphase nucleus, two or- Color Probe is designed to detect deletions is a mixture of a green fluorochrome ange and two green signals are expect- of the long arm of chromosome 20. direct labeled SPEC PTPRT probe hybrid- ed. In a cell with deletions affecting the 20q deletions can occur in various myeloid izing in the CDR at 20q12, an orange 20q12 locus, one or no copy of the green disorders, e.g., myelodysplastic syndromes fluorochrome direct labeled SPEC 20q11 signal will be observed. (MDS), acute myeloid leukemia (AML), probe specific for the chromosomal region and myeloproliferative neoplasms (MPNs). 20q11.21 harboring the DUSP15 gene. In MDS, del(20q) as the sole cytogenetic For an unambigous enumeration of chro- abnormality, is associated with a favorable mosome 20, the SPEC 20q11 is found to prognosis, with better survival, and a lower be suitable. risk for transformation to AML. Del(20q) occurring with additional cytogenetic aberrations predicts a poor prognosis. 20q11 PTPRT The breakpoints of the 20q deletion were

identified to be heterogeneous in several Ideogram of chromosome 20 studies. The minimal common deleted indicating the hybridization locations.

region (CDR) was defined to be flanked RH120697 SHGC-148173 3’ 5’ by the genes PTPRT (20q12) and EYA2 SPEC PTPRT/20q11 Dual Color Probe hybridized (20q13.12). to normal interphase cells as indicated by two PTPRT orange and two green signals in each nucleus. It has been suggested that one or more ~590 kb tumor suppressor genes could be located Cen 20q12 Tel

in the CDR, the deletion or inactivation SPEC PTPRT Probe map (not to scale). of which may play a role in malignant D17S2108 D20S874 growth. However, the target gene(s) remain 3’ 5’ unknown. DUSP15 References Bacher U, et al. (2014) Br J Haematol 164: 822-33. ~420 kb Brezinová J, et al. (2005) Cancer Genet Cytogenet 160: 188-92. Okada M, et al. (2012) Cancer Genet 205: 18-24. Cen 20q11.21 Tel Testa JR, et al. (1978) Blood 52: 868-77. SPEC 20q11 Probe map (not to scale).

Lymphocytes of a myelodysplastic syndrome showing a 20q deletion indicated by one single green and two orange signals in each nucleus.

Material kindly provided from Dr. Saurabh Bhattacharya, Lal PathLabs, India.

Prod. No. Product Label Tests* (Volume) Z-2213-50 ZytoLight SPEC PTPRT/20q11 Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

158 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 158 FE129-1-20 ZytoLight ® SPEC ERG Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC ERG Dual Color The SPEC ERG Dual Color Break Apart In an interphase nucleus of a normal Break Apart Probe is designed to detect Probe is a mixture of two direct labeled cell lacking an aberration involving the aberrations involving the ERG gene at probes hybridizing to the long arm of 21q22.13-q22.2 band, two orange/ 21q22.2 frequently detected in prostate chromosome 21. The orange fluoro- green fusion signals are expected repre- cancers. chrome direct labeled probe hybridizes senting the two normal (non-rearranged) ERG (ETS transcription factor ERG) at 21q22.13-q22.2 proximal to the ERG 21q22.13-q22.2 loci. rearrangements have been observed in gene breakpoint region, the green direct One 21q22.13-q22.2 locus affected 40-60% of prostate cancers identified via labeled probe hybridizes at 21q22.2 by a 21q22.2 deletion resulting in the prostate-specific antigen (PSA) screening. distal to the ERG gene breakpoint region. TMPRSS2-ERG fusion is indicated by the The most common aberration affecting loss of one green signal. ERG is the interstitial deletion of about A signal pattern consisting of one orange/ 3 Mb at the chromosomal region 21q22 ERG green fusion signal, a separate green, found in 90% of the cases. This deletion and a separate orange signal indicates an leads to the fusion of the hormonally Ideogram of chromosome 21 ERG translocation without involvement of indicating the hybridization locations. regulated promoter of the TMPRSS2 TMPRSS2 (e.g. SLC45A3-ERG). (transmembrane serine protease 2) gene RH118947 RH119925 RH119817 SHGC-106900 to the coding region of ERG, resulting in 3’ 5’ overexpression of the ERG transcription ERG factor. However, about 10% of the ERG ~560 kb ~775 kb rearranged prostate cancer cases show Cen 21q22.13-q22.2 Tel alternative fusions, as e.g. SLC45A3-ERG SPEC ERG Probe map (not to scale). or NDRG1-ERG. Several studies detected associations of ERG rearrangements with histomorphologic features as well as characteristic chromosomal copy number changes and gene expression signatures, defining SPEC ERG Dual Color Break Apart Probe a distinct sub-class of prostate cancers hybridized to normal interphase cells as indicated with unfavorable prognosis. Hence, the by two orange/green fusion signals per nucleus. evaluation of the ERG rearrangement status in tissue or urine samples by FISH might be of diagnostic and prognostic relevance. EWSR1-ERG gene fusions present in about 10% of patients with Ewing sarcoma may result from complex genomic rearrangements and may therefore not be detected by FISH analysis or may result in a non-classical translocation signal pattern.

References Esgueva R, et al. (2010) Mod Pathol 23: 539-46. Maire G, et al. (2008) Cancer Genet Cytogenet 181: 81-92. Prostate cancer tissue section with interstitial Nam RK, et al. (2007) Br J Cancer 97: 1690-5. Perner S, et al. (2006) Cancer Res 66: 8337-41. deletion of the chromosomal region 21q22.2 Pflueger D, et al. (2009) Neoplasia 11: 804-11. resulting in the TMPRSS2-ERG fusion as Tomlins SA, et al. (2005) Science 310: 644-8. indicated by the loss of one green signal.

Prod. No. Product Label Tests* (Volume) Z-2138-200 ZytoLight SPEC ERG Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

159 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 159 FE063-1-20 ZytoLight ® SPEC ERG/TMPRSS2 TriCheck™ Probe

Background Probe Description An ERG translocation without involvement The ZytoLight ® SPEC ERG/TMPRSS2 The SPEC ERG/TMPRSS2 TriCheck™ Probe of TMPRSS2 is indicated by a separated TriCheck™ Probe is designed to detect is a mixture of three direct labeled probes orange signal and a blue signal co-localiz- deletions between the ERG and the hybridizing to the chromosomal regions ing with the separate green signal. TMPRSS2 gene at 21q22 resulting in the 21q22.13-q22.3. The orange A non-ERG translocation affecting TMPRSS2-ERG fusion. Furthermore, the fluorochrome direct labeled probe hybrid- TMPRSS2 is indicated by a separated triple color approach allows the detection izes at 21q22.13-q22.2 proximal to the blue signal not co-localizing with the ERG of other translocations affecting either of ERG gene breakpoint region, the green fusion signal. these genes. fluorochrome direct labeled probe hy- ERG (ETS transcription factor ERG) bridizes at 21q22.2 distal to that region, rearrangements have been observed in and the blue fluorochrome direct labeled 40-60% of prostate cancers identified via probe hybridizes at 21q22.3 distal to the prostate-specific antigen (PSA) screening. TMPRSS2 gene region. The most common aberration affecting ERG is the interstitial deletion of about 3 Mb at the chromosomal region 21q22 found in ERG 90% of the cases. This deletion leads to the TMPRSS2 fusion of the hormonally regulated promot- Ideogram of chromosome 21 er of the TMPRSS2 (transmembrane serine indicating the hybridization locations.

protease 2) gene to the coding region of RH118947 RH119925 RH121883 ERG, resulting in overexpression of the ERG RH119817 SHGC-106900 D21S113 SPEC ERG/TMPRSS2 TriCheck™ Probe hybridized to 3’ 5’ 3’5’ transcription factor. The deleted fragment is normal interphase cells as indicated by two orange/ green fusion signals and two blue signals in close sometimes observed as insertion on other ERG TMPRSS2 proximity of the respective fusion signals. chromosomes. However, about 10% of ~560 kb ~775 kb ~895 kb Cen 21q22.13-q22.3 Tel the ERG rearranged prostate cancer cases TM show alternative fusions, as e.g. SPEC ERG/TMPRSS2 TriCheck Probe map (not to scale). SLC45A3-ERG. On the other hand non-ERG translocations fusing TMPRSS2 to other ETS family members, as e.g. TMPRSS2-ETV1, have been found in a few percent of these malignancies. Results Several studies detected associations of ERG In a normal interphase nucleus, two rearrangements with histomorphologic fea- orange/green fusion signals and two tures as well as characteristic chromosomal blue signals in close proximity of the copy number changes and gene expression respective fusion signals are expected signatures defining a distinct sub-class of representing two normal (non-rearranged) Prostate cancer tissue section with one 21q22 locus affected by an interstitial deletion of the prostate cancers with unfavorable prognosis. 21q22.13-q22.3 loci. chromosomal region 21q22.2 resulting in the Hence, the evaluation of the TMPRSS2-ERG One 21q22 locus affected by a 21q22.2 TMPRSS2-ERG fusion as indicated by one separate orange signal co-localizing with one rearrangement status in tissue or urine deletion resulting in the TMPRSS2-ERG fu- blue signal, and the loss of one green signal. samples by FISH might be of diagnostic and sion is indicated by one separate orange prognostic relevance. signal co-localizing with one blue signal, References and the loss of one green signal. Esgueva R, et al. (2010) Mod Pathol 23: 539-46. Nam RK, et al. (2007) Br J Cancer 97: 1690-5. Perner S, et al. (2006) Cancer Res 66: 8337-41. Tomlins SA, et al. (2005) Science 310: 644-8.

Prod. No. Product Label Tests* (Volume) Z-2135-200 ZytoLight SPEC ERG/TMPRSS2 TriCheck Probe •/•/• 20 (200 μl) Related Products Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

160 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 160 FE062-1-20 ZytoLight ® SPEC DiGeorge/Phelan McDermid Dual Color Probe

Background global developmental delay, moderate to Results The ZytoLight ® SPEC DiGeorge/Phelan severe intellectual impairment, absent or In a normal interphase nucleus, two or- McDermid Dual Color Probe is designed severely delayed speech, and neonatal ange and two green signals are expected. to detect deletions affecting the chromo- hypotonia. In a cell with deletion of the HIRA gene lo- somal regions 22q11.21 harboring the cus, a reduced number of orange signals HIRA (a.k.a. TUPLE1) gene and 22q13.33 Probe Description will be observed. In a cell with deletion of harboring the SHANK3 (a.k.a. prosap2) The SPEC DiGeorge/Phelan McDer- the SHANK3 gene locus, a reduced num- gene, respectively.. mid Dual Color Probe is a mixture of ber of green signals will be observed. The 22q11.2 deletion syndrome an orange fluorochrome direct labeled (22q11.2DS), also known as velocardi- probe spanning the HIRA gene region at ofacial syndrome (VCFS) and DiGeorge 22q11.21 and a green fluorochrome di- syndrome, is a genetic disorder caused rect labeled probe spanning the SHANK3 by hemizygous microdeletions on chromo- gene region at 22q13.33. some 22q11.2, with population prevalence of about 1 to 4,000 births. The characteristic phenotype of 22q11.2DS HIRA includes cardiac defects, velopharyngeal SHANK3 insufficiency, immune deficiency due to Ideogram of chromosome 22 indicating the hybridization locations. thymic aplasia, growth restriction, and deficits in cognitive abilities. D22S1567 D22S1664 3’ 5’ The 22q11.2 deletion usually occurs by meiotic non-allelic homologous HIRA Lymphocytes and metaphase chromosomes from a DiGeorge syndrome case showing a HIRA deletion as recombination events between low copy ~315 kb indicated by the loss of one green signal. repeats on chromosome 22q11.2 termed Cen 22q11.21 Tel Kindly provided by Dr. Liehr, Jena, Germany. LCR22. There are eight LCR22s that span SPEC HIRA Probe map (not to scale). the 22q11.2 region termed LCR22A RH93825 RH38881 through LCR22H. The majority (90%) of 5’ 3’ 22q11.2DS patients show a recurrent SHANK3 3 Mb deletion between LCR22A and ~265 kb LCR22D harboring the HIRA gene. Cen 22q13.33 Tel

The 22q13.3 deletion syndrome SPEC SHANK3 Probe map (not to scale). (Phelan-McDermid syndrome) typically results from deletions of 100 kb to 9 Mb involving the distal long arm of chromosome 22. Almost all of these deletions include the gene SHANK3 that encodes a scaffold protein in the postsynaptic den- sities of excitatory synapses, connecting References Lymphocytes and metaphase chromosomes from a membrane-bound receptors to the actin cy- Burnside RD (2015) Cytogenet Genome Res 146: 89-99. Phelan-McDermid syndrome case showing a SHANK3 Morrow BE, et al. (2018) Am J Med Genet A 176: 2070-81. deletion as indicated by the loss of one orange signal. toskeleton. This syndrome is characterized Phelan K & McDermid HE (2012) Mol Syndromol 2: 186-201. Scambler PJ, et al. (1991) Genomics 10: 201-6. by neurological deficits, which include Watt JL, et al. (1985) J Med Genet 22: 283-7. Kindly provided by Dr. Kazmierczak, Bremen, Germany.

Prod. No. Product Label Tests* (Volume) Z-2299-50 ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

161 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 161 FE164-1-20 ZytoLight ® SPEC DiGeorge Triple Color Probe

Background Various deletions of more distal regions Results The ZytoLight ® SPEC DiGeorge Triple (between LCR22D-H) have also been In a normal interphase nucleus, two blue, Color Probe is designed to detect dele- reported and result in phenotypic features two orange, and two green signals are ex- tions affecting the chromosomal regions similar to 22q11.2DS. FISH probes target- pected. In a cell with deletion of the HIRA 22q11.21 harboring the genes HIRA ing MAPK1, which maps to the LCR22D-E and/or the CRKL gene locus, a reduced (a.k.a. TUPLE1) and CRKL as well as region, can be used for the detection of number of blue and/or orange signals 22q11.21-q11.22 harboring the MAPK1 this 22q11.2 distal deletion syndrome. will be observed, respectively. In a cell (a.k.a. PRKM2, ERK) gene. with deletion of the MAPK1 gene locus, a The 22q11.2 deletion syndrome Probe Description reduced number of green signals will be (22q11.2DS), also known as velocardi- The SPEC DiGeorge Triple Color Probe observed. ofacial syndrome (VCFS) and DiGeorge is a mixture of a blue fluorochrome direct syndrome, is a genetic disorder caused by labeled probe spanning the HIRA gene hemizygous microdeletions on region at 22q11.21, an orange fluoro- chromosome 22q11.2, with population chrome direct labeled probe spanning the prevalence of about 1 in 4,000 births. The CRKL gene region at 22q11.21, and a characteristic phenotype of 22q11.2DS in- green fluorochrome direct labeled probe cludes cardiac defects, immune deficiency, spanning the MAPK1 gene region at growth restriction, and deficits in cognitive 22q11.21-q11.22. The MAPK1 targeting abilities. probe additionally serves as a reference The 22q11.2 deletion usually occurs by in case of a deletion of the HIRA and meiotic non-allelic homologous recombi- CRKL gene loci (LCR22A-D). nation events between low copy repeats on chromosome 22q11.2 termed LCR22. HIRA SPEC DiGeorge Triple Color Probe hybridized to CRKL normal interphase cells as indicated by two orange, There are eight LCR22s that span the MAPK1 two green, and two blue signals and to metaphase 22q11.2 region termed LCR22A chromosomes of a normal cell. Ideogram of chromosome 22 through LCR22H. The majority (90%) of indicating the hybridization locations. 22q11.2DS patients show a recurrent D22S947 D22S1662 RH81944 D22S935 3 Mb deletion between LCR22A and 3’ 5’ 5’ 3’ 5’ 3’ LCR22D while 8% harbor a nested HIRA TBX1 CRKL 1.5 Mb deletion (LCR22A-B). Some rare ~865 kb ~355 kb atypical deletions of shorter size and Cen 22q11.21 Tel

in variable locations have also been SPEC HIRA/SPEC CRKL Probe map (not to scale). reported (e.g., LCR22B-D and LCR22C-D). RH111612 SHGC-131036 Classic FISH probes for the detection 3’ 5’ of 22q11.2DS target the HIRA gene MAPK1 mapping to the LCR22A-B region, and ~655 kb thus, miss deletions that occur outside this Cen 22q11.21-q11.22 Tel Lymphocytes and metaphase chromosomes region. The DiGeorge Triple Color Probe from a DiGeorge syndrome case showing a SPEC MAPK1 Probe map (not to scale). HIRA/CRKL deletion as indicated by the loss additionally targets CRKL that maps to the References Ben-Shachar S, et al. (2008) Am J Hum Genet 82: 214-21. of one blue and one orange signal. LCR22C-D region allowing the detection Burnside RD (2015) Cytogenet Genome Res 146: 89-99. Michaelovsky E, et al. (2012) BMC Med Genet 13: 122. Kindly provided by Dr. Liehr, Jena, Germany. of rare deletions. Morrow BE, et al. (2018) Am J Med Genet A 176: 2070-81. Scambler PJ, et al. (1991) Genomics 10: 201-6.

Prod. No. Product Label Tests* (Volume) Z-2289-50 ZytoLight SPEC DiGeorge Triple Color Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

162 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 162 FE160-1-20 ® ZytoLight SPEC IGL Dual Color Break Apart ProbeHome saltwater

Background Probe Description Results The ZytoLight ® SPEC IGL Dual Color The SPEC IGL Dual Color Break In an interphase nucleus lacking a Break Apart Probe is designed to detect Apart Probe is a mixture of a green translocation involving the IGL locus at rearrangements affecting the chromosomal fluorochrome direct labeled probe 22q11.22, two orange/green fusion region 22q11.22 harboring the IGL hybridizing proximal to the IGL breakpoint signals are expected. A signal pattern (immunoglobulin lambda locus, a.k.a., IGλ) region at 22q11.21-q11.22 and an consisting of one orange/green fusion gene cluster region. orange fluorochrome direct labeled probe signal, one orange signal, and a separate Translocations involving the immunoglobulin hybridizing distal to the IGL breakpoint green signal indicates one normal IGL (IG) genes are recurring events of B-cell region at 22q11.22-q11.23. locus and one IGL locus affected by a oncogenesis. In all of these translocations, translocation. an oncogene is activated and overexpressed by juxtaposing this oncogene to IG IGL regulatory sequences. Burkitt lymphoma (BL) is characterized by Ideogram of chromosome 22 indicating the hybridization locations. reciprocal translocations involving the MYC

gene and one of the IG loci. The majority of RH48178 SHGC-145319 translocations involve the immunoglobulin RH111612 D22S1673 5’ 3’ heavy chain (IGH) locus while a minor part involves the immunoglobulin light chain IGL loci, either the kappa light chain (IGK) or ~1 Mb ~355 kb the lambda light chain (IGL). IGK and IGL Cen 22q11.21-q11.23 Tel rearrangements resulting from the variant SPEC IGL Probe map (not to scale).

translocations t(2;8)(p11.2;q24.21) and SPEC IGL Dual Color Break Apart Probe hybridized t(8;22)(q24.21;q11.2), respectively, have to normal interphase cells as indicated by two orange/green fusion signals in each nucleus been detected in up to 25% of BL cases. and to metaphase chromosomes of a normal cell. In non-Hodgkin lymphoma (NHL) harboring IG-MYC rearrangements, the MYC translocation partner is IGK and IGL in 8 and 22% of the cases, respectively. IG translocations have been reported in several B-cell lineage malignancies other than BL including atypical Burkitt/Burkitt-like lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, mantle cell lymphoma, and multiple myeloma. The detection of IGK and IGL involvement in lymphomas by Fluorescence in situ Hybridization may prove a valuable diagnostic and prognostic tool. Cell line with an IGL translocation affecting the References 22q11.21-q11.23 locus as indicated by one Cario G, et al. (2000) Br J Haematol 110: 537-46. Einerson RR, et al. (2006) Leukemia 20: 1790-9. non-rearranged orange/green fusion signal, one Martín-Subero JI, et al. (2002) Int J Cancer 98: 470-4. orange signal, and one separate green signal. Poulsen TS, et al. (2002) Leukemia 16: 2148-55.

Prod. No. Product Label Tests* (Volume) Z-2286-50 ZytoLight SPEC IGL Dual Color Break Apart Probe •/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

163 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 163 FE155-1-20 ZytoLight ® SPEC SMARCB1/22q12 Dual Color Probe

Background Probe Description Results The ZytoLight ® SPEC SMARCB1/22q12 The SPEC SMARCB1/22q12 Dual Color In a normal interphase nucleus, two or- Dual Color Probe is designed for the Probe is a mixture of a green fluorochrome ange and two green signals are expected. detection of deletions of the chromosomal direct labeled SPEC SMARCB1 probe In a cell with deletion of the SMARCB1 region harboring the SMARCB1 gene. hybridizing to the human SMARCB1 gene gene locus, a reduced number of green The SMARCB1 (SWI/SNF related, matrix in the chromosomal region 22q11.23 signals will be observed. Deletions af- associated, actin dependent regulator of and an orange fluorochrome direct fecting only parts of the SMARCB1 gene chromatin, subfamily b, member 1, a.k.a. labeled SPEC 22q12 probe specific for might result in a normal signal pattern with INI1, SNF5, or BAF47) gene is located on the KREMEN1 (kringle containing trans- green signals of reduced size. chromosome 22q11.23 and encodes a membrane protein 1) gene region in tumor suppressor. 22q12.1-q12.2. Due to cross-hybridiza- Rhabdoid tumors are highly malignant tions of chromosome 22 alpha satellites to neoplasms that typically arise in infancy other centromeric regions, probes specific and early childhood. They are classified for 22q12 are frequently used for chromo- as atypical teratoid/rhabdoid tumors (AT/ some 22 copy number detection. RT) when they occur in the CNS or as malignant rhabdoid tumors (MRT) when SMARCB1 they are found in renal or extra-renal sites. 22q12 The vast majority of AT/RTs and MRTs are characterized by loss of function of Ideogram of chromosome 22 indicating the hybridization locations. the SMARCB1 gene due to deletions or

mutations. The molecular alterations are REN35238 D22S1259 SPEC SMARCB1/22q12 Dual Color Probe often bi-allelic resulting in complete loss 5’ 3’ hybridized to normal interphase cells as indicated by two orange and two green signals per nucleus. of this tumor suppressor gene, and thus in SMARCB1

cell cycle progression. ~545 kb Patients with germline alterations of Cen 22q11.23 Tel SMARCB1, including deletions, duplica- SPEC SMARCB1 Probe map (not to scale). tions, and mutations, were found to be SHGC-17392 RH76665 predisposed to malignant rhabdoid tumors 5’ 3’ and schwannomatosis. KREMEN1 Moreover, deletions of the SMARCB1 ~335 kb gene were found to occur in patients with Cen 22q12.1-q12.2 Tel highly aggressive renal medullary carci- SPEC 22q12 Probe map (not to scale). noma (RMC), epithelioid sarcoma, and poorly differentiated sarcoma. The identification of SMARCB1 deletions SPEC SMARCB1/22q12 Dual Color Probe by FISH may represent a powerful adjunc- hybridized to epithelioid sarcoma tissue section tive diagnostic tool useful in the differential with biallelic deletion of the SMARCB1 gene as indicated by missing green signals in the nuclei. diagnosis of rhabdoid tumors. Moreover, References prenatal testing should be performed in Calderaro J, et al. (2012) Histopathology 61: 428-35. Eaton KW, et al. (2011) Pediatr Blood Cancer 56: 7-15. situations where alterations of SMARCB1 Mobley BC, et al. (2010) Acta Neuropathol 120: 745-53. Roberts CW & Biegel JA (2009) Cancer Biol Ther 8: 412-6. have been documented in the family. Sullivan LM, et al. (2013) Mod Pathol 26: 385-92.

Prod. No. Product Label Tests* (Volume) Z-2178-50 ZytoLight SPEC SMARCB1/22q12 Dual Color Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

164 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 164 FE087-1-20 ZytoLight ® SPEC EWSR1 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC EWSR1 Dual Color The SPEC EWSR1 Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled location involving the 22q12.1-q12.2 translocations involving the chromosomal probes hybridizing to the 22q12.1-q12.2 band two orange/green fusion signals region 22q12.2 harboring the EWSR1 band. The orange fluorochrome direct are expected representing two normal (EWS RNA binding protein 1, a.k.a. labeled probe hybridizes proximal and ex- (non-rearranged) 22q12.1-q12.2 loci. A EWS) gene. tends inward into intron 4 of the EWSR1 signal pattern consisting of one orange/ Translocations involving the chromosomal gene, the green fluorochrome direct labe- green fusion signal, one orange signal, region 22q12.2 are found in 90-95% of led probe hybridizes distal to that gene. and a separate green signal indicates patients with Ewing sarcoma or peripheral one normal 22q12.1-q12.2 locus and primitive neuroectodermal tumors (PNET). one 22q12.1-q12.2 locus affected by a Ewing sarcoma is the second most com- 22q12.1-q12.2 translocation. mon, highly malignant bone tumor in children and young adults. The most frequent Ideogram of chromosome 22 translocation involving the EWSR1 gene indicating the hybridization locations.

region is t(11;22)(q24.3;q12.2) juxtapos- RH76665 RH122132 ing the EWSR1 gene in 22q12.2 next to RH103903 RH75124 5’ 3’ the FLI-1 (friend leukemia virus integration 1) locus in 11q24.3. FLI-1 is a member of EWSR1 the ETS family of transcription factors. Less ~480 kb ~400 kb Cen Tel frequently, EWSR1 can also be fused to 22q12.1-q12.2 ERG, a transcription factor closely related SPEC EWSR1 Probe map (not to scale). to FLI-1 but located in 21q22.2. For prognosis and appropriate treatment

it is important to differentiate Ewing sar- SPEC EWSR1 Dual Color Break Apart Probe coma/PNET from classic neuroblastoma, hybridized to normal interphase cells as indicated by two orange/green fusion signals per nucleus. Wilms tumor, and rhabdomyosarcoma. In combination with the histopathological diagnosis, detection of the EWSR1 rearrangements by FISH can be used to confirm the diagnosis of Ewing sarcoma/ PNET.

References Bridge RS, et al. (2006) Mod Pathol 19: 1-8. Delattre O, et al. (1992) Nature 359: 162-5. Lee J, et al. (2005) Cancer Genet Cytogenet 159: 177-80. Rekhi B, et al. (2012) Virchows Arch 461: 687-97. Romeo S & Dei Tos AP (2010) Virchows Arch 456: 219-34. Sandberg AA & Bridge JA (2000) Cancer Genet Cytogenet 123: 1-26. Yang L, et al. (2012) Hum Pathol 43: 1463-70. Zucman J, et al. (1993) EMBO J 12: 4481-7.

Ewing sarcoma tissue section with translocation affecting the 22q12.1-q12.2 locus as indicated by one non-rearranged orange/green fusion signal, one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2096-50 ZytoLight SPEC EWSR1 Dual Color Break Apart Probe / 5 (50 μl) Z-2096-200 ZytoLight SPEC EWSR1 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

165 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 165 FE031-1-20 ZytoLight ® SPEC EWSR1/FLI1 TriCheck™ Probe

Background Probe Description Results The ZytoLight ® SPEC EWSR1/FLI1 The SPEC EWSR1/FLI1 TriCheck™ Probe In an interphase nucleus without TriCheck™ Probe is designed to detect trans- is a mixture of three direct labeled probes FLI1-EWSR1 rearrangement, two green/ locations involving the chromosomal region hybridizing to the 22q12.1-q12.2 and orange fusion signals and two blue signals 22q12.2 harboring the EWSR1 (EWS RNA 11q24.3 bands. The orange fluorochrome are expected. binding protein 1, a.k.a. EWS) gene and direct labeled probe hybridizes proxi- A FLI1-EWSR1 fusion is indicated by one the chromosomal region 11q24.3 har- mal to the EWSR1 breakpoint region at separate orange signal co-localizing with boring the FLI1 (Fli-1 proto-oncogene, ETS 22q12.1-q12.2 and the green fluoro- one blue signal and one separate green transcription factor, a.k.a. EWSR2) gene. chrome direct labeled probe hybridizes signal. Translocations involving the chromosomal distal to the EWSR1 breakpoint region at An EWSR1 translocation without involve- region 22q12.2 are found in 90-95% of 22q12.2. The blue fluorochrome direct ment of FLI1 is indicated by the split of patients with Ewing sarcoma or peripheral labeled probe hybridizes distal to the FLI1 one green/orange fusion signal without primitive neuroectodermal tumors (PNET). gene at 11q24.3. co-localization of the separated orange Ewing sarcoma is the second most com- signal with one blue signal. mon, highly malignant bone tumor in children and young adults. The most frequent EWSR1 translocation involving the EWSR1 gene region is t(11;22)(q24.3;q12.2) juxtaposing the EWSR1 gene in 22q12.2 next to the FLI1 locus. FLI1 is a member of the ETS family of transcription factors. Less frequently, EWSR1 can also be fused to ERG, a transcription factor closely related to FLI1 but located in 21q22.2. For prognosis and appropriate treatment it FLI1 is important to differentiate Ewing sarco- Ideograms of chromosomes 22 (above) and 11 (below) indicating the hybridization locations. ma/PNET from classic neuroblastoma, Ewing sarcoma tissue section with FLI1-EWSR1 fusion as indicated by orange/blue fusion signals. Wilms tumor, and rhabdomyosarcoma. RH76665 RH122132 In combination with the histopathological RH103903 RH75124 5’ 3’ diagnosis, detection of the EWSR1 rearrangements by FISH can be used to confirm EWSR1 the diagnosis of Ewing sarcoma/PNET. ~480 kb ~400 kb Cen 22q12.1-q12.2 Tel References Bridge RS, et al. (2006) Mod Pathol 19: 1-8. SPEC EWSR1 Probe map (not to scale). Delattre O, et al. (1992) Nature 359: 162-5. Lee J, et al. (2005) Cancer Genet Cytogenet 159: 177-80. Romeo S & Dei Tos AP (2010) Virchows Arch 456: 219-34. Sandberg AA & Bridge JA (2000) Cancer Genet Cytogenet 123: 1-26. D11S4612 D11S3381 Zucman J, et al. (1993) EMBO J 12: 4481-7. 5’ 3’

FLI1

~640 kb Cen 11q24.3 Tel

SPEC FLI1 Probe map (not to scale). Ewing sarcoma tissue section with a non-FLI1 EWSR1 rearrangement as indicated by the lack of co-localization of the separated orange signal with one blue signal.

Prod. No. Product Label Tests* (Volume) Z-2183-50 ZytoLight SPEC EWSR1/FLI1 TriCheck Probe •/•/• 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

166 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 166 FE115-1-20 ZytoLight ® SPEC PDGFB Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC PDGFB Dual Color The SPEC PDGFB Dual Color Break Apart In an interphase nucleus lacking a trans- Break Apart Probe is designed for the de- Probe is a mixture of two direct labeled location involving the 22q13.1 band two tection of specific translocations involving probes hybridizing to the 22q13.1 band. orange/green fusion signals are expected the chromosomal region 22q13.1 harbor- The orange fluorochrome direct labeled representing two normal (non-rearranged) ing the PDGFB (a.k.a. SIS) gene. probe hybridizes proximal to the break- 22q13.1 loci. A signal pattern consisting The PDGFB gene (platelet derived growth point region of the PDGFB gene, and the of one orange/green fusion signal, one factor receptor beta) belongs to the green fluorochrome direct labeled probe orange signal, and a separate green platelet-derived growth factor family and hybridizes distal to the breakpoint region signal indicates one normal 22q13.1 encodes a protein which acts as a recep- of the PDGFB gene. locus and one 22q13.1 locus affected by tor tyrosine kinase. a 22q13.1 translocation. The most frequent translocation involving the PDGFB gene is t(17;22)(q21.3;q13.1) juxtaposing the PDGFB gene next to the COL1A1 gene in 17q22. Reciprocal trans- Ideogram of chromosome 22 indicating the hybridization locations. locations involving t(17;22)(q21.3;q13.1) are characteristic for dermatofibrosarcoma RH109956 D22S1155 STDJ358M15T7 D22S575 protuberans (DFSP) patients. DFSP is a 3’ 5’ highly recurrent, infiltrative skin tumor of intermediate malignancy. PDGFB The rearrangements are cytogenetically ~595 kb ~545 kb Cen 22q13.1 Tel characterized by the presence of super- SPEC PDGFB Probe map (not to scale). numerary ring chromosomes containing SPEC PDGFB Dual Color Break Apart Probe hybridi- low-level amplified sequences from chro- zed to normal interphase cells as indicated by two orange/green fusion signals per nucleus. mosomes 17q22-qter and 22q10-q13.1, or unbalanced derivatives of the t(17;22) (q21.3;q13.1) translocation. The rearrangement results in a COL1A1- PDGFB fusion protein which is post-transcriptionally processed to a functional platelet-derived growth factor beta chain (PDGFB) protein. The importance of accurately diagnosing DFSP lies in its intermediate malignant

nature and the availability of a therapy References Broom RJ, et al. (2012) Clin Genitourin Cancer 10: 202-6. with significant anti-neoplastic activity but Labropoulos SV & Razis ED (2007) Biologics 4: 347-53. Dermatofibrosarcoma protuberans tissue section with Patel KU, et al. (2008) Human Pathol 39: 184-93. relatively minor adverse effects for cases Shimizu A, et al. (1999) Cancer Res 59: 3719-23. translocation affecting the 22q13.1 locus as indicated Simon MP, et al. (1997) Nat Genet 15: 95-8. by one non-rearranged orange/green fusion signal, not amenable to surgical excision. Sirvent N, et al. (2003) Genes Chromosomes Cancer 37: 1-19. one orange signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2119-50 ZytoLight SPEC PDGFB Dual Color Break Apart Probe / 5 (50 μl) Z-2119-200 ZytoLight SPEC PDGFB Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

167 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 167 FE040-1-20 ZytoLight ® SPEC CRLF2 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC CRLF2 Dual Color The SPEC CRLF2 Dual Color Break Apart In an interphase nucleus of a normal fe- Break Apart Probe is designed to detect Probe is a mixture of two direct labeled male cell lacking a translocation involving rearrangements involving the chromosom- probes hybridizing to the Xp22.33 and the Xp22.33 band, two orange/green al regions Xp22.33 and Yp11.32 harbor- Yp11.32 band, respectively. The or- fusion signals are expected representing ing the CRLF2 (cytokine receptor-like factor ange fluorochrome direct labeled probe normal (non-rearranged) Xp22.33 loci. 2, a.k.a. CRL2, TSLPR) gene. hybridizes proximal to the CRLF2 gene at In an interphase nucleus of a normal male The CRLF2 protein interacts with IL7R to Xp22.33 and Yp11.32, the green fluoro- cell lacking a translocation involving the form a receptor for TSLP, binding of which chrome direct labeled probe hybridizes Xp22.33 or Yp11.32 band, two orange/ activates cell signaling through JAK/STAT distal to the CRLF2 gene at Xp22.33 and green fusion signals are expected repre- pathways. Yp11.32. senting normal (non-rearranged) Xp22.33 Approximately 7% of patients with B-cell and Yp11.32 loci. precursor ALL (B-ALL) and more than CRLF2 A signal pattern consisting of one orange/ 50% of B-ALL in children with Down green fusion signal, one orange signal, syndrome harbor alterations involving the and a separate green signal indicates one CRLF2 gene. These include the translo- normal Xp22.33 or Yp11.32 locus and cations t(X;14)(p22.33;q32.3) or t(Y;14) one Xp22.33 or Yp11.32 locus affected (p11.32;q32.3) which fuse the entire by a translocation. CRLF2 gene to the immunoglobulin heavy Loss of the orange signals or orange chain enhancer region (IGH-CRLF2). signals of reduced size are the result of Another common alteration is an interstitial Ideogram of chromosome X deletions proximal to the CRLF2 break- indicating the hybridization locations. deletion involving the pseudoautosomal point region. region (PAR1) of the sex chromosomes CRLF2 upstream of CRLF2, juxtaposing the first non-coding exon of P2RY8 to the entire coding region of CRLF2 (P2RY8-CRLF2). Ideogram of chromosome Y These rearrangements, which are often indicating the hybridization locations. accompanied by JAK mutations, result in SY2032 BV209312 overexpression of CRLF2 and were shown A054YD1 DXYS138 to contribute to lymphoid transformation. 3’5’ 3’ 5’

Patients with CRLF2 rearrangements and CRLF2 P2RY8

JAK mutations have a poor event-free and ~735 kb ~165 kb overall survival. Tel Xp22.33 Cen Moreover, the detection of CRLF2 rear- SPEC CRLF2 Probe map (not to scale). rangements by FISH may help in selecting SY2032 BV209312 Bone marrow smear with translocation affecting the CRLF2 gene locus as indicated by one B-ALL patients eligible for therapy with A054YD1 DXYS138 3’5’ 3’ 5’ non-rearranged orange/green fusion signal, one inhibitors of the JAK/STAT pathway. orange signal, and one separate green signal. References CRLF2 P2RY8 Harvey RC, et al. (2010) Blood 115: 5312-21. Mullighan CG, et al. (2009) Nat Genet 41: 1243-6. ~735 kb ~165 kb Roberts KG, et al. (2014) N Engl J Med 371: 1005-15. Russell LJ, et al. (2009) Blood 114: 2688-98. Tel Yp11.32 Cen Tasian SK, et al. (2012) Blood 120: 833-42. SPEC CRLF2 Probe map (not to scale).

Prod. No. Product Label Tests* (Volume) Z-2201-50 ZytoLight SPEC CRLF2 Dual Color Break Apart Probe / 5 (50 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

168 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 168 FE102-1-20 ZytoLight ® SPEC TFE3 Dual Color Break Apart Probe

Background Probe Description Results The ZytoLight ® SPEC TFE3 Dual Color The SPEC TFE3 Dual Color Break Apart In a female interphase nucleus lacking a Break Apart Probe is designed to detect Probe is a mixture of two direct labeled translocation involving the Xp11.23 band translocations involving the chromosomal probes hybridizing to the Xp11.23 band. two orange/green fusion signals are region Xp11.23 harboring the TFE3 (tran- The orange fluorochrome direct labeled expected representing two normal (non-re- scription factor binding to IGHM enhancer probe hybridizes distal to the TFE3 gene, arranged) Xp11.23 loci. In a normal male 3, a.k.a. TFEA) gene. the green fluorochrome direct labeled interphase nucleus one orange/green Translocations involving the chromosomal probe hybridizes proximal to that gene. fusion signal is expected representing one region Xp11.2 are frequently detected normal (non-rearranged) Xp11.23 locus. in renal cell carcinomas (RCCs) which One separate green and separate orange usually affect children and adolescents. signal indicate one Xp11.23 locus affect- The Xp11.2 translocation RCCs represent TFE3 ed by a translocation. a predominant and aggressive subtype in the pediatric age group but can also occur in adults. Macroscopically, Xp11.2 translocation RCCs may mimic conventional clear cell RCCs and thus, differential diagnosis of Xp11.2 translocation RCCs is Ideogram of chromosome X indicating the hybridization locations. clinically important. Additionally, the unbalanced chromo- RH18187 RH103001 RH66279 DXS1426 somal translocation of der(17)t(X;17) 3’ 5’ (p11.23;q25) is cytogenetically characteristic for alveolar soft part sarcoma (ASPS). TFE3 ASPS is a rare high grade mesenchymal ~505 kb ~605 kb Tel Xp11.23 Cen malignancy affecting mainly adolescents. SPEC TFE3 Dual Color Break Apart Probe hybridized SPEC TFE3 Probe map (not to scale). This translocation fuses the TFE3 gene at to normal interphase cells as indicated by two orange/green fusion signals per nucleus. Xp11.23 to the ASPSCR1 (alveolar soft part sarcoma chromosome region, candidate 1, a.k.a ASPL) gene on 17q25.3. Diagnosis of ASPS is often difficult due to histologic overlap with other tumors, particularly in small biopsies. Thus, FISH analysis can improve accuracy of ASPS diagnosis. References Argani P, et al. (2001) Am J Pathol 159: 179-92. Armah HB, et al. (2009) Diagn Pathol 4: 15. Dijkhuizen T, et al. (1995) Genes Chromosomes Cancer 14: 43-50. Ladanyi M, et al. (2001) Oncogene 20: 48-57. Llamas-Velasco M, et al. (2013) Histopathology 63: 122-9. Pflueger D, et al. (2013) Neoplasia 15: 1231-40. Wiliams A, et al. (2011) Virchows Arch 458: 291-300. Wu A, et al. (2008) Histopathology 53: 533-44. Yan BC, et al. (2009) Arch Pathol Lab Med 133: 1026-32. Renal cell carcinoma section with translocation affecting the Xp11.23 locus as indicated by one non-rearranged green/orange fusion signal, one separate green signal, and one separate orange signal.

Prod. No. Product Label Tests* (Volume) Z-2109-50 ZytoLight SPEC TFE3 Dual Color Break Apart Probe / 5 (50 μl) Z-2109-200 ZytoLight SPEC TFE3 Dual Color Break Apart Probe / 20 (200 μl) Related Products Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

169 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 169 FE048-1-20 ZytoLight ® Probes for Chromosome Enumeration

Background ZytoLight ® SPEC Probe Maps The ZytoLight ® Chromosome Enumeration SHGC-141884 RH105871 D13S1574 D13S1558 Probes are designed for identification and 3’ 5’ 5’ 3’ enumeration of human chromosomes in in- WARS2 HAO2 terphase cells and as an adjunct to stand- ~285 kb ~690 kb ard karyotyping in metaphases. These Tel 1p12 Cen Cen 13q12.11 Tel probes will produce sharp, bright signals SPEC 1p12 Probe map (not to scale). SPEC 13q12 Probe map (not to scale). specific for each individual chromosome. The ZytoLight ® SPEC 1p12 Probe is The ZytoLight ® SPEC 13q12 Probe is CEN Probe Description designed to hybridize in close proximity of designed to hybridize in close proximity For most chromosomes, direct labeled centromere 1 at 1p12 harboring WARS2 of centromere 13 at 13q12.11. Since ZytoLight ® CEN ™ Probes hybridizing to and HA02. Since chromosomes 1, 5, and chromosomes 13 and 21 share the same highly repetitive human satellite DNA se- 19 share the same repetitive sequences, repetitive sequences, they cannot be differ- quences mainly located at the centromeric they cannot be differentiated by probes entiated by probes detecting centromere regions of chromosomes are applicable. detecting centromere specific repeats. specific repeats.

SPEC Probe Description D2S2776 RH121214 SHGC-154558 RH93602 As several chromosomes share the 3’ 5’ 5’ 3’ same repetitive sequences resulting in cross-hybridization signals, they AFF3 NEDD4L cannot be differentiated by centromere ~490 kb ~765 kb Cen 2q11.2 Tel Cen 18q21.31-q21.32 Tel specific probes. Instead, these chromosomes can be identified by direct labeled SPEC 2q11 Probe map (not to scale). SPEC 18q21 Probe map (not to scale). ZytoLight ® SPEC ™ Probes hybridizing in The ZytoLight ® SPEC 2q11 Probe is The SPEC 18q21 Probe, included in the close proximity to the respective satellite specific for the AFF3 (AF4/FMR2 family, ZytoLight ® SPEC 18/CEN X/Y Triple DNA sequences or to other chromosome member 3) gene region in 2q11.2. Due to Color Probe, is specific for NEDD4L specific loci. cross-hybridizations of chromosome 2 al- (NEDD4 like E3 ubiquitin protein ligase) pha satellites to other centromeric regions, gene region in 18q21.31-q21.32. Results probes specific for 2q11 are frequently In a normal interphase nucleus, two sig- used for chromosome 2 copy number nals are expected using Chromosome Enu- detection. meration Probes specific for autosomes. Using chromosome Y specific probes will RH67343 RH82989 RH103587 D21S338 result in normal male cells in one signal 5’3’ and in normal female cells in no signal. Using chromosome X specific probes will ZAR1 result in normal male cells in one signal ~435 kb ~410 kb Tel 4p11 Cen Cen 21q22.13-q22.2 Tel and in normal female cells in two signals per nucleus. Other signal patterns indicate SPEC 4p11 Probe map (not to scale). SPEC 21q22 Probe map (not to scale). numerical aberrations of the respective The ZytoLight ® SPEC 4p11 Probe is The ZytoLight ® SPEC 21q22 Probe hybrid- chromosome. designed to hybridize in close proximity of izes to the so-called Down Syndrome Criti- centromere 4 at 4p11 harboring the ZAR1 cal Region on 21q22.13-q22.2 commonly (zygote arrest 1) gene. For an unambig- duplicated in cases with partial trisomy uous enumeration of chromosome 4 the 21. Since chromosomes 13 and 21 share SPEC 4p11 is found to be more suitable. the same repetitive sequences, they cannot be differentiated by probes detecting centromere specific repeats.

170 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 170 FE001-1-20 Prod. No. Product Alpha/Class. Sat. Chr. Band Label Tests* (Volume) Z-2101-200 ZytoLight SPEC 1p12 Probe - 1p12 20 (200 μl) Z-2049-200 ZytoLight SPEC 2q11 Probe - 2q11.2 20 (200 μl) Z-2001-200 ZytoLight CEN 3 Probe D3Z1 3p11.1-q11.1 20 (200 μl) Z-2083-200 ZytoLight SPEC 4p11 Probe - 4p11 20 (200 μl) Z-2002-200 ZytoLight CEN 6 Probe D6Z1 6p11.1-q11 20 (200 μl) Z-2003-200 ZytoLight CEN 7 Probe D7Z1 7p11.1-q11.1 20 (200 μl) Z-2004-50/-200 ZytoLight CEN 8 Probe D8Z2 8p11.1-q11.1 5/20 (50/200 μl) Z-2067-200 ZytoLight CEN 9 Probe III D9Z3 9q12 20 (200 μl) Z-2079-200 ZytoLight CEN 10 Probe D10Z1 10p11.1-q11.1 20 (200 μl) Z-2005-200 ZytoLight CEN 11 Probe D11Z1 11p11.11-q11 20 (200 μl) Z-2050-200 ZytoLight CEN 12 Probe D12Z3 12p11.1-q11 20 (200 μl) Z-2085-200 ZytoLight SPEC 13q12 Probe - 13q12.11 20 (200 μl) Z-2095-50/-200 ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe D18Z1 13q12.11/18p11.1/21q22.13-q22.2 •/•/ 5/20 (50/200 μl) Z-2164-200 ZytoLight SPEC 13/21 Dual Color Probe - 13q12.11/21q22.13-q22.2 / 20 (200 μl) Z-2006-200 ZytoLight CEN 17 Probe D17Z1 17p11.1-q11.1 20 (200 μl) Z-2007-200 ZytoLight CEN 18 Probe D18Z1 18p11.1-q11.1 20 (200 μl) Z-2163-200 ZytoLight SPEC 18/CEN X/Y Triple Color Probe DXZ1/DYZ3 18q21.31-q21.32/Xp11.1-q11.1/Yp11.1-q11.1 •/•/ 20 (200 μl) Z-2086-200 ZytoLight SPEC 21q22 Probe - 21q22.13-q22.2 20 (200 μl) Z-2180-200 ZytoLight SPEC 21/CEN X/Yq12 Triple Color Probe DXZ1/ III DYZ1 21q22.13-q22.2/Xp11.1-q11.1/Yq12 •/ /• 20 (200 μl) Z-2008-200 ZytoLight CEN X Probe DXZ1 Xp11.1-q11.1 20 (200 μl) Z-2010-200 ZytoLight CEN Yq12 Probe III DYZ1 Yq12 20 (200 μl) Z-2123-200 ZytoLight CEN Y (DYZ3) Probe DYZ3 Yp11.1-q11.1 20 (200 μl) Z-2016-50/-200 ZytoLight CEN X/Yq12 Dual Color Probe DXZ1/III DYZ1 Xp11.1-q11.1/Yq12 /• 5/20 (50/200 μl) Z-2120-200 ZytoLight CEN X/Y Dual Color Probe DXZ1/ DYZ3 Xp11.1-q11.1/Yp11.1-q11.1 / 20 (200 μl) Related Products Z-2279-20 ZytoLight Aneuploidy Panel 18/X/Y and 13/21 20 Incl. ZytoLight SPEC 18/CEN X/Y Triple Color Probe, 0.2 ml (Z-2163-200); ZytoLight SPEC 13/21 Dual Color Probe, 0.2 ml (Z-2164-200) Z-2104-5 ZytoLight Aneuploidy Panel X/Y and 13/18/21 5 Incl. ZytoLight CEN X/Yq12 Dual Color Probe, 0.05 ml (Z-2016-50); ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe, 0.05 ml (Z-2095-50) Z-2104-20 ZytoLight Aneuploidy Panel X/Y and 13/18/21 20 Incl. ZytoLight CEN X/Yq12 Dual Color Probe, 0.2 ml (Z-2016-200); ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe, 0.2 ml (Z-2095-200) Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

171 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 171 FE001-1-20 ZytoLight ® Aneuploidy Panel 18/X/Y and 13/21

Background Probe Description Results The ZytoLight ® Aneuploidy Panel 18/X/Y The ZytoLight ® Aneuploidy Panel 18/X/Y In an interphase nucleus of a normal cell and 13/21 is designed for chromosome and 13/21 is comprised of the ZytoLight ® using the ZytoLight ® SPEC 13/21 Dual enumeration of the chromosomes 13, 18, SPEC 18/CEN X/Y Triple Color Probe Color Probe, two green and two orange 21, X, and Y. hybridizing to chromosome 18 specific signals are expected. Trisomies of the autosomes 13, 18, or 21 sequences at 18q21.31-q21.32 and to In an interphase nucleus of a normal cell, (Down Syndrome) are common genomic the alpha satellites of the chromosomes X using the ZytoLight ® SPEC 18/CEN X/Y aberrations. Aberrant numbers of the gon- (DYZ1) and Y (DYZ3), and of the Triple Color Probe, two blue signals are osomes X and Y are resulting in disorders ZytoLight ® SPEC 13/21 Dual Color expected. Two green signals are expect- of sex development (DSD). Diseases such Probe hybridizing to chromosome 13 ed in a normal female cell, or one single as Ulrich-Turner-Syndrome (45, X) or Triple and 21 specific sequences at 13q12.11 green and one single orange signal is X Syndrome (47, XXX) may cause severe and 21q22.13-q22.2, respectively. Both expected in a normal male cell. developmental and metabolic disorders. probes are approved to be used with a Other signal patterns indicate numerical The prevalence of chromosomal abnormal- hybridization time of 2 hours on cytologi- aberration of the respective chromosomes. ities detectable in the newborn, including cal specimens. chromosome 13, 18, 21, X, and Y, is D13S1574 D13S1558 about 0.92%.

References Gillenberg C, (1998) J Autism Dev Disord 28: 415-25. Jacobs PA, et al. (1992) J Med Genet 29: 103-8. ~690 kb Cen 13q12.11 Tel

SPEC 13q12 Probe map (not to scale).

SHGC-154558 RH93602 5’ 3’

NEDD4L SPEC 18/CEN X/Y Triple Color Probe hybridized to interphase nuclei of normal male cells and to ~765 kb chromosomes of a metaphase spread. Cen 18q21.31-q21.32 Tel

SPEC 18q21 Probe map (not to scale).

RH103587 D21S338

~410 kb Cen 21q22.13-q22.2 Tel

SPEC 21q22 Probe map (not to scale).

SPEC 13/21 Dual Color Probe hybridized to interphase cells with trisomy of chromosome 21.

Prod. No. Product Label Tests* (Volume) Z-2279-20 ZytoLight Aneuploidy Panel 18/X/Y and 13/21 20 (200 μl) Related Products Z-2164-200 ZytoLight SPEC 13/21 Dual Color Probe •/• 20 (200 μl) Z-2163-200 ZytoLight SPEC 18/CEN X/Y Triple Color Probe •/•/• 20 (200 μl) Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

172 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 172 FE145-1-20 ZytoLight ® Aneuploidy Panel X/Y and 13/18/21

Background Probe Description Results The ZytoLight ® Aneuploidy Panel X/Y and The ZytoLight ® Aneuploidy Panel X/Y and In an interphase nucleus, using the 13/18/21 is designed for enumeration of 13/18/21 is comprised of the ZytoLight ® ZytoLight ® CEN X/Yq12 Dual Color the chromosomes 13, 18, 21, X, and Y. CEN X/Yq12 Dual Color Probe hybridiz- Probe, two orange signals are expected Trisomies of the autosomes 13, 18, or 21 ing to the alpha satellites of chromosome in a normal female cell whereas one single (Down Syndrome) are common genomic X (DYZ1) and to the classical satellite orange and one single green signal is aberrations. Aberrant numbers of the gon- III of chromosome Y (DYZ1), and of the expected in a normal male cell. osomes X and Y are resulting in disorders ZytoLight ® SPEC 13/CEN 18/SPEC In an interphase nucleus of a normal cell, of sex development (DSD). Diseases such 21 Triple Color Probe hybridizing to the using the ZytoLight ® SPEC 13/CEN 18/ as Ulrich-Turner-Syndrome (45, X) or Triple chromosome 13 and 21 specific sequenc- SPEC 21 Triple Color Probe, two green, X Syndrome (47, XXX) may cause severe es at 13q12.11 and 21q22.13-q22.2, two blue, and two orange signals are developmental and metabolic disorders. respectively, and to the alpha satellites expected. The prevalence of chromosomal abnormal- of chromosome 18. Both probes are Other signal patterns indicate numerical ities detectable in the newborn including approved to be used with a hybridization aberrations of the respective chromosomes. chromosomes 13, 18, 21, X, and Y, is time of 2 hours on cytological specimens. about 0.92%. D13S1574 D13S1558 References Gillenberg C, (1998) J Autism Dev Disord 28: 415-25. Jacobs PA, et al. (1992) J Med Genet 29: 103-8.

~690 kb Cen 13q12.11 Tel

SPEC 13q12 Probe map (not to scale).

RH103587 D21S338

SPEC 13/CEN 18/ SPEC 21 Triple Color Probe hybridized to normal interphase cells. ~410 kb Cen 21q22.13-q22.2 Tel

SPEC 21q22 Probe map (not to scale).

CEN X/Yq12 Dual Color Probe hybridized to metaphase chromosomes of a normal male cell.

Prod. No. Product Label Tests* (Volume) Z-2104-5/20 ZytoLight Aneuploidy Panel X/Y and 13/18/21 5/20 (50/200 μl) Related Products Z-2095-50/-200 ZytoLight SPEC 13/CEN 18/ SPEC 21 Triple Color Probe •/•/• 5/20 (50/200 μl) Z-2016-50/-200 ZytoLight CEN X/Yq12 Dual Color Probe •/• 5/20 (50/200 μl) Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

173 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 173 FE144-1-20 Page

Method Introduction - FlexISH ® 175

Probes, sorted by Chromosomes 176 f. sorted by Gene Names 178 sorted by Indication 178

Product Data Sheets 179 ff.

Accessories 184 f.

FISH Reagents, Fluorochromes and Filter Recommendations 186 ff.

174 174 Simply Adapt the Hybridization Time to your Needs!

F ISH® Bringing to Your FISH

Introduction Advantages of FlexISH ® FlexISH ® Kit - Convenient Solution FlexISH ® products are designed for • Hybridization time can be varied All FlexISH ® probes can be combined identification of chromosomal aberrations between 2 hours and overnight. with the FlexISH ®-Tissue Implementation on various specimens by FISH. Using the • With a hybridization temperature of Kit to obtain reliable results already FlexISH ® products gives you the flexibility 37°C the FlexISH ® protocol is fully within 4.5 hours. to choose between a 1-day (2 h hybridiza- compatible with routine workflows in The FlexISH ® protocol can also be tion) or a 2-day (overnight hybridization) pathology laboratories. incorporated into the routine workflow protocol by adapting the hybridization • Short hybridization time does not with overnight hybridization providing time just according to your individual negatively affect the performance, the highest flexibility. needs! specimen quality or diagnostic result1.

High-Quality FISH Results with flexible Hybridization Time

There is an excellent correlation between the FISH results obtained after overnight and short hybridization periods with regard to signal brightness, signal-to-noise ratio, and the diagnostic result1.

Standard FISH workflow

FlexISH ® FISH workflow

from 2 hours to overnight hybridization

0 h 2 h 4 h 6 h 8 h 10 h 12 h 14 h 16 h 18 h 20 h

Dewaxing Heat Pretreatment Proteolysis Denaturation Hybridization Stringency Wash Mounting

References 1 Brockhoff G, et al. (2016) Histopathology 69: 635-46.

175 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 175 MFLE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

1 no probes available yet

2 2p23 FlexISH ALK/ROS1 DistinguISH™ Probe Z-2203-50/-200 50/200 µl 179

3 3q27 FlexISH BCL2/BCL6 DistinguISH™ Probe Z-2283-50/-200 50/200 µl 180

4-5 no probes available yet

6 6q22.1 FlexISH ALK/ROS1 DistinguISH™ Probe Z-2203-50/-200 50/200 µl 179

7 no probes available yet

8 8q24.21 FlexISH MYC/IGH TriCheck™ Probe Z-2293-50 50 µl 181

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

176 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 176 IFLE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

9 no probes available yet

10 10p11.2 FlexISH RET/KIF5B TriCheck™ Probe Z-2269-50/-200 50/200 µl 182 10q11.2 FlexISH RET/KIF5B TriCheck™ Probe Z-2269-50/-200 50/200 µl 182

11-13 no probes available yet

14 14q32.3 FlexISH MYC/IGH TriCheck™ Probe Z-2293-50 50 µl 181

15-16 no probes available yet

17 17q12 FlexISH ERBB2/CEN 17 Dual Color Probe Z-2166-50/-200 50/200 µl 183

18 18q21.3 FlexISH BCL2/BCL6 DistinguISH™ Probe Z-2283-50/-200 50/200 µl 180

19-22 no probes available yet

X, Y no probes available yet

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

177 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 177 IFLE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

ALK CD246 FlexISH ALK/ROS1 DistinguISH™ Probe Z-2203-50/-200 50/200 µl 179

BCL2 Bcl-2, PPP1R50 FlexISH BCL2/BCL6 DistinguISH™ Probe Z-2283-50/-200 50/200 µl 180

BCL6 ZNF51, LAZ3 FlexISH BCL2/BCL6 DistinguISH™ Probe Z-2283-50/-200 50/200 µl 180

ERBB2 HER2, HER-2, NEU FlexISH ERBB2/CEN 17 Dual Color Probe Z-2166-50/-200 50/200 µl 183

IGH IGH@ FlexISH MYC/IGH TriCheck™ Probe Z-2293-50 50 µl 181

KIF5B KNS1 FlexISH RET/KIF5B TriCheck™ Probe Z-2269-50/-200 50/200 µl 182

MYC CMYC, bHLHe39, c-Myc FlexISH MYC/IGH TriCheck™ Probe Z-2293-50 50 µl 181

RET HSCR1, CDHF12 FlexISH RET/KIF5B TriCheck™ Probe Z-2269-50/-200 50/200 µl 182

ROS1 MCF3, ROS FlexISH ALK/ROS1 DistinguISH™ Probe Z-2203-50/-200 50/200 µl 179

Indication Index Indication Product Name Product No. Quantity Page

Solid Tumors Breast Cancer FlexISH ERBB2/CEN 17 Dual Color Probe Z-2166-50/-200 50/200 µl 183

Gastrointestinal Cancer FlexISH ERBB2/CEN 17 Dual Color Probe Z-2166-50/-200 50/200 µl 183

Lung Cancer FlexISH ALK/ROS1 DistinguISH™ Probe Z-2203-50/-200 50/200 µl 179 FlexISH RET/KIF5B TriCheck™ Probe Z-2269-50/-200 50/200 µl 182 FlexISH ERBB2/CEN 17 Dual Color Probe Z-2166-50/-200 50/200 µl 183

Hematology Specific Probes Non-Hodgkin Lymphoma, other FlexISH BCL2/BCL6 DistinguISH™ Probe Z-2283-50/-200 50/200 µl 180 FlexISH MYC/IGH TriCheck™ Probe Z-2293-50 50 µl 181

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

178 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 178 IFLE001-1-20 FlexISH ® ALK/ROS1 DistinguISH ™ Probe

Background Probe Description Results The FlexISH ® ALK/ROS1 DistinguISH™ The FlexISH ® ALK/ROS1 DistinguISH™ In an interphase nucleus without ALK or Probe is designed to detect rearrange- Probe is a mixture of five direct labeled ROS1 rearrangements, two ALK specific ments involving the chromosomal region probes hybridizing to the 2p23.1-p23.2 green/orange fusion signals and two 2p23.1-p23.2 and 6q22.1 harboring the and 6q22.1-q22.2 bands. The orange ROS1 specific green/orange/blue fusion ALK (ALK receptor tyrosine kinase, a.k.a. fluorochrome direct labeled probe frac- signals are expected. An ALK rearrange- CD246) and ROS1 (ROS proto-oncogene tions hybridize distal to the ALK and ROS1 ment is indicated by one separate orange 1, receptor tyrosine kinase) gene, re- breakpoint regions, the green direct signal and/or one separate green signal, spectively. Using this probe, it is possible labeled probe fractions hybridize proximal both not co-localizing with blue signals. to simultaneously detect ALK and ROS1 to the ALK and ROS1 breakpoint regions. A ROS1 rearrangement is indicated by rearrangements and, additionally, to The blue fluorochrome direct labeled one separate green signal, and/or one discriminate between possible aberrations probe hybridizes distal and proximal to separate orange signal both co-localizing affecting these chromosomal regions. the ROS1 breakpoint region. with blue signals. Both, the ALK as well as the ROS1 gene, encode for transmembrane receptor ALK tyrosine kinases. Rearrangements affecting the ALK or the ROS1 gene locus are frequently found in non-small cell lung cancer (NSCLC). The most frequent ALK rearrangement in NSCLC is the inversion [inv(2)(p21p23)] affecting the genes ALK and EML4, both located on chromosome 2. ROS1 The ROS1 gene is evolutionary closely related to the ALK family which forms part H3122 cell line which shows two green/orange/blue of the scientific basis of using inhibitors of fusion signals and one orange/green fusion signal. ALK as inhibitors of ROS1. ALK and ROS1 An ALK rearrangement is indicated by one separate orange and one separate green signal, positive NSCLC patients benefit from a Ideograms of chromosomes 2 (left) and 6 (right) indicating the hybridization locations. both not co-localizing with blue signals. tyrosine kinase targeted therapy, like, e.g., crizotinib. References Birchmaier C, et al. (1987) Proc Natl Acad Sci U S A 84: 9270-4. Bos M, et al. (2013) Lung Cancer 81: 142-3. Sasaki T, et al. (2010) Eur J Cancer 46: 1773-80. Shaw AT, et al. (2014) N Engl J Med 371: 1963-71.

RH27490 D6S2158 SHGC-130752 SHGC-142586 D2S2934 D2S405 RH69070 RH104060 RH12489 SHGC-142979 D6S1391E RH78348 3’ 5’ 3’ 5’ 3’ 5’

ALK ROS1 GOPC ~715 kb ~215 kb ~210 kb ~635 kb ~590 kb ~680 kb Tel 2p23.1-p23.2 Cen Cen 6q22.1-q22.2 Tel HCC78 cell line which shows two green/orange fusion signals and one green/orange/blue fusion ALK Probe map (not to scale). ROS1 Probe map (not to scale). signal. ROS1 rearrangement is indicated by one separate orange and one separate green signal, both co-localizing with blue signals.

Prod. No. Product Label Tests* (Volume) Z-2203-50 FlexISH ALK/ROS1 DistinguISH Probe / / 5 (50 μl) Z-2203-200 FlexISH ALK/ROS1 DistinguISH Probe / / 20 (200 μl) Related Products Z-2182-5 FlexISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; 5x FlexISH Wash Buffer, 150 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2182-20 FlexISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; 5x FlexISH Wash Buffer, 500 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

179 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 179 FLE001-1-20 FlexISH ® BCL2/BCL6 DistinguISH™ Probe

Background Probe Description Results The FlexISH ® BCL2/BCL6 DistinguISH™ The FlexISH ® BCL2/BCL6 DistinguISH™ In an interphase nucleus without BCL2 or Probe is designed to detect rearrange- Probe is a mixture of five direct labeled BCL6 rearrangements, two BCL2 specific ments involving the chromosomal regions probes hybridizing to the 18q21.33-q22.1 green/orange fusion signals and two 18q21.33 and 3q27.3 harboring the BCL2 and 3q27.3-q28 bands. The green fluo- BCL6 specific green/orange/blue fusion (BCL2 apoptosis regulator, a.k.a. PPP1R50) rochrome direct labeled probes hybridize signals are expected. A BCL2 rearrange- gene and the BCL6 (BCL6 transcription proximal to the BCL2 and BCL6 break- ment is indicated by one separate green repressor, a.k.a. ZNF51, LAZ3) gene, point regions, and the orange fluoro- and one separate orange signal, both not respectively. Using this probe, it is possible chrome direct labeled probes hybridize co-localizing with blue signals. A BCL6 to simultaneously detect BCL2 and BCL6 re- distal to the BCL2 and BCL6 breakpoint rearrangement is indicated by one separate arrangements and, additionally, to discrim- regions. The blue fluorochrome direct labe- green and one separate orange signal, inate between possible aberrations affecting led probe hybridizes distal and proximal both co-localizing with blue signals. these chromosomal regions, individually. to the BCL6 breakpoint region. BCL2 encodes for a mitochondrial membrane protein that regulates apoptosis and is expressed in B-cells. BCL6 encodes for a protein that acts as a transcriptional repressor involved in the regulation of lymphoid development and function. BCL2 and BCL6 rearrangements are frequently found in various Non-Hodgkin lymphomas. Additionally, BCL2 Lymphoma tissue which shows two green/orange/blue BCL2 and BCL6 rearrangements are known fusion signals and one green/orange fusion signal. BCL2 rearrangement is indicated by one separate to be concurrent with MYC rearrangements. green and one separate orange signal, both not co- BCL6 localizing with blue signals. MYC rearrangements with either BCL2 or Specimen kindly provided by Dr. Rontogianni, Athens, Greece. Ideograms of chromosomes 3 (left) and BCL6 co-aberration are so-called double-hit 18 (right) indicating the hybridization locations. B-cell lymphomas (DHL) known to be highly RH26523 RH123158 aggressive with poor prognosis. Rarely, D3S4060 RH29459 triple-hit B-cell lymphomas (THL) showing 3’ 5’

simultaneous rearrangements of MYC, BCL2, BCL6 and BCL6 occur. According to the ~825 kb ~665 kb revised 4th edition of the WHO classification Cen 3q27.3-q28 Tel

of tumors of haematopoietic and lymphoid BCL6 Probe map (not to scale). tissues (2017) DHL and THL are classified DLBCL tissue which shows one green/orange/blue as high-grade B-cell lymphoma with MYC D18S91 RH44852 fusion signal and one green/orange fusion signal. SHGC-147772 REN12166 BCL6 rearrangement is indicated by one 3’ 5’ and BCL2 and/or BCL6 rearrangements. separate green and one separate orange signal, Hence, detection of BCL2 and/or BCL6 both colocalizing with blue signals. Additionally, BCL2 one separate orange and one separate green rearrangements using Fluorescence in situ ~735 kb ~665 kb signal indicate a further BCL2 positivity, confirming a BCL2/BCL6 co-rearrangement. Hybridization (FISH) may be of diagnostic Cen 18q21.33-q22.1 Tel References and prognostic relevance. Swerdlow SH, et al. (ed.) (2017) WHO Classification of Tumours of BCL2 Probe map (not to scale). Haematopoietic and Lymphoid Tissues (Revised 4th Edition).

Prod. No. Product Label Tests* (Volume) Z-2283-50 FlexISH BCL2/BCL6 DistinguISH Probe / / 5 (50 μl) Z-2283-200 FlexISH BCL2/BCL6 DistinguISH Probe / / 20 (200 μl) Related Products Z-2182-5 FlexISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; 5x FlexISH Wash Buffer, 150 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2182-20 FlexISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; 5x FlexISH Wash Buffer, 500 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

180 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 180 FLE003-1-20 FlexISH ® MYC/IGH TriCheck™ Probe

Background Probe Description Results The FlexISH ® MYC/IGH TriCheck™ Probe is The FlexISH ® MYC/IGH TriCheck™ In an interphase nucleus without designed to detect the translocation t(8;14) Probe is a mixture of three direct labeled rearrangements of the MYC/IGH loci, two (q24.21;q32.3) affecting the MYC gene in probes hybridizing to the 8q24.21 green/orange fusion signals and two blue the chromosomal region 8q24.21 and the and 14q32.33 bands. The orange signals are expected. IGH locus in 14q32.33. Moreover, using fluorochrome direct labeled probe A MYC-IGH fusion is indicated by one this probe it is possible to discriminate hybridizes proximal to the MYC gene separate green signal and one separate between MYC-IGH translocations and MYC region, and the green fluorochrome direct orange signal, both co-localizing with translocations involving fusion partners labeled probe hybridizes distal to the blue signals. A MYC translocation other than IGH. MYC gene region. The blue fluorochrome without involvement of IGH is indicated The MYC proto-oncogene (a.k.a. CMYC) direct labeled probe spans the known by separated orange and green signals encodes a transcription factor essential for breakpoints of IGH. without co-localization of the separated cell growth and proliferation and is broadly signals with blue signals. implicated in tumorigenesis. Translocations involving the MYC gene are considered to be cytogenetic hallmarks for Burkitt lymphoma (BL) but are also found in other types of lymphomas. The most frequent translocation involving the MYC gene region t(8;14) (q24.21;q32.3) can be found in approx. 80% of the BL cases and juxtaposes the MYC gene next to IGH (immunoglobulin heavy locus). Further translocations affecting the MYC gene are t(8;22) (q24.21;q11.2) and t(2;8)(p11.2;q24.21), Non-Hodgkin lymphoma tissue section with t(8;14) as indicated by one separate green both of which involve one of the two and one separate orange signal, and Ideograms of chromosomes 8 (above) and one additional blue signal. immunoglobulin light chain loci. All three 14 (below) indicating the hybridization locations. translocations bring the MYC gene under the control of a regulatory element from SHGC-130556 D8S1701 D8S1090 RH75025 one of the immunoglobulin loci resulting in 5’3’ constitutive overexpression of MYC. MYC Large B-cell lymphoma patients with MYC- ~475 kb ~560 kb IG have shorter overall survival compared Cen 8q24.21 Tel

with both MYC translocation with non-IG MYC Probe map (not to scale). translocation partner gene as well as absence of MYC translocation. Thus, the BV448316 RH119884 3’ 5’ 3’ 5’ detection of MYC translocation partner by FISH may prove a valuable diagnostic and JAG2 IGH prognostic tool. ~1.5 Mb Cen 14q32.33 Tel Non-Hodgkin lymphoma tissue section with translo- References cation of the MYC gene without IGH involvement as May P, et al. (2010) Cancer Genet Cytogenet 198: 71-5. IGH Probe map (not to scale). Pedersen MØ, et al. (2014) Eur J Haematol 92: 42-8. indicated by one separate green and one separate Perkins AS & Friedberg JW (2008) Hematology Am Soc Hematol Educ Program: 341-8. orange signal, without an additional blue signal. Veronese ML, et al. (1995) Blood 85: 2132-8.

Prod. No. Product Label Tests* (Volume) Z-2293-50 FlexISH MYC/IGH TriCheck Probe / / 5 (50 μl) Related Products Z-2182-5 FlexISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; 5x FlexISH Wash Buffer, 150 ml; DAPI/DuraTect-Solution, 0.2 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

181 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 181 FLE005-1-20 FlexISH ® RET/KIF5B TriCheck™ Probe

Background Probe Description Results The FlexISH ® RET/KIF5B TriCheck™ Probe is The FlexISH ® RET/KIF5B TriCheck™ In an interphase nucleus without designed to detect inversions involving the Probe is a mixture of three direct labeled rearrangements of the KIF5B/RET locus, two chromosomal region 10q11.21 harboring probes hybridizing to the 10q11.21 and green/orange fusion signals and two blue the rearranged during transfection (RET) 10p11.22 bands. The green fluorochrome signals are expected. gene and the chromosomal region direct labeled probe hybridizes distal A KIF5B-RET inversion is indicated by one 10p11.22 harboring the kinase family to the RET gene region, and the orange separate green signal, one separate orange member 5B (KIF5B) gene. Moreover, using fluorochrome direct labeled probe signal, and an additional blue signal. this probe it is possible to discriminate hybridizes proximal to the RET gene A RET translocation is indicated by between KIF5B-RET inversions and RET region. The blue fluorochrome direct separated orange and green signals translocations involving fusion partners other labeled probe spans the KIF5B gene without an additional blue signal. than KIF5B (e.g., BCR, FGFR1OP, and PTC). region. KIF5B-RET inversion with deletion of the RET rearrangements, including inversions 5’-RET sequences is indicated by loss of and translocations, are found in non-small one orange signal and co-localization of cell lung cancer (NSCLC) with an incidence KIF5B the isolated green signal with a blue signal. RET of 1-2%. The pericentric inversion of chromosome 10 [inv(10)(p11.2q11.2)] leads to a fusion transcript of the KIF5B gene and the RET proto-oncogene and, thus, forms a chimeric protein. The resulting Ideogram of chromosome 10 homo-dimerization of the coiled-coil indicating the hybridization locations. domains of KIF5B causes an aberrant D10S2320 RH69007 activation of the receptor tyrosine kinase D10S141 RH101258 (RTK) of RET, a mechanism known from 5’ 3’ FlexISH RET/KIF5B TriCheck™ Probe on KIF5B-ALK fusion which is also found RET normal interphase cells with non-rearranged in non-small cell lung adenocarcinoma RET loci (two green/orange fusion signals), and ~170 kb ~485 kb non-rearranged KIF5B loci (two blue signals). (LADC). LADC patients with KIF5B-RET Cen 10q11.21 Tel

fusions are commonly tested negative for RET Probe map (not to scale). LADC common driver mutations in the EGFR, KRAS, and ALK genes. RH92798 RH2783 3’ 5’ Since in vitro studies have shown that NSCLC patients presenting a KIF5B-RET KIF5B fusion are less sensitive to vandetanib ~1.5 Mb Tel 10p11.22 Cen treatment compared to patients with KIF5B independent RET-fusions, FISH analysis can KIF5B Probe map (not to scale). sustain the treatment decision. NSCLC tissue section with a KIF5B-RET inversion as indicated by one green, one separated orange, and References an additional blue signal. Gautschi O, et al. (2013) J Thorac Oncol 8: e43-4. Ju YS, et al. (2012) Genome Res 22: 436-45. Specimen kindly provided by Dr. Schildhaus, Essen, Germany. Kohno T, et al. (2012) Nat Med 18: 375-7. Tsuta K, et al. (2014) Br H Cancer 110: 1571-8. Yoh K, et al. (2017) Lancet Respir Med 5: 42-50.

Prod. No. Product Label Tests* (Volume) Z-2269-50 FlexISH RET/KIF5B TriCheck Probe / / 5 (50 μl) Z-2269-200 FlexISH RET/KIF5B TriCheck Probe / / 20 (200 μl) Related Products Z-2182-5 FlexISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; 5x FlexISH Wash Buffer, 150 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2182-20 FlexISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; 5x FlexISH Wash Buffer, 500 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

182 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 182 FLE004-1-20 FlexISH ® ERBB2/CEN 17 Dual Color Probe

Background Probe Description Results The FlexISH ® ERBB2/CEN 17 Dual Color The ERBB2/CEN 17 Dual Color Probe is In a normal interphase nucleus, two green Probe is designed for the detection of a mixture of a green fluorochrome direct and two orange signals are expected. In a ERBB2 gene amplification frequently ob- labeled ERBB2 probe specific for the chro- cell with amplification of the ERBB2 gene served in solid malignant neoplasms, e.g., mosomal region 17q12-q21.1 harboring locus, multiple copies of the green signal breast cancer samples. the ERBB2 gene and an orange fluoro- or green signal clusters will be observed. The ERBB2 gene (a.k.a. HER2 and NEU) is chrome direct labeled CEN 17 probe located in the chromosomal region 17q12 specific for the alpha satellite centromeric and encodes a 185-190 kDa transmem- region of chromosome 17 (D17Z1). brane glycoprotein, p185, acting as a cellular growth factor receptor. The p185 protein belongs to the EGFR (epidermal CEN 17 (D17Z1) ERBB2 growth factor receptor) subgroup of the RTK (receptor tyrosine kinase) superfamily also including EGFR (ERBB1), ERBB3 Ideogram of chromosome 17 (HER3), and ERBB4 (HER4). indicating the hybridization locations. Amplification of the proto-oncogene ERBB2, observed in approximately 20% of all HUMUT6435 D17S1179 5’ 3’ breast cancer samples, has been correlated with a poor prognosis of the disease. ERBB2 FlexISH ERBB2/CEN 17 Dual Color Probe hybridized for 2 hours on an endometrial carcinoma Similar results have been obtained for a ~610 kb tissue section with ERBB2 (green) amplification. Cen 17q12-q21.1 Tel variety of other malignant neoplasms, e.g., ovarian cancer, stomach cancer, and carci- ERBB2 Probe map (not to scale). nomas of the salivary gland.

FlexISH ERBB2/CEN 17 Dual Color Probe hybridized overnight on an endometrial carcinoma tissue section with ERBB2 (green) amplification.

Prod. No. Product Label Tests* (Volume) Z-2166-50 FlexISH ERBB2/CEN 17 Dual Color Probe / 5 (50 μl) Z-2166-200 FlexISH ERBB2/CEN 17 Dual Color Probe / 20 (200 μl) Related Products Z-2182-5 FlexISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; 5x FlexISH Wash Buffer, 150 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2182-20 FlexISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; 5x FlexISH Wash Buffer, 500 ml; DAPI/DuraTect-Solution, 0.8 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

183 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 183 FLE002-1-20 FISH Accessories

ZytoLight ® Implementation Kits For the detection of ZytoLight ® Probes Prod. No. Product Tests Z-2028-5 ZytoLight FISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 25x Wash Buffer A, 50 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2028-20 ZytoLight FISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 25x Wash Buffer A, 100 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

ZytoLight ® Wash Buffers Prod. No. Product

WB-0001-560 Wash Buffer SSC, 560 ml

WB-0002-50 25x Wash Buffer A, 50 ml

WB-0003-50 20x SSC Solution, 50 ml

WB-0005-50 20x Wash Buffer TBS, 50 ml

WB-0007-500 Cytology Stringency Wash Buffer SSC, 500 ml

WB-0008-500 Cytology Wash Buffer SSC, 500 ml

FlexISH ® Implementation Kits For the detection of FlexISH ® Probes Prod. No. Product Tests Z-2182-5 FlexISH-Tissue Implementation Kit 5 Incl. Heat Pretreatment Solution Citric, 150 ml; Pepsin Solution, 1 ml; 5x FlexISH Wash Buffer, 150 ml; DAPI/DuraTect-Solution, 0.2 ml Z-2182-20 FlexISH-Tissue Implementation Kit 20 Incl. Heat Pretreatment Solution Citric, 500 ml; Pepsin Solution, 4 ml; 5x FlexISH Wash Buffer, 500 ml; DAPI/DuraTect-Solution, 0.8 ml Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 20

The FlexISH ®-Tissue Implementation Kit can be used for FFPE samples and the ZytoLight ® FISH-Cytology Implementation Kit for cytology specimens in combination with any FlexISH ® FISH probe.

FlexISH ® Wash Buffers Prod. No. Product

WB-0007-500 Cytology Stringency Wash Buffer SSC, 500 ml

WB-0008-500 Cytology Wash Buffer SSC, 500 ml

WB-0010-500 5x FlexISH Wash Buffer, 500 ml

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

184 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 184 AFI004-1-20 FISH Accessories

FISH Pretreatment Reagents Prod. No. Product

ES-0001-4 Pepsin Solution, 4 ml

ES-0001-8 Pepsin Solution Set, 2x 4 ml

ES-0001-50 Pepsin Solution, 50 ml

ES-0001-1000 Pepsin Solution, 1000 ml

ES-0002-4 Cytology Pepsin Solution, 4 ml

ES-0002-50 Cytology Pepsin Solution, 50 ml

PT-0001-1000 Heat Pretreatment Solution Citric, 1000 ml PT-0006-100 Formaldehyde Dilution Buffer Set

Incl. 10x MgCl2, 50 ml; 10x PBS, 50 ml

Ancillary Reagents Prod. No. Product

E-4005-50 Fixogum Rubber Cement, 50 g

E-4005-125 Fixogum Rubber Cement, 125 g

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

185 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 185 AFI004-1-20 DAPI/DuraTect™ Solutions

Product Description ZytoVision‘s DAPI/Antifade Mounting ZytoVision‘s DAPI/Antifade Mounting ZytoVision‘s DAPI/Antifade Mounting Solutions are ready-to-use mounting media Solutions are optimized to be used on Solutions prevent permanent loss of that are applied directly to fluorescently tissue or cell specimens that have been fluorescence and protect fluorescent dyes labeled tissue or cell specimens on mi- hybridized with any available ZytoLight ®, from photobleaching during fluorescence croscope slides. They contain the nuclear FlexISH ®, or ZytoMation ® FISH Probe. microscopy. counterstain DAPI (4‘, 6-diamidino-2-phe- They are all particularly compatible with nylindole) which produces a blue fluores- the ZytoVision fluorochromes ZyGreen™, cence when bound to DNA. ZyOrange™, ZyBlue™, ZyGold™ and ZyRed™.

Prod. No. Product Concentration Storage Temperature Description

MT-0007-0.8 DAPI/DuraTect-Solution, 0.8 ml 150 ng DAPI/ml 2...8ºC · Best overall signal protection · Superior signal stability of mounted tissue sections (≤3 months at 2...21ºC)

MT-0008-0.8 DAPI/DuraTect-Solution (ultra), 0.8 ml 1360 ng DAPI/ml 2...8ºC · Best overall signal protection · Superior signal stability of mounted tissue sections (≤3 months at 2...21ºC) · Recommended when a more intense DAPI stain is desired

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

186 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 186 AFI001-1-20 ZyBlack™ Quenching Solution

Without ZyBlack™ Quenching Solution With ZyBlack™ Quenching Solution

Kidney tissue section hybridized with the ZytoLight ® SPEC PTEN/CEN 10 Dual Color Probe.

Product Description ZyBlack™ Quenching Solution is a ready- One of the major concerns of Fluores- ZyBlack™ Quenching Solution reduces to-use solution to reduce autofluorescence cence in situ Hybridization (FISH)-based autofluorescence without adversely on both formalin-fixed paraffin-embedded diagnostic assays is the interference by affecting tissue integrity or specific and frozen sections. autofluorescence. Several types of tissue fluorescence signals. It can be easily incorporated into the tend to emit intense autofluorescence, manual FISH protocol by applying it after including brain, liver, kidney and myocar- the proteolytic pretreatment. dium, making it difficult to evaluate FISH results.

Prod. No. Product Volume BS-0002-8 ZyBlack Quenching Solution 8 ml

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

187 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 187 AFI003-1-20 ZytoVision Fluorochromes and Filter Recommendations

ZytoVision Fluorochromes Fluorochrome Excitation Emission Equivalent to Two factors that mainly influence • ZyBlue™ 418 nm 467 nm DEAC FISH analyses: ZyGreen™ 503 nm 528 nm FITC · Fluorochromes of the FISH probes • ZyGold™ 532 nm 553 nm Rhodamine 6G · Appropriate filter sets • • ZyOrange™ 547 nm 572 nm Rhodamine • ZyRed™ 580 nm 599 nm TexasRed ®

Recommended Filter Sets Prod. No Product Detected Fluorochrome All filter sets are produced by well E-4030-1 DAPI Single Bandpass Filter Set v2 DAPI known manufacturers and have a E-4026-1 ZyBlue™ Single Bandpass Filter Set v2 superior-signal-to-noise ratio! • E-4012-1 ZyGreen™ Single Bandpass Filter Set v2 • E-4027-1 ZyGold™ Single Bandpass Filter Set v2 • E-4013-1 ZyOrange™ Single Bandpass Filter Set v2 • E-4017-1 ZyRed™ Single Bandpass Filter Set v2 • E-4016-1 ZyGreen™/ZyOrange™ Dual Bandpass Filter Set v2 •/• E-4010-1 DAPI/ZyGreen™/ZyOrange™ Triple Bandpass Filter Set DAPI/•/• E-4028-1 ZyBlue™/ZyGreen™/ZyOrange™ Triple Bandpass Filter Set •/•/•

Fluorescence Filter Holder Prod. No Product Compatible for Microscopes e.g.* The filter sets need to be E-4111-1 ZEISS Fluorescence Filter Holder „FL EC P&C“ Zeiss: Axio Imager, Axiostar plus, Axioskop 40 assembled in fluorescence E-4113-1 ZEISS Fluorescence Filter Holder „FL“ Zeiss: Axioplan 2, Axioskop 2, Axiophot 2 filter holder specific for the E-4121-1 OLYMPUS Fluorescence Filter Holder „U-MF 2“ Olympus: AX, AX70, BX41, BX50, BX51 respective microscope. E-4122-1 OLYMPUS Fluorescence Filter Holder „U-FF“ Olympus: BX43, BX53, BX63 E-4131-1 LEICA Fluorescence Filter Holder „DM K“ Leica: DM4000-6000, DMI4000-6000 E-4141-1 NIKON Fluorescence Filter Holder „C-FL“ Nikon: Eclipse 50i, Eclipse 80i, Eclipse TI *If your model is not listed, please contact [email protected]

Microscope Specifications In order to provide you with the best possible service, please provide us with the following details: · Microscope manufacturer · Type or model of microscope · Approx. age of microscope

These filter sets, optimized for ZytoLight ® and FlexISH ® FISH probes, will significantly increase brightness and quality of your FISH results!

188 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 188 AFI002-1-20 ZytoMation Products for automated FISH Page

Method Introduction - ZytoMation ® 190

Probes, sorted by Chromosomes 191 sorted by Gene Names 192 sorted by Indication 192

Product Data Sheets 193 f.

FISH Reagents, Fluorochromes and Filter Recommendations 186 ff.

189 189 ZytoMation Products for automated FISH

Fully automated Probes for the BOND™ Systems!

Introduction Advantages of ZytoMation ® Workflow The ZytoMation ® probes combine the • High sensitivity and specificity on fully To successfully use the ZytoMation ® known high quality of the ZytoVision automated Leica BOND™ Systems probes, the Leica BOND™ FISH Kit probes for Fluorescence in situ • Ready-to-use probes (DS9636) is required. Hybridization (FISH) with an automated • Reduced hands-on time Before starting the Leica BOND™ System, workflow. They are designed for fully • Fully automated 5 h protocol the Leica BOND™ FISH Kit has to be automated FISH to detect genetic aber- complemented with the BOND™ Enzyme rations such as translocations and ampli- Pretreatment Kit and the ready-to-use fications in formalin-fixed, paraffin- Compatible BOND™ Systems ZytoMation ® FISH probe, transferred embedded tissue sections on the Leica • Leica BOND™-III to the BOND™ Titration Kit. Prior to BOND™ Systems. • Leica BOND™-MAX evaluation, the hybridized slides should be • Leica BOND™ RXm mounted using a DAPI/DuraTect™-Solution (MT-0007-0.8/MT-0008-0.8).

Workflow Schedule

Dewaxing 1.

2. Heat Pretreatment

Stringency Wash 6.

3. Proteinase K Digestion

Denaturation and Hybridization 5.

4. Probe Application

™ Leica BOND -III RUO Leica BOND™ RXm Leica BOND™-MAX

190 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 190 MME001-1-20 ZytoMation Products for automated FISH Chromosome Index Chr. Band Product Name Product No. Quantity Page

1-5 no probes available yet

6 6q22.1 ZytoMation ROS1 Dual Color Break Apart FISH Probe Z-2298-5.1ML 5.1 ml 193

7-16 no probes available yet

17 17q12 ZytoMation ERBB2/CEN 17 Dual Color FISH Probe Z-2292-5.1ML 5.1 ml 194

18-22 no probes available yet

X, Y no probes available yet

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

191 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 191 IME001-1-20 ZytoMation Products for automated FISH Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

ERBB2 HER2, HER-2, NEU ZytoMation ERBB2/CEN 17 Dual Color FISH Probe Z-2292-5.1ML 5.1 ml 194

ROS1 MCF3, ROS ZytoMation ROS1 Dual Color Break Apart FISH Probe Z-2298-5.1ML 5.1 ml 193

Indication Index Indication Product Name Product No. Quantity Page

Solid Tumors Breast Cancer ZytoMation ERBB2/CEN 17 Dual Color FISH Probe Z-2292-5.1ML 5.1 ml 194

Gastrointestinal Cancer ZytoMation ERBB2/CEN 17 Dual Color FISH Probe Z-2292-5.1ML 5.1 ml 194

Lung Cancer ZytoMation ERBB2/CEN 17 Dual Color FISH Probe Z-2292-5.1ML 5.1 ml 194 ZytoMation ROS1 Dual Color Break Apart FISH Probe Z-2298-5.1ML 5.1 ml 193

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

192 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 192 IME001-1-20 ZytoMation Products for automated FISH

ZytoMation ® ROS1 Dual Color Break Apart FISH Probe

Background Probe Description Results The ZytoMation ® ROS1 Dual Color Break The ROS1 Dual Color Break Apart FISH In an interphase nucleus lacking an ab- Apart FISH Probe is designed to detect Probe is a mixture of two direct labeled erration involving the 6q22.1 band, two translocations involving the chromosomal probes hybridizing to the 6q22.1 band. orange/green fusion signals are expected region 6q22.1 harboring the ROS pro- The orange fluorochrome direct labeled representing two normal (non-rearranged) to-oncogene 1, receptor tyrosine kinase probe hybridizes distal, the green fluoro- 6q22.1 loci. A signal pattern consisting (ROS1, a.k.a. MCF3) gene. chrome direct labeled probe hybridizes of one orange/green fusion signal, one The ROS1 gene is located on 6q22.1 proximal to the ROS1 breakpoint region orange signal, and a separate green and encodes a receptor tyrosine kinase. at 6q22.1. signal indicates one normal 6q22.1 locus Translocations affecting ROS1 have been and one 6q22.1 locus affected by a detected in glioblastoma, cholangiocar- translocation. cinoma, and non-small cell lung cancer Isolated green signals are the result of (NSCLC). deletions distal to the ROS1 breakpoint In NSCLC several ROS1 translocation region or are due to unbalanced translo- partners have been detected all of which cations affecting this chromosomal region. result in the fusion of variably truncated ROS1 forms of e.g. TPM3, SDC4, SLC34A2, CD74, EZR, or LRIG3 to the kinase domain of ROS1. GOPC has also been Ideogram of chromosome 6 found to be fused to ROS1 in NSCLC. indicating the hybridization locations. GOPC-ROS1 fusions result from interstitial RH69070 RH104060 deletion of approx. 240 kb on 6q22.1. D6S1391E RH78348 ROS1 rearrangements are thought to 3’ 5’ 3’ 5’

define a molecular subset of NSCLC with ROS1 GOPC

distinct clinical characteristics that are ~715 kb ~215 kb similar to those observed in patients with Cen 6q22.1 Tel

ALK rearranged NSCLC. ROS1 Probe map (not to scale). First evidence suggests that administration ROS1 Dual Color Break Apart FISH Probe of ROS1 kinase inhibitors may represent a hybridized to normal interphase cells as indicated very effective therapeutic strategy in by two orange/green fusion signals per nucleus. NSCLC patients harboring activating ROS1 rearrangements. Accordingly, detection of ROS1 rearrangements using Fluorescence in situ Hybridization might be a helpful tool for the identification of patients likely to respond to ROS1 kinase targeting therapies.

References Bergethon K, et al. (2012) J Clin Oncol 30: 863-70. Birchmaier C, et al. (1987) Proc Natl Acad Sci U S A 84: 9270-74. Bos M, et al. (2013) Lung Cancer 81: 142-3. Lee SE, et al. (2015) Mod Pathol 28: 468-79. Rikova K, et al. (2007) Cell 131: 1190-203. Rimkunas VM, et al. (2012) Clin Cancer Res 18: 4449-57. Suehara Y, et al. (2012) Clin Cancer Res 18: 6599-608. Takeuchi K, et al. (2012) Nat Med 18: 378-81.

Lung cancer tissue section with translocation of the ROS1 gene as indicated by one non-rearranged orange/green fusion signal, one orange and one separate green signal.

Prod. No. Product Label Tests* (Volume) Z-2298-5.1ML ZytoMation ROS1 Dual Color Break Apart FISH Probe / up to 20 (5.1 ml)

* Using 240 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

193 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 193 ME002-1-20 ZytoMation Products for automated FISH

ZytoMation ® ERBB2/CEN 17 Dual Color FISH Probe

Background Probe Description Results The ZytoMation ® ERBB2/CEN 17 Dual The ERBB2/CEN 17 Dual Color FISH In a normal interphase nucleus, two or- Color FISH Probe is designed for the Probe is a mixture of an orange fluoro- ange and two green signals are expected. detection of ERBB2 gene amplification chrome direct labeled CEN 17 probe In a cell with amplification of the ERBB2 frequently observed in solid malignant specific for the alpha satellite centromeric gene locus, multiple copies of the green neoplasms e.g. breast cancer samples. region of chromosome 17 (D17Z1) and a signal or green signal clusters will be The ERBB2 gene (a.k.a. HER2 and NEU) green fluorochrome direct labeled ERBB2 observed. is located in the chromosomal region probe specific for the chromosomal region 17q12 and encodes a 185-190 kDa 17q12-q21.1 harboring the ERBB2 gene. transmembrane glycoprotein, p185, acting as a cellular growth factor receptor. ERBB2 is a member of the Erb-b2 receptor CEN 17 (D17Z1) ERBB2 tyrosine kinase (RTK) family, also including EGFR (ERBB1, HER1), ERBB3 (HER3), and ERBB4 (HER4). Ideogram of chromosome 17 Amplification of the proto-oncogene indicating the hybridization locations. ERBB2, observed in approximately 20% HUMUT6435 D17S1179 of all breast cancer samples, has been 5’ 3’ correlated with a poor prognosis of the disease. ERBB2 Similar results have been obtained for a ~610 kb ERBB2/CEN 17 Dual Color FISH Probe hybridized Cen 17q12-q21.1 Tel to normal interphase cells as indicated by two variety of other malignant neoplasms e.g. green and two orange signals in each nucleus. ovarian cancer, stomach cancer, and ERBB2 Probe map (not to scale). carcinomas of the salivary gland.

References Baselga J, et al. (1999) Semin Oncol 26: 78-83. Brunello E, et al. (2012) Histopathology 60: 482-8. Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Coussens L, et al. (1985) Science 230: 1132-9. Ettl T, et al. (2012) Br J Cancer 106: 719-26. Hwang CC, et al. (2011) Histopathology 59: 984-92. Hynes NE & Stern DF (1994) Biochim Biophys Acta 1198: 165-84. Moelans CB, et al. (2011) Crit Rev Oncol Hematol 80: 380-92. Park JB, et al. (1989) Cancer Res 49: 6605-9. Popescu NC, et al. (1989) Genomics 4: 362-6. Sassen A, et al. (2008) Breast Cancer Res 10: R2. Slamon DJ, et al. (1987) Science 235: 177-82. Voutsas IF, et al. (2013) Int J Radiat Biol 89: 319-25. Wolff AC, et al. (2013) J Clin Oncol 31: 3997-4013.

Breast cancer tissue section with amplification of the ERBB2 gene locus as indicated by multiple copies of the green signal in each nucleus.

Prod. No. Product Label Tests* (Volume) Z-2292-5.1ML ZytoMation ERBB2/CEN 17 Dual Color FISH Probe / up to 20 (5.1 ml)

* Using 240 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

194 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 194 ME001-1-20 Page

Method Introduction - ZytoDot ® 196 - ZytoDot 2C ® 197

Probes, sorted by Chromosomes 198 ff. sorted by Gene Names 202 f. sorted by Indication 204 f.

Product Data Sheets 206 ff.

Accessories 244 f.

195 195 Reliable and Simple Detection of Genomic Alterations using Light Microscopy!

Introduction CISH: A reliable Alternative to FISH High Signal-to-Noise Ratio The ZytoDot ® products are designed for High concordance between CISH and The ZytoDot ® probes are processed by the detection of aneuploidies and gene FISH ranging from 92-100% has been the unique ZytoVision ® Repeat Subtraction amplifications by Chromogenicin situ shown by numerous international studies Technique resulting in advanced specificity Hybridization (CISH) in formalin-fixed, for ERBB2 amplification. and less background. No further blocking paraffin-embedded tissue sections, cell of repetitive sequences is needed! samples, blood or bone marrow smears, Advantages of CISH and metaphase chromosome spreads. • Quick and easy interpretation of results ZytoDot ® Kits – Convenient Solutions comparable to IHC For making CISH analysis reliable and • Simultaneous observation of tissue user-friendly, all ZytoDot ® CISH probes morphology and CISH signals can be combined with the ZytoDot ® CISH • Storage of slides at room temperature - Implementation Kit (C-3018-40) which CISH signals are permanent includes all necessary pretreatment solu- • No costly fluorescent microscope tions, wash buffers, antibodies, chromo- needed genic substrates, counterstaining solution, mounting solution and a detailed protocol to perform successful CISH experiments. Additionally, for ERBB2 a complete kit including the probe and all necessary reagents is available.

The ZytoDot ® system uses Digoxigenin (DIG)-labeled probes 1 which are, after blocking 2, detected using a Mouse-anti-DIG antibody 3. This antibody is detected by a polymerized HRP-Goat-anti-Mouse antibody 4. The enzymatic reaction of DAB 5 leads to the formation of strong permanent brown signals that can be visualized by light microscopy using a 40x objective.

Protocol Overview

Pepsin Denature Apply Blocking Apply 2nd Pretreatment Apply Probe Wash Wash Wash Stain and cover Analyze slides digestion & Hybridize & 1st Antibody Antibody

196 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 196 MDE001-1-20 ® TM ZytoDot 2C – 2-Color CISH for the Detection of Genomic Alterations

Introduction Advantages of ZytoDot ® 2C™ ZytoDot ® 2C™ Kits – The ZytoDot ® 2C™ products are designed • Simultaneous observation of tissue mor- Standardized Solutions for the simultaneous detection of two dif- phology and CISH signals at 40x using For making CISH analysis reliable and ferent genomic targets by Chromogenic in light microscopy user-friendly, a complete ZytoDot ® 2C™ situ Hybridization (CISH) in formalin-fixed, • Two targets detected simultaneously kit is available for the detection of ERBB2 paraffin-embedded tissue sections, cell • High contrasting distinct red and green amplification. This kit includes the probe, samples, and blood or bone marrow signals all necessary pretreatment solutions, wash smears. This two color system is especially • Quick and easy interpretation of results buffers, antibodies, chromogenic substrates, useful for the differentiation of aneuploi- comparable to IHC counterstaining and mounting solution, dies from gene amplifications, and the • Standardized and complete kits and a detailed protocol. detection of deletions and translocations. • No costly fluorescent microscopeneeded For other targets, any separately available ZytoDot ® 2C™ probe can be combined with the ZytoDot ® 2C™ CISH Implemen- tation Kit resulting in target specific kit solutions.

The ZytoDot ® 2C™ system uses DIG- and DNP-labeled probe cocktails targeting different genomic sections 1 which are detected using a Mouse- anti-DIG/Rabbit-anti-DNP cocktail 2. These antibodies are detected by a unique cocktail of polymerized HRP- Goat-anti-Mouse/AP-Goat-anti-Rabbit antibodies 3. The enzymatic reaction of AP-Red 4 and HRP-Green 5 leads to the formation of strong permanent red respectively green signals that can be visualized by light microscopy using a 40x objective.

Protocol Overview

Pepsin Denature Apply Antibody Apply HRP-/ Pretreatment Apply Probe Wash Wash Wash Stain and cover Analyze slides digestion & Hybridize Cocktail AP-Polymer Cocktail

197 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 197 MDE002-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

1 1p36.3 ZytoDot 2C Glioma 1p/19q Probe Set C-3076-10/-40 10/40 tests 206 ZytoDot 2C SPEC 1p36/1q25 Probe C-3036-100/-400 100/400 µl 207 1p12 ZytoDot SPEC 1p12 Probe C-3035-400 400 µl 242 f. 1q25.3 ZytoDot 2C Glioma 1p/19q Probe Set C-3076-10/-40 10/40 tests 206 ZytoDot 2C SPEC 1p36/1q25 Probe C-3036-100/-400 100/400 µl 207

2 2p24 ZytoDot SPEC MYCN Probe C-3029-400 400 µl 209 2p23 ZytoDot 2C SPEC ALK Break Apart Probe C-3055-100/-400 100/400 µl 210 2p21 ZytoDot 2C SPEC EML4 Break Apart Probe C-3059-400 400 µl 211 2q11.2 ZytoDot SPEC 2q11 Probe C-3051-400 400 µl 242 f.

3 3p11.1-q11.1 ZytoDot CEN 3 Probe C-3045-400 400 µl 242 f. 3q27 ZytoDot 2C SPEC BCL6 Break Apart Probe C-3074-100 100 µl 212

4-5 no probes available yet

6 6p11.1-q11 ZytoDot CEN 6 Probe C-3002-400 400 µl 242 f. 6q22.1 ZytoDot 2C SPEC ROS1 Break Apart Probe C-3063-100/-400 100/400 µl 213

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

198 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 198 IDE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

7 7p11.2 ZytoDot SPEC EGFR Probe C-3007-400 400 µl 214 ZytoDot 2C SPEC EGFR/CEN 7 Probe C-3033-100/-400 100/400 µl 215 7p11.1-q11.1 ZytoDot CEN 7 Probe C-3008-400 400 µl 242 f. 7q31.2 ZytoDot 2C SPEC MET/CEN 7 Probe C-3057-400 400 µl 216

8 8p11.2 ZytoDot 2C SPEC FGFR1/CEN 8 Probe C-3050-400 400 µl 217 8p11.1-q11.1 ZytoDot CEN 8 Probe C-3016-400 400 µl 242 f. 8q24.21 ZytoDot SPEC MYC Probe C-3013-400 400 µl 218 ZytoDot 2C SPEC MYC Break Apart Probe C-3066-400 400 µl 219

9 9p21 ZytoDot 2C SPEC CDKN2A/CEN 9 Probe C-3067-400 400 µl 220

10 10q11.2 ZytoDot 2C SPEC RET Break Apart Probe C-3064-100/-400 100/400 µl 221 10q23.3 ZytoDot 2C SPEC PTEN/CEN 10 Probe C-3053-400 400 µl 222 10q26.1 ZytoDot 2C SPEC FGFR2/CEN 10 Probe C-3056-400 400 µl 223

11 11q13.3 ZytoDot 2C SPEC CCND1 Break Apart Probe C-3075-100 100 µl 224

12 12p11.1-q11 ZytoDot CEN 12 Probe C-3014-400 400 µl 242 f. 12q13.3 ZytoDot 2C SPEC DDIT3 Break Apart Probe C-3047-100 100 µl 225 12q14 ZytoDot 2C SPEC CDK4/CEN 12 Probe C-3062-400 400 µl 226 12q15 ZytoDot SPEC MDM2 Probe C-3012-400 400 µl 227 ZytoDot 2C SPEC MDM2/CEN 12 Probe C-3049-100/-400 100/400 µl 228

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

199 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 199 IDE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

13 13q12.1 ZytoDot SPEC 13q12 Probe C-3052-400 400 µl 242 f. 13q14.1 ZytoDot 2C SPEC FOXO1 Break Apart Probe C-3065-100 100 µl 229

14 14q32.3 ZytoDot 2C SPEC IGH Break Apart Probe C-3071-100 100 µl 230

15 no probes available yet

16 16p11.2 ZytoDot 2C SPEC FUS Break Apart Probe C-3054-100 100 µl 231

17 17p13 ZytoDot 2C SPEC USP6 Break Apart Probe C-3077-100 100 µl 232 17p11.1-q11.1 ZytoDot CEN 17 Probe C-3006-400 400 µl 242 f. 17q12 ZytoDot SPEC ERBB2 Probe C-3001-400 400 µl 233 ZytoDot SPEC ERBB2 Probe Kit C-3003-40 40 tests 233 ZytoDot 2C SPEC ERBB2/CEN 17 Probe C-3032-100/-400 100/400 µl 234 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit C-3022-10/-40 10/40 tests 234 ZytoDot 2C SPEC ERBB2/D17S122 Probe C-3068-100 100 µl 235 17q21.2 ZytoDot 2C SPEC TOP2A/CEN 17 Probe C-3040-400 400 µl 236

18 18q11.2 ZytoDot 2C SPEC SS18 Break Apart Probe C-3046-100 100 µl 237 18q21.3 ZytoDot 2C SPEC BCL2 Break Apart Probe C-3073-100 100 µl 238 ZytoDot 2C SPEC MALT1 Break Apart Probe C-3072-100 100 µl 239

19 19p13.3 ZytoDot 2C Glioma 1p/19q Probe Set C-3076-10/-40 10/40 tests 206 ZytoDot 2C SPEC 19q13/19p13 Probe C-3037-100/-400 100/400 µl 208 19q13.3 ZytoDot 2C Glioma 1p/19q Probe Set C-3076-10/-40 10/40 tests 206 ZytoDot 2C SPEC 19q13/19p13 Probe C-3037-100/-400 100/400 µl 208

20 no probes available yet

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

200 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 200 IDE001-1-20 Chromosome Index Chr. Band Product Name Product No. Quantity Page

21 21q22.1-q22.2 ZytoDot SPEC 21q22 Probe C-3026-400 400 µl 242 f. 21q22.2 ZytoDot 2C SPEC ERG Break Apart Probe C-3058-400 400 µl 240

22 22q12.2 ZytoDot 2C SPEC EWSR1 Break Apart Probe C-3043-100 100 µl 241

X Xp11.1-q11.1 ZytoDot CEN X Probe C-3025-400 400 µl 242 f. ZytoDot 2C CEN X/Y Probe C-3048-400 400 µl 242 f.

Y Yp11.1-q11.1 ZytoDot 2C CEN X/Y Probe C-3048-400 400 µl 242 f. Yq12 ZytoDot CEN Yq12 Probe C-3020-400 400 µl 242 f.

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

201 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 201 IDE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

ALK CD246 ZytoDot 2C SPEC ALK Break Apart Probe C-3055-100/-400 100/400 µl 210

BCL2 Bcl-2, PPP1R50 ZytoDot 2C SPEC BCL2 Break Apart Probe C-3073-100 100 µl 238

BCL6 ZNF51, LAZ3 ZytoDot 2C SPEC BCL6 Break Apart Probe C-3074-100 100 µl 212

CCND1 BCL1, PRAD1 ZytoDot 2C SPEC CCND1 Break Apart Probe C-3075-100 100 µl 224

CDK4 PSK-J3 ZytoDot 2C SPEC CDK4/CEN 12 Probe C-3062-400 400 µl 226

CDKN2A p16, ARF, INK4 ZytoDot 2C SPEC CDKN2A/CEN 9 Probe C-3067-400 400 µl 220

DDIT3 CHOP, GADD153 ZytoDot 2C SPEC DDIT3 Break Apart Probe C-3047-100 100 µl 225

EGFR HER1, ERBB1 ZytoDot SPEC EGFR Probe C-3007-400 400 µl 214 ZytoDot 2C SPEC EGFR/CEN 7 Probe C-3033-100/-400 100/400 µl 215

EML4 ROPP120 ZytoDot 2C SPEC EML4 Break Apart Probe C-3059-400 400 µl 211

ERBB2 HER2, HER-2, NEU ZytoDot SPEC ERBB2 Probe C-3001-400 400 µl 233 ZytoDot SPEC ERBB2 Probe Kit C-3003-40 40 tests 233 ZytoDot 2C SPEC ERBB2/CEN 17 Probe C-3032-100/-400 100/400 µl 234 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit C-3022-10/-40 10/40 tests 234 ZytoDot 2C SPEC ERBB2/D17S122 Probe C-3068-100 100 µl 235

ERG erg-3, p55 ZytoDot 2C SPEC ERG Break Apart Probe C-3058-400 400 µl 240

EWSR1 EWS ZytoDot 2C SPEC EWSR1 Break Apart Probe C-3043-100 100 µl 241

FGFR1 FLT2, BFGFR ZytoDot 2C SPEC FGFR1/CEN 8 Probe C-3050-400 400 µl 217

FGFR2 BEK, CD332 ZytoDot 2C SPEC FGFR2/CEN 10 Probe C-3056-400 400 µl 223

FOXO1 FKHR, FKH1 ZytoDot 2C SPEC FOXO1 Break Apart Probe C-3065-100 100 µl 229

FUS FUS1 ZytoDot 2C SPEC FUS Break Apart Probe C-3054-100 100 µl 231

IGH IGH@ ZytoDot 2C SPEC IGH Break Apart Probe C-3071-100 100 µl 230

MALT1 MLT ZytoDot 2C SPEC MALT1 Break Apart Probe C-3072-100 100 µl 239

MDM2 HDM2 ZytoDot SPEC MDM2 Probe C-3012-400 400 µl 227 ZytoDot 2C SPEC MDM2/CEN 12 Probe C-3049-100/-400 100/400 µl 228

MET HGFR, RCCP2 ZytoDot 2C SPEC MET/CEN 7 Probe C-3057-400 400 µl 216

MYC CMYC, bHLHe39, c-Myc ZytoDot SPEC MYC Probe C-3013-400 400 µl 218 ZytoDot 2C SPEC MYC Break Apart Probe C-3066-400 400 µl 219

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

202 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 202 IDE001-1-20 Gene Index

HUGO Synonym Product Name Product No. Quantity Page Name

MYCN NMYC, N-myc ZytoDot SPEC MYCN Probe C-3029-400 400 µl 209

PTEN MMAC1, TEP1 ZytoDot 2C SPEC PTEN/CEN 10 Probe C-3053-400 400 µl 222

RET HSCR1, CDHF12 ZytoDot 2C SPEC RET Break Apart Probe C-3064-100/-400 100/400 µl 221

ROS1 MCF3, ROS ZytoDot 2C SPEC ROS1 Break Apart Probe C-3063-100/-400 100/400 µl 213

SS18 SYT, SSXT ZytoDot 2C SPEC SS18 Break Apart Probe C-3046-100 100 µl 237

TOP2A TOP2 ZytoDot 2C SPEC TOP2A/CEN 17 Probe C-3040-400 400 µl 236

USP6 Tre-2, TRE17 ZytoDot 2C SPEC USP6 Break Apart Probe C-3077-100 100 µl 232

The Gene Index list includes only those probes directed against DNA sequences assigned to known genes. It does not contain probes directed against other genomic sequences as e.g. repetitive satellite DNA sequences. For a complete overview of all ZytoDot ® probes, please refer to the Chromosome Index.

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

203 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 203 IDE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Solid Tumors Brain and Neural Tumors ZytoDot 2C Glioma 1p/19q Probe Set C-3076-10/-40 10/40 tests 206 ZytoDot 2C SPEC 1p36/1q25 Probe C-3036-100/-400 100/400 µl 207 ZytoDot 2C SPEC 19q13/19p13 Probe C-3037-100/-400 100/400 µl 208 ZytoDot 2C SPEC CDKN2A/CEN 9 Probe C-3067-400 400 µl 220 ZytoDot SPEC EGFR Probe C-3007-400 400 µl 214 ZytoDot 2C SPEC EGFR/CEN 7 Probe C-3033-100/-400 100/400 µl 215 ZytoDot 2C SPEC MET/CEN 7 Probe C-3057-400 400 µl 216 ZytoDot SPEC MYCN Probe C-3029-400 400 µl 209 ZytoDot 2C SPEC PTEN/CEN 10 Probe C-3053-400 400 µl 222

Breast Cancer ZytoDot SPEC EGFR Probe C-3007-400 400 µl 214 ZytoDot 2C SPEC EGFR/CEN 7 Probe C-3033-100/-400 100/400 µl 215 ZytoDot SPEC ERBB2 Probe C-3001-400 400 µl 233 ZytoDot SPEC ERBB2 Probe Kit C-3003-40 40 tests 233 ZytoDot 2C SPEC ERBB2/CEN 17 Probe C-3032-100/-400 100/400 µl 234 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit C-3022-10/-40 10/40 tests 234 ZytoDot 2C SPEC ERBB2/D17S122 Probe C-3068-100 100 µl 235 ZytoDot 2C SPEC FGFR1/CEN 8 Probe C-3050-400 400 µl 217 ZytoDot 2C SPEC FGFR2/CEN 10 Probe C-3056-400 400 µl 223 ZytoDot SPEC MYC Probe C-3013-400 400 µl 218 ZytoDot 2C SPEC TOP2A/CEN 17 Probe C-3040-400 400 µl 236

Cervical Cancer ZytoDot SPEC MYC Probe C-3013-400 400 µl 218

Gastrointestinal Cancer ZytoDot SPEC ERBB2 Probe C-3001-400 400 µl 233 ZytoDot SPEC ERBB2 Probe Kit C-3003-40 40 tests 233 ZytoDot 2C SPEC ERBB2/CEN 17 Probe C-3032-100/-400 100/400 µl 234 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit C-3022-10/-40 10/40 tests 234 ZytoDot 2C SPEC ERBB2/D17S122 Probe C-3068-100 100 µl 235 ZytoDot SPEC MDM2 Probe C-3012-400 400 µl 227 ZytoDot 2C SPEC MDM2/CEN 12 Probe C-3049-100/-400 100/400 µl 228

Lung Cancer ZytoDot 2C SPEC ALK Break Apart Probe C-3055-100/-400 100/400 µl 210 ZytoDot 2C SPEC EML4 Break Apart Probe C-3059-400 400 µl 211 ZytoDot SPEC EGFR Probe C-3007-400 400 µl 214 ZytoDot 2C SPEC EGFR/CEN 7 Probe C-3033-100/-400 100/400 µl 215 ZytoDot SPEC ERBB2 Probe C-3001-400 400 µl 233 ZytoDot SPEC ERBB2 Probe Kit C-3003-40 40 tests 233 ZytoDot 2C SPEC ERBB2/CEN 17 Probe C-3032-100/-400 100/400 µl 234 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit C-3022-10/-40 10/40 tests 234 ZytoDot 2C SPEC ERBB2/D17S122 Probe C-3068-100 100 µl 235 ZytoDot 2C SPEC FGFR1/CEN 8 Probe C-3050-400 400 µl 217 ZytoDot 2C SPEC FGFR2/CEN 10 Probe C-3056-400 400 µl 223 ZytoDot 2C SPEC MET/CEN 7 Probe C-3057-400 400 µl 216 ZytoDot 2C SPEC RET Break Apart Probe C-3064-100/-400 100/400 µl 221 ZytoDot 2C SPEC ROS1 Break Apart Probe C-3063-100/-400 100/400 µl 213

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

204 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 204 IDE001-1-20 Indication Index Indication Product Name Product No. Quantity Page

Prostate Cancer ZytoDot 2C SPEC ERG Break Apart Probe C-3058-400 400 µl 240 ZytoDot 2C SPEC PTEN/CEN 10 Probe C-3053-400 400 µl 222

Salivary Gland Tumors ZytoDot 2C SPEC EWSR1 Break Apart Probe C-3043-100 100 µl 241

Sarcomas ZytoDot 2C SPEC ALK Break Apart Probe C-3055-100/-400 100/400 µl 210 ZytoDot 2C SPEC CDK4/CEN 12 Probe C-3062-400 400 µl 226 ZytoDot 2C SPEC DDIT3 Break Apart Probe C-3047-100 100 µl 225 ZytoDot 2C SPEC EWSR1 Break Apart Probe C-3043-100 100 µl 241 ZytoDot 2C SPEC FOXO1 Break Apart Probe C-3065-100 100 µl 229 ZytoDot 2C SPEC FUS Break Apart Probe C-3054-100 100 µl 231 ZytoDot SPEC MDM2 Probe C-3012-400 400 µl 227 ZytoDot 2C SPEC MDM2/CEN 12 Probe C-3049-100/-400 100/400 µl 228 ZytoDot SPEC MYC Probe C-3013-400 400 µl 218 ZytoDot 2C SPEC SS18 Break Apart Probe C-3046-100 100 µl 237 ZytoDot 2C SPEC USP6 Break Apart Probe C-3077-100 100 µl 232

Hematology Specific Probes Acute Myelogenous Leukemia (AML) ZytoDot CEN 8 Probe C-3016-400 400 µl 242 f.

Chronic Lymphocytic Leukemia (CLL) ZytoDot 2C SPEC BCL2 Break Apart Probe C-3073-100 100 µl 238 ZytoDot 2C SPEC CCND1 Break Apart Probe C-3075-100 100 µl 224 ZytoDot SPEC MYC Probe C-3013-400 400 µl 218

Chronic Myelogenous Leukemia (CML) ZytoDot CEN 8 Probe C-3016-400 400 µl 242 f.

Multiple Myeloma ZytoDot 2C SPEC CCND1 Break Apart Probe C-3075-100 100 µl 224 ZytoDot 2C SPEC IGH Break Apart Probe C-3071-100 100 µl 230

Myelodysplastic Syndrome (MDS) ZytoDot CEN 8 Probe C-3016-400 400 µl 242 f.

Non-Hodgkin Lymphoma, other ZytoDot 2C SPEC BCL2 Break Apart Probe C-3073-100 100 µl 238 ZytoDot 2C SPEC BCL6 Break Apart Probe C-3074-100 100 µl 212 ZytoDot 2C SPEC CCND1 Break Apart Probe C-3075-100 100 µl 224 ZytoDot 2C SPEC IGH Break Apart Probe C-3071-100 100 µl 230 ZytoDot 2C SPEC MALT1 Break Apart Probe C-3072-100 100 µl 239 ZytoDot 2C SPEC MYC Break Apart Probe C-3066-400 400 µl 219

Genetics Sex Mismatched Bone-Marrow Transplantant Management ZytoDot CEN X Probe C-3025-400 400 µl 242 f. ZytoDot CEN Yq12 Probe C-3020-400 400 µl 242 f. ZytoDot 2C CEN X/Y Probe C-3048-400 400 µl 242 f.

Prenatal, Postnatal, and Preimplantation Genetics ZytoDot SPEC 21q22 Probe C-3026-400 400 µl 242 f. ZytoDot CEN X Probe C-3025-400 400 µl 242 f. ZytoDot CEN Yq12 Probe C-3020-400 400 µl 242 f. ZytoDot 2C CEN X/Y Probe C-3048-400 400 µl 242 f.

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

205 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 205 IDE001-1-20 ZytoDot ® 2C Glioma 1p/19q Probe Set

Background Probe Description Results Deletions affecting the short arm of chro- The ZytoDot ® 2C Glioma 1p/19q Probe Using the SPEC 1p36/1q25 Probe or the mosome 1 (1p36) and the long arm of Set includes the ZytoDot ® 2C SPEC SPEC 19q13/19p13 Probe in a normal chromosome 19 (19q13) are frequently 1p36/1q25 Probe and the ZytoDot ® 2C interphase nucleus, two red and two green found in human gliomas. SPEC 19q13/19p13 Probe for the detec- signals are expected. In a cell with dele- According to the 2016 WHO criteria tion of both 1p36 and 19q13 loci. tions affecting the 1p36 or 19q13 locus, for classification of tumors of the central one or no copy of the red signal will be ZytoDot ® 2C SPEC 1p36/1q25 Probe nervous system, the detection of 1p/19q observed. D1S1384 SHGC-31390 loss is required for the diagnosis of 3’ 5’ WHO grade II or III “oligodendroglioma, CHD5 IDH-mutant and 1p/19q codeleted”. Since both, astrocytomas and oligodendroglio- ~365 kb Tel 1p36.31 Cen mas, can exhibit IDH mutations, evaluation SPEC 1p36 Probe map (not to scale). of 1p/19q status plays a critical role in differentiating astrocytoma from oligoden- RH70423 D1S2355 droglioma. Oligodendroglioma morphology, IDH-mutant genotype, and 1p/19q codeletion ~190 kb are associated with better response to Cen 1q25.3 Tel chemotherapy and improved survival. SPEC 1q25 Probe map (not to scale). SPEC 1p36/1q25 Probe hybridized to glioma tissue section with 1p36 deletion as Hence, determination of 1p and 19q sta- ZytoDot ® 2C SPEC 19q13/19p13 Probe indicated by one red signal in each nucleus. tus may aid in therapeutic decisions and predict outcome in patients with diffuse RH76260 PMC196911P4 gliomas.

References Barbashina V, et al. (2005) Clin Cancer Res 11: 1119-28. ~145 kb Cairncross JG, et al. (1998) J Natl Cancer Inst 90: 1473-9. Cairncross G, et al. (2013) J Clin Oncol 31: 337-43. Tel 19p13.3 Cen Griffin CA, et al. (2006) J Neuropathol Exp Neurol 65: 988-94. Louis DN, et al. (ed.) (2016) WHO Classification of Tumours of the SPEC 19p13 Probe map (not to scale). Central Nervous System (Revised 4th Edition). Reifenberger G, et al. (2017) Nat Rev Clin Oncol 14: 434-52. Rosenberg JE, et al. (1996) Oncogene 13: 2483-5. RH68396 WI5375 Smith JS, et al. (1999) Oncogene 18: 4144-52. 5’ 3’ 5’ 3’ 5’ 3’ Smith JS, et al. (2000) Genes Chromosomes Cancer 29: 16-25. GLTSCR1 GLTSCR2 CRX

~480 kb Cen 19q13.32-q13.33 Tel SPEC 19q13/19p13 Dual Color Probe hybridized to glioma tissue section with 19q13 deletion as SPEC 19q13 Probe map (not to scale). indicated by one red signal in each nucleus.

Images kindly provided by Prof. W. Müller, University Leipzig, Germany.

Prod. No. Product Label Tests* (Volume) C-3076-10 ZytoDot 2C Glioma 1p/19q Probe Set 10 Incl. ZytoDot 2C SPEC 1p36/1q25 Probe, 0.1 ml; ZytoDot 2C SPEC 19q13/19p13 Probe, 0.1 ml

C-3076-40 ZytoDot 2C Glioma 1p/19q Probe Set 40 Incl. ZytoDot 2C SPEC 1p36/1q25 Probe, 0.4 ml; ZytoDot 2C SPEC 19q13/19p13 Probe, 0.4 ml Related Products C-3036-100/-400 ZytoDot 2C SPEC 1p36/1q25 Probe DNP/DIG 10/40 (100 µl/400 µl) C-3037-100/-400 ZytoDot 2C SPEC 19q13/19p13 Probe DNP/DIG 10/40 (100 µl/400 µl) C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

206 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 206 DE044-1-20 ZytoDot ® 2C SPEC 1p36/1q25 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC 1p36/1q25 The ZytoDot ® 2C SPEC 1p36/1q25 In a normal interphase nucleus, using the Probe is designed for the detection of 1p Probe is a mixture of a Dinitrophenyl-labe- ZytoDot ® 2C SPEC 1p36/1q25 Probe deletions. led 1p36 probe specific for the smallest in combination with ZytoDot ® 2C CISH Deletions affecting the short arm of chro- region of consistent deletion (SRD) of Implementation Kit, two red (1p) and two mosome 1 (1p) are frequently found in chromosome 1 defined in neuroblastoma green (1q) signals are expected. In a cell human gliomas and neuroblastomas, but at 1p36.31 and a Digoxigenin-labeled with deletions affecting the 1p36 locus, also in breast, lung, endometrial, ovarian, 1q25 probe specific for 1q25.3. one or no copy of the red signal will be and colorectal carcinomas. observed. Deletions affecting the long arm of chromosome 19 (19q) are frequently found in human malignant gliomas as well as in neuroblastomas and epithelial ovarian cancers. Combined loss of the complete 1p/19q chromosome arms, caused by an unbalanced t(1;19)(q10;p10) translocation, is characteristic of oligodendrogliomas. According to the 2016 WHO criteria for classification of tumors of the central nervous system, the detection of 1p/19q loss is required for the diagnosis of WHO grade II or III “oligodendroglioma, Ideogram of chromosome 1 SPEC 1p36/1q25 Probe hybridized to indicating the hybridization locations. glioma tissue section with 1p36 deletion as IDH-mutant and 1p/19q codeleted”. Since indicated by one red signal in each nucleus. D1S1384 SHGC-31390 both, astrocytomas and oligodendroglio- Image kindly provided by Prof. W. Müller, University Leipzig, Germany. 3’ 5’ mas, can exhibit IDH mutations, evaluation of 1p/19q status plays a critical role in CHD5 differentiating astrocytoma from oligoden- ~365 kb droglioma. Tel 1p36.31 Cen SPEC 1p36 Probe map (not to scale). References Oligodendroglioma morphology, IDH-mu- Barbashina V, et al. (2005) Clin Cancer Res 11: 1119-28. Cairncross JG, et al. (1998) J Natl Cancer Inst 90: 1473-9. tant genotype, and 1p/19q codeletion RH70423 D1S2355 Cairncross G, et al. (2013) J Clin Oncol 31: 337-43. Caron H, et al. (1996) N Engl J Med 334: 225-30. are associated with better response to Griffin CA, et al. (2006) J Neuropathol Exp Neurol 65: 988-94. Louis DN, et al. (ed.) (2016) WHO Classification of Tumours of the chemotherapy and improved survival. Central Nervous System (Revised 4th Edition). Lass U, et al. (2013) Brain Pathol 23: 311-8. Hence, determination of 1p and 19q sta- ~190 kb Ragnarsson G, et al. (1999) Br J Cancer 79: 1468-74. Reifenberger G, et al. (2017) Nat Rev Clin Oncol 14: 434-52. tus may aid in therapeutic decisions and Cen 1q25.3 Tel Rosenberg JE, et al. (1996) Oncogene 13: 2483-5. Smith JS, et al. (1999) Oncogene 18: 4144-52. predict outcome in patients with diffuse SPEC 1q25 Probe map (not to scale). Smith JS, et al. (2000) Genes Chromosomes Cancer 29: 16-25. gliomas.

Prod. No. Product Label Tests* (Volume) C-3036-100 ZytoDot 2C SPEC 1p36/1q25 Probe DNP/DIG 10 (100 μl) C-3036-400 ZytoDot 2C SPEC 1p36/1q25 Probe DNP/DIG 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

207 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 207 DE026-1-20 ZytoDot ® 2C SPEC 19q13/19p13 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC 19q13/19p13 The ZytoDot ® 2C SPEC 19q13/19p13 Using the ZytoDot ® 2C SPEC 19q13/ Probe is designed for the detection of 19q Probe is a mixture of a Dinitrophenyl-la- 19p13 Probe in combination with the deletions. beled 19q13 probe specific for the ZytoDot ® 2C CISH Implementation Kit, Deletions affecting the short arm of chro- region of common deletion in gliomas at two red (19q) and two green (19p) sig- mosome 1 (1p) are frequently found in 19q13.32-q13.33 and a Digoxigenin-la- nals are expected in a normal interphase human gliomas and neuroblastomas, but beled 19p13 probe specific for 19p13.3. nucleus. In a cell with deletions affecting also in breast, lung, endometrial, ovarian, the 19q13 locus, one or no copy of the and colorectal carcinomas. red signal will be observed. Deletions affecting the long arm of chromosome 19 (19q) are frequently found in human malignant gliomas as well as in neuroblastomas and epithelial ovarian Ideogram of chromosome 19 indicating the hybridization locations. cancers. Combined loss of the complete 1p/19q RH76260 PMC196911P4 chromosome arms, caused by an unbalanced t(1;19)(q10;p10) translocation, is characteristic of oligodendrogliomas. ~145 kb According to the 2016 WHO criteria Tel 19p13.3 Cen for classification of tumors of the central SPEC 19p13 Probe map (not to scale).

nervous system, the detection of 1p/19q RH68396 WI5375 loss is required for the diagnosis of 5’ 3’ 5’ 3’ 5’ 3’ SPEC 19q13/19p13 Dual Color Probe hybridized WHO grade II or III “oligodendroglioma, GLTSCR1 GLTSCR2 CRX to glioma tissue section with 19q13 deletion as indicated by one red signal in each nucleus. IDH-mutant and 1p/19q codeleted”. Since ~480 kb both, astrocytomas and oligodendroglio- Cen 19q13.32-q13.33 Tel Image kindly provided by Prof. W. Müller, University Leipzig, Germany.

mas, can exhibit IDH mutations, evaluation SPEC 19q13 Probe map (not to scale). of 1p/19q status plays a critical role in differentiating astrocytoma from oligodendroglioma.

Oligodendroglioma morphology, IDH-mu- References Barbashina V, et al. (2005) Clin Cancer Res 11: 1119-28. tant genotype, and 1p/19q codeletion Cairncross JG, et al. (1998) J Natl Cancer Inst 90: 1473-9. Cairncross G, et al. (2013) J Clin Oncol 31: 337-43. are associated with better response to Caron H, et al. (1996) N Engl J Med 334: 225-30. Griffin CA, et al. (2006) J Neuropathol Exp Neurol 65: 988-94. chemotherapy and improved survival. Louis DN, et al. (ed.) (2016) WHO Classification of Tumours of the Central Nervous System (Revised 4th Edition). Hence, determination of 1p and 19q sta- Lass U, et al. (2013) Brain Pathol 23: 311-8. Ragnarsson G, et al. (1999) Br J Cancer 79: 1468-74. tus may aid in therapeutic decisions and Reifenberger G, et al. (2017) Nat Rev Clin Oncol 14: 434-52. Rosenberg JE, et al. (1996) Oncogene 13: 2483-5. predict outcome in patients with diffuse Smith JS, et al. (1999) Oncogene 18: 4144-52. gliomas. Smith JS, et al. (2000) Genes Chromosomes Cancer 29: 16-25.

Prod. No. Product Label Tests* (Volume) C-3037-100 ZytoDot 2C SPEC 19q13/19p13 Probe DNP/DIG 10 (100 μl) C-3037-400 ZytoDot 2C SPEC 19q13/19p13 Probe DNP/DIG 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

208 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 208 DE043-1-20 ZytoDot ® SPEC MYCN Probe

Background Probe Description Results The ZytoDot ® SPEC MYCN Probe is The ZytoDot ® SPEC MYCN Probe is a In normal cells, two distinct dot-shaped designed for the detection of MYCN Digoxigenin-labeled probe specific for the signals per nucleus will be observed. Nu- amplification which represents the most MYCN gene at 2p24.3, processed by the clei with amplification of the MYCN gene powerful unfavorable prognostic factor for unique ZytoVision ® Repeat Subtraction locus or aneuploidy of chromosome 2 will neuroblastoma. Less frequently amplifica- Technique resulting in advanced specificity show multiple dots or large signal clusters. tions are found in retinoblastoma, small and less background. cell lung cancer, astrocytoma and other tumors derived from the neuroectoderm. MYCN The MYCN (MYCN proto-oncogene, bHLH transcription factor, a.k.a. NMYC) gene is located in the chromosomal region 2p24.3 and encodes a 62-64 kDa transcription factor normally expressed in the developing nervous system and other selected tissues. The MYCN oncogene is amplified in about 25% of primary neuroblastomas and 90% of tumor-derived cell lines. Additional copies are rarely located at the Ideogram of chromosome 2 Normal nuclei each with two MYCN signals. normal locus but are detected as double indicating the hybridization locations. minute chromosomes or homogeneously SHGC-140713 GDB:315969 staining regions. 5’3’ Amplification of the MYCN gene is strongly associated with rapid tumor pro- MYCN gression, advanced stages of the disease, ~370 kb Tel 2p24.3 Cen and poor prognosis. Hence, amplification status is increasingly being used for strat- SPEC MYCN Probe map (not to scale). ification of patients to different treatment protocols.

References Kaneko M, et al. (1998) Med Pediatr Oncol 31: 1-7. Lee WH, et al. (1984) Nature 309: 458-60. Maris JM, et al. (2007) Lancet 369: 2106-20. Slamon DJ, et al. (1986) Science 232: 768-72. Suita S, et al. (2007) J Pediatr Surg 42: 489-93. Thorner PS, et al. (2006) Am J Surg Pathol 30: 635-42.

Prod. No. Product Label Tests* (Volume) C-3029-400 ZytoDot SPEC MYCN Probe DIG 40 (400 μl) Related Products C-3018-40 ZytoDot CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

209 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 209 DE010-1-20 ZytoDot ® 2C SPEC ALK Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC ALK Break Apart The ZytoDot ® 2C SPEC ALK Break Apart In an interphase nucleus of a normal Probe is designed to detect rearrangements Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the involving the chromosomal region 2p23.2 beled probe and a Dinitrophenyl-labeled 2p23.2 band, using the ZytoDot ® 2C harboring the ALK (ALK receptor tyrosine probe hybridizing to the 2p23.2 band. CISH Implementation Kit, two red/green kinase, a.k.a. CD246) gene. The Digoxigenin-labeled probe hybridizes fusion signals are expected representing ALK encodes a transmembrane receptor proximal to the ALK gene breakpoint re- two normal (non-rearranged) 2p23.2 loci. tyrosine kinase. This gene exerts charac- gion at 2p23.2, the Dinitrophenyl-labeled A signal pattern consisting of one red/ teristic oncogenic activities through fusion probe hybridizes distal to the ALK gene green fusion signal, one red signal, and a to several gene partners or mutations both breakpoint region at 2p23.2. separate green signal indicates one nor- in hematopoietic and non-hematopoietic mal 2p23.2 locus and one 2p23.2 locus solid tumors. affected by a translocation or inversion. Translocations affecting the ALK gene EML4-ALK inversion with deletion of locus are frequently found in anaplastic 5’-ALK sequences is indicated by one or large cell lymphoma (ALCL), an aggres- multiple isolated red signals. sive non-Hodgkin lymphoma arising from T-cells. The most frequent translocation t(2;5) results in a fusion with the NPM1 (nucleophosmin a.k.a. nucleolar phos- phoprotein B23, numatrin) gene located on chromosome 5q35. This rearrangement results in a NPM1/ALK fusion protein, which is constitutively activated through Ideogram of chromosome 2 autophosphorylation, and that in turn indicating the hybridization locations. mediates malignant cell transformation by activating downstream effectors like e.g. D2S2934 D2S405 RH12489 RH35185 STAT3. 3’ 5’ Additionally, inversions affecting the ALK ALK gene located on the short arm of chro- Lung carcinoma tissue section with translocation ~210 kb ~220 kb affecting the 2p23.2 locus as indicated by mosome 2 [inv(2)(p21p23)] have been one red/green fusion (non-rearranged) signal, Tel 2p23.2 Cen frequently detected in non-small cell lung one red signal, and one separate green signal. SPEC ALK Probe map (not to scale). cancer (NSCLC) and lead to the formation of EML4-ALK fusion transcripts.

ALK kinase targeted therapies may repre- References Inamura K, et al. (2009) Mod Pathol 22: 508-15. sent a very effective therapeutic strategy Koivunen JP, et al. (2008) Clin Cancer Res 14: 4275-83. Martelli MP, et al. (2009) Am J Pathol 174: 661-70. in NSCLC patients carrying EML4-ALK Palmer RH, et al. (2009) Biochem J 420: 345-61. Perner S, et al. (2008) Neoplasia 10: 298-302. rearrangements. Rodig SJ, et al. (2009) Clin Cancer Res 15: 5216-23. Sasaki T, et al. (2010) Eur J Cancer 46: 1773-80. Schildhaus HU, et al. (2013) Mod Pathol 26: 1468-77. von Laffert M, et al. (2014) J Thorac Oncol 9: 1464-9. Wagner F, et al. (2014) J Clin Pathol 67: 403-7. Zhang Q, et al. (2007) Nat Med 11: 1341-8.

Prod. No. Product Label Tests* (Volume) C-3055-100 ZytoDot 2C SPEC ALK Break Apart Probe DIG/DNP 10 (100 μl) C-3055-400 ZytoDot 2C SPEC ALK Break Apart Probe DIG/DNP 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

210 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 210 DE025-1-20 ZytoDot ® 2C SPEC EML4 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC EML4 Break Apart The ZytoDot ® 2C SPEC EML4 Break Apart In an interphase nucleus of a normal Probe is designed to detect rearrange- Probe is a mixture of a Digoxigenin-labe- cell lacking a translocation involving the ments involving the chromosomal region led and a Dinitrophenyl-labeled probe hy- 2p21 band, using the ZytoDot ® 2C CISH 2p21 harboring the EML4 (echinoderm bridizing to the 2p21 band. The DNP-la- Implementation Kit, two red/green fusion microtubule-associated protein-like 4, beled probe hybridizes proximal to the signals are expected representing two nor- a.k.a. ROPP120) gene. EML4 gene breakpoint region at 2p21, mal (non-rearranged) 2p21 loci. A signal Inversions in the short arm of chromosome the DIG-labeled probe hybridizes distal to pattern consisting of one red/green fusion 2 [inv(2)(p21p23)] have been frequently the EML4 gene breakpoint region. signal, one red signal, and a separate detected in non-small cell lung cancer green signal indicates one normal 2p21 (NSCLC) and lead to the formation of locus and one 2p21 locus affected by a EML4-ALK fusion transcripts. A few reports translocation or inversion. also identified these fusion transcripts in breast, gastric, and colorectal cancers. EML4 belongs to the family of echinoderm microtubule-associated protein-like proteins. The EML4-ALK fusion transcripts comprise variably truncated N-terminal portions of the EML4 gene and the intracellular signaling domain of the ALK receptor tyrosine kinase (a.k.a. CD246). It was found that EML4 mediates ligand-in- Ideogram of chromosome 2 dependent dimerization of ALK, resulting indicating the hybridization locations. in constitutive kinase activity. EML4-ALK was demonstrated to possess transforming D2S2932 RH81694 RH120737 D2S1482 SPEC EML4 Break Apart Probe hybridized to activity in vitro and in vivo. 5’ 3’ normal interphase cells as indicated by two The EML4-ALK fusion transcript is found red/green fusion signals pert nucleus. EML4 in about 5% of NSCLC, predominantly ~125 kb ~590 kb adenocarcinomas, and is considered to Tel 2p21 Cen be mutually exclusive to EGFR or KRAS SPEC EML4 Probe map (not to scale). mutations. The detection of the inversion by in situ Hybridization might represent a valuable tool to identify a subpopulation of NSCLC likely to respond to ALK kinase targeting therapies. References Choi YL, et al. (2008) Cancer Res 69: 4971-6. Inamura K, et al. (2009) Mod Pathol 22: 508-15. Lin E, et al. (2009) Mol Cancer Res 7: 1466-76. Perner S, et al. (2008) Neoplasia 10: 298-302. Rodig SJ, et al. (2009) Clin Cancer Res 15: 5216-23. Schildhaus HU, et al. (2013) Mod Pathol 26: 1468-77. Shaw AT, et al. (2009) J Clin Oncol 27: 4247-53. Soda M, et al. (2007) Nature 448: 561-6.

Prod. No. Product Label Tests* (Volume) C-3059-400 ZytoDot 2C SPEC EML4 Break Apart Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

211 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 211 DE031-1-20 ZytoDot ® 2C SPEC BCL6 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC BCL6 Break Apart The ZytoDot ® 2C SPEC BCL6 Break Apart In an interphase nucleus of a normal Probe is designed for the detection of Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the translocations involving the chromosomal beled and a Dinitrophenyl-labeled probe 3q27.3-q28 band, using the ZytoDot ® 2C region 3q27.3 harboring the BCL6 (BCL6 hybridizing to the 3q27.3-q28 band. The CISH Implementation Kit, two red/green transcription repressor, a.k.a. ZNF51, DNP-labeled probe hybridizes distal to the fusion signals are expected representing LAZ3) gene. BCL6 gene at 3q27.3-q28, the DIG-labe- two normal (non-rearranged) 3q27.3-q28 The BCL6 protein acts as a transcriptional led probe hybridizes proximal to the BLC6 loci. A signal pattern consisting of one repressor that is involved in the regulation gene at 3q27.3. red/green fusion signal, one red signal, of lymphoid development and function. and a separate green signal indicates Chromosomal rearrangements of the one normal 3q27.3-q28 locus and one BCL6 gene region were found to occur 3q27.3-q28 locus affected by a translo- in different types of non-Hodgkin lympho- cation. ma (NHL), including diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL). The most common BCL6 translocation t(3;14)(q27;q32.3) results in the IGH-BCL6 gene fusion. In addition, more than 20 partner loci have been identified BCL6 including immunoglobulin (Ig) genes but also a number of non-Ig genes. As a result Ideogram of chromosome 3 indicating the hybridization locations. of these translocations, the rearranged BCL6 gene comes under the control of the RH122573 RH123158 D3S2492 RH106114 promoter of the partner gene leading to 3’ 5’ deregulated expression of BCL6. In DLBCL, the most common histologic BCL6 subtype of NHL, BCL6 translocations rep- ~185 kb ~195 kb SPEC BCL6 Break Apart Probe hybridized Cen 3q27.3-q28 Tel to normal interphase cells as indicated by resent one of the most frequent cytogenetic two red/green fusion signals per nucleus. SPEC BCL6 Probe map (not to scale). abnormality, occurring in 20% to 40% of the cases. Several studies reported a correlation of BCL6 translocation with an inferior overall survival. Moreover, DLBCL which are positive for both BCL6 and MYC rearrangements have been shown to have an extremely poor prognosis. Hence, the detection of BCL6 rearrangements by CISH may help in predicting the clinical outcome in patients with NHL. References Akyurek N, et al. (2012) Cancer 118: 4173-83. Cady FM, et al. (2008) J Clin Oncol 26: 4814-9. Ohno H (2004) Histol Histopathol 19: 637-50. Ohno H (2006) J Clin Exp Hematop 46: 43-53.

Prod. No. Product Label Tests* (Volume) C-3074-100 ZytoDot 2C SPEC BCL6 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

212 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 212 DE040-1-20 ZytoDot ® 2C SPEC ROS1 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC ROS1 Break The ZytoDot ® 2C SPEC ROS1 Break In an interphase nucleus of a normal Apart Probe is designed to detect translo- Apart Probe is a mixture of a Digoxi- cell lacking an aberration involving the cations involving the chromosomal region genin-labeled and a Dinitrophenyl-labeled 6q22.1 band, using the ZytoDot ® 2C 6q22.1 harboring the ROS proto-onco- probe hybridizing to the 6q22.1 band. CISH Implementation Kit, two red/green gene 1, receptor tyrosine kinase (ROS1, The DNP-labeled probe hybridizes distal fusion signals are expected representing a.k.a. MCF3) gene. to the ROS1 gene breakpoint region at two normal (non-rearranged) 6q22.1 loci. The ROS1 gene is located on 6q22.1 6q22.1, the DIG-labeled probe hybridizes A signal pattern consisting of one red/ and encodes a receptor tyrosine kinase. proximal to the ROS1 gene breakpoint green fusion signal, one red signal, and a Translocations affecting ROS1 have been region. separate green signal indicates one nor- detected in glioblastoma, cholangiocar- mal 6q22.1 locus and one 6q22.1 locus cinoma, and non-small cell lung cancer affected by a translocation. (NSCLC). Isolated green signals are the result of In NSCLC several ROS1 translocation deletions distal to the ROS1 breakpoint partners have been detected all of which region or are due to unbalanced translo- result in the fusion of variably truncated cations affecting this chromosomal region. forms of e.g. TPM3, SDC4, SLC34A2, ROS1 CD74, EZR, or LRIG3 to the kinase domain of ROS1. GOPC has also been found to be fused to ROS1 in NSCLC. Ideogram of chromosome 6 GOPC-ROS1 fusions result from interstitial indicating the hybridization locations. deletion of approx. 240 kb on 6q22.1. RH69070 RH104060 ROS1 rearrangements have been exclu- BV208912 RH78348 sively detected in adenocarcinoma of the 3’ 5’ 3’ 5’

lung and are thought to define a molecu- ROS1 GOPC

lar subset of NSCLC with distinct clinical ~180 kb ~215 kb characteristics that are similar to those Cen 6q22.1 Tel SPEC ROS1 Break Apart Probe hybridized to normal interphase cells as indicated by observed in patients with ALK rearranged SPEC ROS1 Probe map (not to scale). two red/green fusion signals per nucleus. NSCLC. First evidence suggests that administration of ROS1 kinase inhibitors may represent a very effective therapeutic strategy in NSCLC patients harboring activating ROS1 rearrangements. Accordingly, detection of ROS1 rearrangements using

Chromogenic in situ Hybridization might References Bergethon K, et al. (2012) J Clin Oncol 30: 863-70. be a helpful tool for the identification of Bos M, et al. (2013) Lung Cancer 81: 142-3. Rikova K, et al. (2007) Cell 131: 1190-203. patients likely to respond to ROS1 kinase Rimkunas VM, et al. (2012) Clin Cancer Res 18: 4449-57. Suehara Y, et al. (2012) Clin Cancer Res 18: 6599-608. targeting therapies. Takeuchi K, et al. (2012) Nat Med 18: 378-81. Lung cancer tissue section with rearrangement of the ROS1 gene as indicated by isolated green signals.

Prod. No. Product Label Tests* (Volume) C-3063-100 ZytoDot 2C SPEC ROS1 Break Apart Probe DIG/DNP 40 (400 μl) C-3063-400 ZytoDot 2C SPEC ROS1 Break Apart Probe DIG/DNP 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

213 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 213 DE022-1-20 ZytoDot ® SPEC EGFR Probe

Background Probe Description Results The ZytoDot ® SPEC EGFR Probe is The ZytoDot ® SPEC EGFR Probe is a In normal cells, two distinct dot-shaped designed for the detection of EGFR gene Digoxigenin-labeled probe specific for the signals per nucleus will be observed. amplification frequently observed in solid EGFR gene at 7p11.2, processed by the Nuclei with amplification of the EGFR neoplasms including non-small cell lung unique ZytoVision ® Repeat Subtraction gene locus or aneuploidy of chromo- cancer (NSCLC) and glioblastoma. The Technique resulting in advanced specificity some 7 will show multiple dots or large EGFR gene (a.k.a. ERBB1 and HER1) and less background. signal clusters. is located in the chromosomal region 7p11.2 and encodes a transmembrane glycoprotein acting as a cellular growth factor receptor. Overexpression of EGFR has been shown in a number of tumor entities and is associated with poor prognosis. EGFR copy number identified byin situ Hybridization is thought to be a molecular predictor in Ideogram of chromosome 7 neoplasms. indicating the hybridization locations. References Balla P, et al. (2011) Histopathology 59: 376-89. Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. RH122798 SHGC-146751 Ettl T, et al. (2012) Hum Pathol 43: 921-31. 5’ 3’ Gont A, et al. (2013) Oncotarget 4: 1266-79. Isola J & Tanner M (2004) Methods Mol Med 97: 133-44. Normal nuclei each with two EGFR signals. Kondo I & Shimizu N (1983) Cytogenet Cell Genet 35: 9-14. EGFR Laurent-Puig P, et al. (2009) J Clin Oncol 27: 5924-30. Marquez A, et al. (2004) Diagn Mol Pathol 13: 1-8. ~345 kb Miyazawa H, et al. (2008) Cancer Sci 99: 595-600. Tel 7p11.2 Cen Oikawa M, et al. (2013) J Oral Pathol Med 42: 424-34. Thomas F, et al. (2007) Clin Cancer Res 13: 7086-92. Tsukamoto T, et al. (1991) Int J Dev Biol 35: 25-32. SPEC EGFR Probe map (not to scale). Walker F, et al. (2009) Hum Pathol 40: 1517-27. Yoo SB, et al. (2010) Lung Cancer 67: 301-5. Zaczek A, et al. (2005) Histol Histopathol 20: 1005-15.

Cancer cells with multiple EGFR signals in sputum sample from a NSCLC patient.

Prod. No. Product Label Tests* (Volume) C-3007-400 ZytoDot SPEC EGFR Probe DIG 40 (400 μl) Related Products C-3018-40 ZytoDot CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

214 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 214 DE002-1-20 ZytoDot ® 2C SPEC EGFR/CEN 7 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC EGFR/CEN 7 The ZytoDot ® 2C SPEC EGFR/CEN 7 In a normal interphase nucleus, using the Probe is designed for the simultaneous Probe is a mixture of a Digoxigenin-labe- ZytoDot ® 2C CISH Implementation Kit two detection of EGFR and centromere 7 in led probe specific for the EGFR gene at green and two red signals are expected. formalin-fixed, paraffin-embedded tissue 7p11.2 and a Dinitrophenyl-labeled CEN In a cell with amplification of the EGFR sections and cell samples. 7 probe specific for the alpha satellite cen- gene locus, multiple copies of the green The EGFR gene (a.k.a. ERBB1 and HER1) tromeric region of chromosome 7 (D7Z1). signal or green signal clusters will be is located in the chromosomal region observed. 7p11.2 and encodes a transmembrane glycoprotein acting as a cellular growth factor receptor. Overexpression of EGFR has been shown in a number of tumor entities and is associated with poor prognosis. EGFR copy number identified byin situ Hybridization is thought to be a molecular predictor in Ideogram of chromosome 7 neoplasms. indicating the hybridization locations. References Balla P, et al. (2011) Histopathology 59: 376-89. RH70813 SHGC-148394 Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. 5’ 3’ Ettl T, et al. (2012) Hum Pathol 43: 921-31. Isola J & Tanner M (2004) Methods Mol Med 97: 133-44. Kondo I & Shimizu N (1983) Cytogenet Cell Genet 35: 9-14. EGFR Marquez A, et al. (2004) Diagn Mol Pathol 13: 1-8. Tsukamoto T, et al. (1991) Int J Dev Biol 35: 25-32. ~200 kb Normal nuclei each with two EGFR (green) and two Zaczek A, et al. (2005) Histol Histopathol 20: 1005-15. Tel 7p11.2 Cen centromere 7 (red) signals.

SPEC EGFR Probe map (not to scale).

Trisomy of chromosome 7 as indicated by three EGFR (green) and three CEN 7 (red) signals in each nucleus.

Prod. No. Product Label Tests* (Volume) C-3033-100 ZytoDot 2C SPEC EGFR/CEN 7 Probe DIG/DNP 10 (100 μl) C-3033-400 ZytoDot 2C SPEC EGFR/CEN 7 Probe DIG/DNP 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

215 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 215 DE011-1-20 ZytoDot ® 2C SPEC MET/CEN 7 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC MET/CEN 7 Probe The ZytoDot ® 2C SPEC MET/CEN 7 In a normal interphase nucleus, using the is designed for the detection of MET gene Probe is a mixture of a Dinitrophenyl-la- ZytoDot ® 2C CISH Implementation Kit two amplifications found in a variety of human beled CEN 7 probe specific for the alpha red (CEN 7) and two green (MET) signals tumors. satellite centromeric region of chromo- are expected. In a cell with amplification The MET gene (a. k. a. c-Met) is located some 7 (D7Z1) and a Digoxigenin-labeled of the MET gene locus, multiple copies of in the chromosomal region 7q31.2 and probe specific for the chromosomal region the green signal or green signal clusters encodes a transmembrane tyrosine kinase 7q31.2 harboring the MET gene. will be observed. receptor for the hepatocyte growth factor (HGF). HGF and MET play an important role in angiogenesis and tumor growth. Activation or upregulation of MET was CEN 7 (D7Z1) found in a number of carcinomas including lung, breast, colorectal, prostate, and gastric carcinomas as well as in gliomas, MET melanomas and some sarcomas. MET overexpression is known as a negative prognostic indicator in patients with Ideogram of chromosome 7 indicating the hybridization locations. various carcinomas, multiple myeloma, or glioma. Therefore, several inhibitors of the REN66320 STSG598383 5’ 3’ HGF/MET signaling pathway are being Lung cancer tissue section with multiple studied and developed as potent therapies MET copies of chromosome 7 (red) and extra to inhibit angiogenesis and tumor growth. ~420 kb MET signals (green) in the nuclei. Recently, it was shown that MET ampli- Cen 7q31.2 Tel fication leads to resistance to gefitinib SPEC MET Probe map (not to scale). or erlotinib in lung cancer by driving ERBB3-dependent activation of the PI3K pathway.

References Cooper CS, et al. (1984) Nature 311: 29-32. Engelman JA, et al. (2007) Science 316: 1039-43. Garcia S, et al. (2007) Int J Oncol 31: 49-58. Hara T, et al. (1998) Lab Invest 78: 1143-53.

Prod. No. Product Label Tests* (Volume) C-3057-400 ZytoDot 2C SPEC MET/CEN 7 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

216 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 216 DE028-1-20 ZytoDot ® 2C SPEC FGFR1/CEN 8 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC FGFR1/CEN The ZytoDot ® 2C SPEC FGFR1/CEN 8 In a normal interphase nucleus, using the 8 Probe is designed for the detection Probe is a mixture of a Digoxigenin-labe- ZytoDot ® 2C CISH Implementation Kit two of FGFR1 gene amplification frequently led probe specific for the FGFR1 gene at green (FGFR1) and two red (CEN 8) sig- observed in malignant tumors e.g. breast 8p11.23-p11.22 and a Dinitrophenyl-la- nals are expected. In a cell with an ampli- and prostate cancer and oral squamous beled CEN 8 probe specific for the alpha fication of the FGFR1 gene locus, multiple cell carcinoma (OSCC). satellite centromeric region of chromo- copies of the green signal or green signal The FGFR1 (fibroblast growth factor recep- some 8 (D8Z2). clusters will be observed. tor 1) gene is located in the chromosomal region 8p11.23-p11.22 and encodes a transmembrane receptor tyrosine kinase. FGFR1 Amplification of the FGFR1 gene, ob- CEN 8 (D8Z2) served in approximately 10% of all breast cancer samples, has revealed to be an independent prognostic factor for overall survival. FGFR1 is believed to emerge as a potential therapeutic target for lobular Ideogram of chromosome 8 breast carcinomas. indicating the hybridization locations.

In prostate cancer, FGFR1 gene amplifica- D8S135 D8S414 tion seems to be an important step during 3’ 5’ 3’ 5’

the transmission to hormone resistance. WHSC1L1 FGFR1 In OSCC, FGFR1 gene amplification, Breast carcinoma tissue section with FGFR1 ~270 kb amplification as indicated by large green clusters. observed in nearly 20% of all cases, is Tel 8p11.22-p11.23 Cen indicated to contribute to oral carcinogen- SPEC FGFR1 Probe map (not to scale). esis at an early stage of development.

References Edwards J, et al. (2003) Clin Cancer Res 9: 5271-81. Elbauomy Elsheikh S, et al. (2007) Breast Cancer Res 9: R23. Freier K, et al. (2007) Oral Oncology 43: 60-6. Lee PL, et al. (1989) Science 245: 57-60. Lacroix-Triki M, et al. (2010) J Pathol 222: 282-98. Swoboda A, et al. (2011) Genes Chromosomes Cancer 50: 680-8. Turner N, et al. (2010) Cancer Res 70: 2085-94. Wetterskog D, et al. (2012) J Pathol 226: 84-96.

Prod. No. Product Label Tests* (Volume) C-3050-400 ZytoDot 2C SPEC FGFR1/CEN 8 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

217 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 217 DE024-1-20 ZytoDot ® SPEC MYC Probe

Background Probe Description Results The ZytoDot ® SPEC MYC Probe is The ZytoDot ® SPEC MYC Probe is a In normal cells, two distinct dot-shaped designed for the detection of MYC gene Digoxigenin-labeled probe specific for the signals per nucleus will be observed. amplification frequently observed in malig- MYC gene region at 8q24.21, processed Nuclei with amplification of the MYC gene nant tumors e.g. breast and endometrial by the unique ZytoVision ® Repeat Sub- locus or polysomy of chromosome 8 cancer. traction Technique resulting in advanced will show multiple dots or large signal The proto-oncogene MYC (a.k.a. CMYC) specificity and less background. clusters. is located in the chromosomal region 8q24.21 and encodes a nuclear transcription factor displaying high-affinity, site specific DNA-binding capacity when com- plexed with its cellular partners. Thus, the MYC protein is involved in proliferation, growth, differentiation, and apoptosis. MYC Amplification of the chromosomal MYC gene region has been detected in many Ideogram of chromosome 8 indicating the hybridization locations. types of malignant neoplasms e.g. breast,

lung, head, colon, kidney, neck, ovary, SHGC-140056 SHGC-144185 bladder, and endometrial cancers. It was 5’3’

shown that MYC amplification occurs MYC Tetrasomy of chromosome 8 as indicated in advanced, widespread tumors or in ~290 kb by four MYC signals per nucleus. aggressive, primary tumors. In non-small Cen 8q24.21 Tel cell lung cancer (NSCLC) and breast SPEC MYC Probe map (not to scale). cancer, for example, MYC amplification was strongly associated with lymph node status. Accordingly, the MYC gene can be considered as a powerful prognostic marker. Additionally, malignant cutaneous angiosarcomas but not benign and atypical vascular lesions occurring after radiother- apy of breast cancer are characterized by amplification of the MYC gene. The presence of MYC amplification is thus of considerable diagnostic importance for the distinction of malignant from atypical postradiation vascular neoplasms of the skin. References Alves Rde C, et al. (2014) J Cancer Res Clin Oncol 140: 2021-5. Butt AJ, et al. (2005) Endocr Relat Cancer 12: 47-59. Dalla-Favera R, et al. (1982) Proc Natl Acad Sci U S A 79: 6497-501. Deming SL, et al. (2000) Br J Cancer 83: 1688-95. Hara T, et al. (1998) Lab Invest 78: 1143-53. Kubokura H, et al. (2001) Ann Thorac Cardiovasc Surg 7: 197-203. Mentzel T, et al. (2012) Mod Pathol 25: 75-95. Rummukainen JK, et al. (2001) Lab Invest 81: 1445-51. Schraml P, et al. (1999) Clin Cancer Res 5: 1966-75. Yokota J, et al. (1986) Science 231: 261-5.

Prod. No. Product Label Tests* (Volume) C-3013-400 ZytoDot SPEC MYC Probe DIG 40 (400 μl) Related Products C-3018-40 ZytoDot CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

218 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 218 DE004-1-20 ZytoDot ® 2C SPEC MYC Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC MYC Break Apart The ZytoDot ® 2C SPEC MYC Break Apart In an interphase nucleus of a normal Probe is designed to detect transloca- Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the tions involving the chromosomal region beled and a Dinitrophenyl-labeled probe 8q24.21 band, using the ZytoDot ® 2C 8q24.21 harboring the MYC gene. hybridizing to the 8q24.21 band. The CISH Implementation Kit, two red/green The MYC proto-oncogene (MYC proto-on- DNP-labeled probe hybridizes proximal fusion signals are expected representing cogene, bHLH transcription factor, a.k.a. to the MYC gene breakpoint region at two normal (non-rearranged) 8q24.21 CMYC) encodes a transcription factor 8q24.21, the DIG-labeled probe hybrid- loci. A signal pattern consisting of one essential for cell growth and proliferation izes distal to the MYC gene breakpoint red/green fusion signal, one red signal, and is broadly implicated in tumorigen- region. and a separate green signal indicates one esis. Translocations involving the MYC normal 8q24.21 locus and one 8q24.21 gene are considered to be cytogenetic locus affected by a translocation. hallmarks for Burkitt lymphoma but are Alternative break points particularly also found in other types of lymphomas. observed in variant MYC translocations The most frequent translocation involv- t(8;22) and t(2;8) might result in different ing the MYC gene region is t(8;14) signal patterns. (q24.21;q32.3) juxtaposing the MYC gene in 8q24.21 next to the IGH (immunoglobulin heavy chain) locus in Ideogram of chromosome 8 indicating the hybridization locations. 14q32.33. Further translocations affecting the MYC gene are t(8;22)(q24.21;q11.2) SHGC-130556 RH75025 D8S1090 RH118867 and t(2;8)(p11.2;q24.21), both of which 5’ 3’ involve one of the two immunoglobulin MYC light chain loci. All three translocations bring the MYC gene under the control ~475 kb ~475 kb Cen 8q24.21 Tel of a regulatory element from one of the SPEC MYC Probe map (not to scale). immunoglobulin loci resulting in SPEC MYC Break Apart Probe hybridized to normal interphase cells as indicated by constitutive overexpression of MYC. two red/green fusion signals per nucleus. References Boerma EG, et al. (2009) Leukemia 23: 225-34. Dalla-Favera R, et al. (1982) Proc Natl Acad Sci U S A 79: 6497-501. Haralambieva E, et al. (2004) Genes Chromosomes Cancer 40: 10-8. Veronese ML, et al. (1995) Blood 85: 2132-8. Walker BA, et al. (2014) Blood Cancer J 4: e191.

Non-Hodgkin lymphoma tissue section with translocation affecting the 8q24.21 locus as indicated by one red/green fusion (non-rearranged) signal, one red signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) C-3066-400 ZytoDot 2C SPEC MYC Break Apart Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

219 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 219 DE032-1-20 ZytoDot ® 2C SPEC CDKN2A/CEN 9 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC CDKN2A/CEN 9 The ZytoDot ® 2C SPEC CDKN2A/CEN 9 In a normal interphase nucleus, using the Probe is designed for the detection of Probe is a mixture of a Digoxigenin-labe- ZytoDot ® 2C CISH Implementation Kit two CDKN2A deletions frequently observed in led probe specific for the CDKN2A gene green (CDKN2A) and two red (CEN 9) most tumor cell lines as well as in primary at 9p21.3 and a Dinitrophenyl-labeled signals are expected. In a cell with human malignancies. CEN 9 probe specific for the classical deletion of the CDKN2A gene locus, a The CDKN2A gene, often referred to as satellite III region of chromosome 9 (D9Z3) reduced number of green signals will be p16 or INK4a/ARF, is located in the chro- at 9q12. observed. Deletions affecting only parts mosomal region 9p21.3. Using alternative of the CDKN2A gene might result in a first exons and an alternative reading normal signal pattern with green signals CDKN2A frame, the gene encodes for two distinct of reduced size. tumor suppressor proteins p16INK4a CEN 9 (D9Z3) and p14ARF, both involved in cell cycle regulation. CDKN2A has been identified as a major susceptibility gene for melanoma. The tumor suppressor gene CDKN2A is inactivated by homozygous deletions Ideogram of chromosome 9 indicating the hybridization locations. with high frequency in a variety of human primary tumors e.g. bladder and renal cell D9S1749 RH98751 5’ 3’ 3’ 5’ 3’5’ carcinoma, prostate and ovarian adenocarcinoma, non-small cell lung cancer, MTAP sarcoma, glioma, mesothelioma, and CDKN2A CDKN2B ~315 kb melanoma. Furthermore, deletion of the Tel 9p21.3 Cen

CDKN2A gene is found in up to 80% of SPEC CDKN2A Probe map (not to scale). SPEC CDKN2A/CEN 9 Probe hybridized to T-cell acute lymphoblastic leukemia cases normal interphase cells as indicated by two red and two green signals per nucleus. and is associated with poor prognosis and relapse of the disease.

References Arif Q & Husain AN (2015) Arch Pathol Lab Med 139: 978-80. Cowan JM et al. (1988) J NatI Cancer Inst 80: 1159-64. Holley T, et al. (2012) PLoS One 7: e50586. Hussussian CJ, et al. (1994) Nat Genet 8: 15-21. Kamb A, et al. (1994) Science 264: 436-40. Nobori T, et al. (1994) Nature 368: 753-6. Quelle DE, et al. (1995) Cell 83: 993-1000. Rocco JW & Sidransky D (2001) Exp Cell Res 264: 42-55. Schopmeyer K, et al. (1999) Neoplasia 1: 128-37. Schwarz S, et al. (2008) Cytometry A 73: 305-11. Sharpless NE (2005) Mutat Res 576: 22-38.

Prod. No. Product Label Tests* (Volume) C-3067-400 ZytoDot 2C SPEC CDKN2A/CEN 9 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

220 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 220 DE034-1-20 ZytoDot ® 2C SPEC RET Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC RET Break Apart The ZytoDot ® 2C SPEC RET Break Apart In an interphase nucleus of a normal Probe is designed to detect translocations Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the involving the chromosomal region 10q11.21 beled and a Dinitrophenyl-labeled probe 10q11.21 band, using the ZytoDot ® 2C harboring the RET (ret proto-oncogene) hybridizing to the 10q11.21 band. The CISH Implementation Kit, two red/green gene. RET encodes a tyrosine kinase (TK) DNP-labeled probe hybridizes proximal fusion signals are expected representing receptor. to the RET gene breakpoint region at two normal (non-rearranged) 10q11.21 Translocations involving RET were first 10q11.21, the DIG-labeled probe hy- loci. A signal pattern consisting of one described in papillary thyroid carcinoma bridizes distal to the RET gene breakpoint red/green fusion signal, one red signal, (PTC) where somatic rearrangements result region. and a separate green signal indicates in the fusion of its TK catalytic domain with one normal 10q11.21 locus and one an N-terminal dimerization domain encoded 10q11.21 locus affected by a transloca- by various fusion partner genes. tion or inversion. More recently, recurrent inversions RET [inv(10)(p11.2q11.2)] fusing the coiled-coil domains of the kinesin family member 5B (KIF5B) gene to the RET kinase domain have been detected in lung adenocarcinoma. Ideogram of chromosome 10 The resulting KIF5B-RET fusion protein can indicating the hybridization locations. form homodimers through the coiled-coil D10S141 RH102119 domains of KIF5B, causing an aberrant acti- D10S2320 D10S1179 vation of the TK of RET, a mechanism known 5’ 3’

from KIF5B-ALK fusions which is also found RET

in lung adenocarcinoma. ~170 kb ~170 kb Since in vitro studies showed transforming Cen 10q11.21 Tel

activity of KIF5B-RET which could be sup- SPEC RET Probe map (not to scale). SPEC RET Break Apart Probe hybridized to pressed by a TK inhibitor, it was assumed normal interphase cells as indicated by two red/green fusion signals per nucleus. that the chimeric oncogene might be a promising molecular target for the treatment of lung cancer. The same holds true for the BCR-RET and FGFR1OP-RET fusion genes in chronic myelomonocytic leukemia (CMML) generated by two balanced translocations t(10;22) (q11.2;q11.2) and t(6;10)(q27;q11.2), respectively. References Ballerini P, et al. (2012) Leukemia 26: 2384-9. Gautschi O, et al. (2013) J Thorac Oncol 8: e43-4. Ju YS, et al. (2012) Genome Res 22: 436-45. Kohno T, et al. (2012) Nat Med 18: 375-7. Nikiforov YE (2002) Endocr Pathol 13: 3-16. Takahashi M, et al. (1985) Cell 42: 581-8. Takeuchi K, et al. (2012) Nat Med 18: 378-81.

Prod. No. Product Label Tests* (Volume) C-3064-100 ZytoDot 2C SPEC RET Break Apart Probe DIG/DNP 10 (100 μl) C-3064-400 ZytoDot 2C SPEC RET Break Apart Probe DIG/DNP 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

221 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 221 DE035-1-20 ZytoDot ® 2C SPEC PTEN/CEN 10 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC PTEN/CEN 10 The ZytoDot ® 2C SPEC PTEN/CEN 10 In a normal interphase nucleus, using the Probe is designed for the detection of Probe is a mixture of a Dinitrophenyl-labe- ZytoDot ® 2C CISH Implementation Kit PTEN deletions frequently observed in led CEN 10 probe specific for the alpha two red (CEN 10) and two green (PTEN) many tumor types, including renal, mela- satellite centromeric region of chromo- signals are expected. In a cell with a noma, endometrial, breast, prostate, lung, some 10 (D10Z1) and a Digoxigenin-la- deletion of the PTEN gene locus a reduced bladder, and thyroid cancer but also in beled probe specific for the chromosomal number of green signals will be observed. hematological neoplasms. region 10q23.2-q23.31 harboring the Deletions affecting only parts of the PTEN The tumor suppressor gene PTEN (phos- PTEN gene. gene might result in normal signal pattern phatase and tensin homolog), often with green signals of reduced size. referred to as MMAC1 (mutated in multiple advanced cancers 1), is located CEN 10 (D10Z1) on 10q23.31 and encodes a 47 kDa dual-specificity phosphatase that has both lipid and protein phosphatase activity. Its PTEN inactivation results in constitutive activation of the PI3K/AKT pathway and in subse- Ideogram of chromosome 10 quent increase in protein synthesis, cell indicating the hybridization locations. cycle progression, migration, and survival. RH17273 RH74520 Deletions affecting the long arm of chro- 5’ 3’ mosome 10 have been detected in 30 to 50% of early and advanced stage spo- PTEN radic melanomas and about 40 to 70% ~315 kb Cen 10q23.2-q23.31 Tel of prostate cancers. In both tumor entities Prostate cancer tissue section with deletion of the SPEC PTEN Probe map (not to scale). PTEN gene as indicated by one green signal. loss of PTEN has been associated with poor clinical outcome. Currently, several drugs targeting the PI3K/AKT pathway for the therapy of solid tumors have entered clinical trials. References Dahia PLM, et al. (1999) Hum Mol Genet 8: 185-93. Healy E, et al. (1998) Oncogene 16: 2213-8. Li J, et al. (1997) Science 275: 1943-7. Swoboda A, et al. (2011) Genes Chromosomes Cancer 50: 680-8. Weng LP, et al. (2001) Hum Mol Genet 10: 599-604. Yoshimoto M, et al. (2006) Cancer Genet Cytogenet 169: 128-37. Yoshimoto M, et al. (2007) Br J Cancer 97: 678-85.

Prod. No. Product Label Tests* (Volume) C-3053-400 ZytoDot 2C SPEC PTEN/CEN 10 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

222 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 222 DE023-1-20 ZytoDot ® 2C SPEC FGFR2/CEN 10 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC FGFR2/CEN 10 The ZytoDot ® 2C SPEC FGFR2/CEN 10 In a normal interphase nucleus, using Probe is designed for the detection of Probe is a mixture of a Dinitrophenyl-labe- the ZytoDot ® 2C CISH Implementation FGFR2 gene amplifications frequently led CEN 10 probe specific for the alpha Kit two red (CEN 10) and two green observed in breast cancer as well as in satellite centromeric region of chromosome (FGFR2) signals are expected. Nuclei with gastric cancer. 10 (D10Z1) and a Digoxigenin-labeled amplification of the FGFR2 gene locus The FGFR2 (fibroblast growth factor probe specific for the chromosomal region at 10q26.12-q26.13 or polysomy of receptor 2, a.k.a. BEK) gene is located 10q26.12-q26.13 harboring the FGFR2 chromosome 10 will show multiple copies on chromosome 10q26.13 and encodes gene. of the green signal or large green signal splice variants of the receptor tyrosine clusters. kinases FGFR2b and FGFR2c. Amplification of the FGFR2 gene leads to CEN 10 (D10Z1) overexpression of the FGFR2 protein and subsequently to signal activation. Addition- ally, during the amplification process the C-terminal deletion of FGFR2 can occur FGFR2 due to exclusion of the last exon from Ideogram of chromosome 10 the FGFR2 amplicon. Both, overexpres- indicating the hybridization locations. sion and deletion of the last exon result CHLC.ATA44G07 SHGC-146065 in FGFR2 signaling activation based on 3’ 5’ constitutive phosphorylation of the FRS2 FGFR2 adaptor molecule. ~410 kb The process of ligand independent FGFR2 Cen 10q26.12-q26.13 Tel Breast carcinoma tissue section signaling leads to a more severe malig- SPEC FGFR2 Probe map (not to scale). with FGFR2 (green) amplification. nant phenotype of these tumors. Moreo- ver, high FGFR2 expression is correlated with poor overall survival (OS) and poor disease-free survival (DFS) rates in breast cancer patients. Consequently, FGFR2 gene amplification detected by Chromoge- nic in situ Hybridization might be used as a prognostic marker in breast cancer.

References Azuma K, et al. (2011) Biochem Biophys Res Commun 407: 219-24. Katoh M (2010) Expert Rev Anticancer Ther 10: 1375-9. Katoh Y & Katoh M (2009) Int J Mol Med 23: 307-11. Moffa AB, et al. (2004) Mol Cancer Res 2: 643-52. Sun S, et al. (2012) J Surg Oncol 105: 773-9

Prod. No. Product Label Tests* (Volume) C-3056-400 ZytoDot 2C SPEC FGFR2/CEN 10 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

223 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 223 DE027-1-20 ZytoDot ® 2C SPEC CCND1 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC CCND1 Break The ZytoDot ® 2C SPEC CCND1 Break In an interphase nucleus of a normal Apart Probe is designed to detect translo- Apart Probe is a mixture of a Digoxi- cell lacking a translocation involving the cations involving the chromosomal region genin-labeled and a Dinitrophenyl-labeled 11q13.3 band, using the ZytoDot ® 2C 11q13.3 harboring the CCND1 gene. probe hybridizing to the 11q13.3 band. CISH Implementation Kit, two red/green The CCND1 gene (cyclin D1, a.k.a. The DNP-labeled probe hybridizes distal fusion signals are expected representing PRAD1) encodes a regulatory subunit of to the CCND1 gene breakpoint region at two normal (non-rearranged) 11q13.3 cyclin-dependent kinases. 11q13.3, the DIG-labeled probe hybridiz- loci. A signal pattern consisting of one Translocations involving the chromosomes proximal to the CCND1 gene break- red/green fusion signal, one red signal, al region t(11;14)(q13.3;q32.3) are point region. and a separate green signal indicates one considered to be characteristic for mantle normal 11q13.3 locus and one 11q13.3 cell lymphomas (MCL) but have also been locus affected by a translocation. identified in other lymphoproliferative disorders (LPDs), such as B-prolymphocytic leukemia, and, less frequently, in plasma CCND1 cell myelomas, B-cell chronic lymphocytic leukemia, and in splenic lymphomas with villous lymphocytes (SLVL). Ideogram of chromosome 11 The t(11;14) rearrangement often leads indicating the hybridization locations. to overexpression of the CCND1 protein. RH47914 PMC133966P1 Determination of translocations involving RH51714 RH48109 the chromosomal region 11q13.3 can 5’ 3’

also help to distinguish MCL from other CCND1

chronic lymphoproliferative disorders. ~185 kb ~580 kb Since the course of MCL is aggressive, Cen 11q13.3 Tel SPEC CCND1 Break Apart Probe hybridized and its response to chemotherapy is poor, SPEC CCND1 Probe map (not to scale). to normal interphase cells as indicated by differential diagnosis is clinically impor- two red/green fusion signals per nucleus. tant. Additionally, it was also shown that a renal oncocytoma (RO) specific breakpoint is located in band 11q13.3, involving the CCND1 locus. The histologic features of RO may overlap with those of chromophobe renal cell carcinoma (ChRCC). CISH can be used as a diagnostic tool for differentiation of RO from ChRCC. References Bentz JS, et al. (2004) Cancer 102: 124-31. Bosch F, et al. (1997) Cancer 82: 567-75. Sinke RJ, et al. (1997) Cancer Genet Cytogenet 96: 95-101. Sukov WR, et al. (2007) Hum Pathol 40: 1296-303. Tarsitano M, et al. (2009) Cancer Genet Cytogenet 195: 164-7. Tsujimoto Y, et al. (1984) Science 224: 1403-6. Vaandrager JW, et al. (1996) Blood 4: 1177-82. Mantle cell lymphoma tissue section with translocation affecting the 11q13.3 locus as indicated by one non-rearranged red/green fusion signal, one red signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) C-3075-100 ZytoDot 2C SPEC CCND1 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

224 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 224 DE042-1-20 ZytoDot ® 2C SPEC DDIT3 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC DDIT3 Break The ZytoDot ® 2C SPEC DDIT3 Break In an interphase nucleus of a normal Apart Probe is designed to detect translo- Apart Probe is a mixture of a Digoxi- cell lacking a translocation involving cations involving the chromosomal region genin-labeled and a Dinitrophenyl-labeled the 12q13.3-q14.1 band, using the 12q13.3 harboring the DDIT3 (DNA dam- probe hybridizing to the 12q13.3-q14.1 ZytoDot ® 2C CISH Implementation Kit, age inducible transcript 3) gene (a.k.a. band. The DNP-labeled probe hybridiz- two red/green fusion signals are expected CHOP, GADD153) in formalin-fixed, es proximal to the DDIT3 gene and the representing two normal (non-rearranged) paraffin-embedded tissue sections or cell DIG-labeled probe hybridizes distal to that 12q13.3-q14.1 loci. A signal pattern con- samples. gene. sisting of one red/green fusion signal, one The DDIT3 gene encodes for a stress-in- red signal, and a separate green signal induced dominant-negative inhibitor of dicates one normal 12q13.3-q14.1 locus the transcription factors C/EBP and and one 12q13.3-q14.1 locus affected by LAP. DDIT3 is consistently rearranged in DDIT3 a translocation or inversion. myxoid liposarcomas (MLS). The most frequent translocation involving the DDIT3 gene region is t(12;16)(q13.3;p11.2) and occurs in about 90% of patients with MLS. Ideogram of chromosome 12 The rearrangement results in a fusion gene indicating the hybridization locations. comprising the 5´ part of the FUS (fused in sarcoma) gene, located in 16p11.2, and RH39898 D9S2056 RH52872 RH110894 the complete coding region of the DDIT3 3’5’ gene. The FUS-DDIT3 fusion protein acts DDIT3 as an abnormal transcription factor and ~480 kb ~460 kb development of myxoid liposarcomas is Cen 12q13.3-q14.1 Tel thus regarded as a consequence of dereg- SPEC DDIT3 Probe map (not to scale). ulated FUS-DDIT3 target genes. SPEC DDIT3 Break Apart Probe hybridized Differential diagnosis of liposarcomas and to normal interphase cells as indicated by two red/green fusion signals per nucleus. accurate classification, the latter being especially important with regard to appropriate treatment and prognosis, are often problematic. Therefore, detection of DDIT3 rearrangements via ISH analysis is a valuable tool to confirm the histopathological diagnosis of myxoid liposarcoma.

References Ǻman P, et al. (1992) Genes Chromosomes Cancer 5: 278-85. Andersson M, et al. (2010) BMC Cancer 10: 249-58. Germano G, et al. (2010) Cancer Res 70: 2235-44. Meis-Kindblom JM, et al. (2001) Virchows Arch 439: 141-51. Panagopoulos I, et al. (1994) Cancer Res 54: 6500-3. Ron D & Habener JF (1992) Genes Dev 6: 439-53.

Myxoid liposarcoma tissue section with translocation affecting the 12q13.3-q14.1 locus as indicated by one non-rearranged red/green fusion signal, one red signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) C-3047-100 ZytoDot 2C SPEC DDIT3 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

225 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 225 DE017-1-20 ZytoDot ® 2C SPEC CDK4/CEN 12 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC CDK4/CEN 12 The ZytoDot ® 2C SPEC CDK4/CEN 12 In a normal interphase nucleus, using the Probe is designed for the detection of Probe is a mixture of a Digoxigenin-labe- ZytoDot ® 2C CISH Implementation Kit, CDK4 gene amplifications. The cyclin-de- led probe specific for the chromosomal two green (CDK4) and two red (CEN pendent kinase 4 (CDK4) gene is located region 12q13.3-q14.1 harboring the 12) signals are expected. In a cell with in the chromosomal region 12q14.1, CDK4 gene and a Dinitrophenyl-labeled amplification of the CDK4 gene locus or ~10 Mb centromeric to the murine double CEN 12 probe specific for the alpha satel- polysomy of chromosome 12, multiple minute (MDM2) gene and is frequently lite centromeric region of chromosome 12 copies of the green signal or green signal coamplified with MDM2 in different malig- (D12Z3). clusters will be observed. nancies. In a complex with cyclin D1 (CCND1), the CDK4 encoded serine/threonine kinase CEN 12 (D12Z3) phosphorylates the retinoblastoma protein CDK4 1 (RB1) which in turn leads to the release of the EF2 transcription factor and subsequently to an upregulation of genes which are required for progression through the Ideogram of chromosome 12 S-, G2-, and M-phases of the cell cycle. indicating the hybridization locations. Due to amplification of the respective chromosomal region, CDK4 is overexpressed RH94136 RH45075 3’ 5’ in many human tumors such as soft tissue sarcomas, osteosarcomas (OS), and CDK4 ~310 kb Liposarcoma tissue section with CDK4 gliomas. In glioblastomas, the lack of amplification as indicated by large green clusters. Cen 12q13.3-q14.1 Tel amplification of several genes like CDK4 was recognized to be associated with a SPEC CDK4 Probe map (not to scale). longer survival time. In OS, coamplification of MDM2 and CDK4, located in two discontinuous regions, occurs frequently in parosteal OS and less often in classical high-grade OS. Although MDM2/CDK4 coamplification is not restricted to atypical lipomatous tumors/well-differentiated liposarcomas (ALT/ WDLPS) and dedifferentiated liposarcomas (DDLPS), its detection is a strong cri- terion for distinguishing these tumor types References from other undifferentiated sarcomas and Binh MB, et al. (2005) Am J Surg Pathol 29: 1340-7. Fischer U, et al. (2010) Int J Cancer 126: 2594-602. even from carcinomas and lymphomas. Lee SE, et al. (2014) Histol Histopathol 29: 127-38. Lopes MA, et al. (2001) Oral Oncol 37: 566-71. Moreover, CDK4 amplification is a poor Mejia-Guerrero S, et al. (2010) Genes Chromosomes Cancer 49: 518-25. Sirvent A, et al. (2007) Am J Surg Pathol 31: 1476-89. prognostic factor in WDLPS and DDLPS. Wunder JS, et al. (1999) Oncogene 18: 783-8.

Prod. No. Product Label Tests* (Volume) C-3062-400 ZytoDot 2C SPEC CDK4/CEN 12 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

226 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 226 DE021-1-20 ZytoDot ® SPEC MDM2 Probe

Background Probe Description Results The ZytoDot ® SPEC MDM2 Probe is de- The ZytoDot ® SPEC MDM2 Probe is a In normal cells, two distinct dot-shaped sig- signed for the detection of MDM2 gene Digoxigenin-labeled probe specific for the nals per nucleus will be observed. Nuclei amplifications found in more than 10% of MDM2 gene region at 12q15, processed with amplification of the MDM2 gene human tumors. by the the unique ZytoVision ® Repeat Sub- locus or polysomy of chromosome 12 will The MDM2 (MDM2 proto-oncogene) gene traction Technique resulting in advanced show multiple dots or large signal clusters. is located in the chromosomal region specificity and less background. 12q15 and encodes for an E3 ubiquitin ligase which acts as a major negative regulator of the tumor suppressor p53. Due to the amplification of the respective chromosomal region, MDM2 is overexpressed in many human tumors such as soft tissue sarcomas, osteosarcomas, gliomas, NSCLC, gastric and breast Ideogram of chromosome 12 carcinomas. Well-differentiated liposarco- indicating the hybridization locations. mas (WDLPS), the most common soft tissue tumors in adults, are characterized by the B611O2/T7 B611O2/SP6 5’ 3’ amplification of 12q-derived chromosomal Normal nuclei each with two MDM2 signals. material, harboring the MDM2 oncogene MDM2 while lipomas show balanced transloca- ~240 kb Cen Tel tions involving 12q13-15. Accordingly, 12q15 detection of the 12q14-15 amplification SPEC MDM2 Probe map (not to scale). is regarded as a valuable tool for the differential diagnosis between well-differentiated liposarcomas and lipomas. Fur- thermore, detection of the MDM2 amplification might have prognostic relevance in gastrointestinal stromal tumors (GIST), the most common primary mesenchymal tumor of the gastrointestinal tract.

References Korcheva VB, et al. (2011) Appl Immunhistochem Mol Morphol 19: 119-25. Larousserie F, et al. (2013) Eur J Radiol 82: 2149-53. Luan SL, et al. (2010) J Pathol 222: 166-79. Momand J, et al. (1992) Cell 69: 1237-45. Oliner JD, et al. (1992) Nature 358: 80-3. Pedeutour F, et al. (1994) Genes Chromosomes Cancer 10: 85-94. Pedeutour F, et al. (2004) Bull Cancer 91: 317-23. Poaty H, et al. (2012) PLoS One 7: e29426. Toledo F & Wahl GM (2006) Nat Rev Cancer 6: 909-23. Tornillo L, et al. (2005) Lab Invest 85: 921-31. Vassilev LT (2007) Trends Mol Med 13: 23-31.

Prod. No. Product Label Tests* (Volume) C-3012-400 ZytoDot SPEC MDM2 Probe DIG 40 (400 μl) Related Products C-3018-40 ZytoDot CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

227 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 227 DE013-1-20 ZytoDot ® 2C SPEC MDM2/CEN 12 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC MDM2/CEN 12 The ZytoDot ® 2C SPEC MDM2/CEN 12 In a normal interphase nucleus,using the Probe is designed for the simultaneous Probe is a mixture of a Digoxigenin-labeled ZytoDot ® 2C CISH Implementation Kit two detection of MDM2 and centromere 12 in probe specific for MDM2 gene at 12q15 red and two green signals are expected. formalin-fixed, paraffin-embedded tissue and a Dinitrophenyl-labeled CEN 12 In a cell with amplification of the MDM2 sections or cell samples. probe specific for the alpha satellite gene locus, multiple copies of the green The MDM2 (MDM2 proto-oncogene) gene centromeric region of chromosome 12 signal or green signal clusters will be is located in the chromosomal region (D12Z3). observed. 12q15 and encodes for an E3 ubiquitin ligase which acts as a major negative regulator of the tumor suppressor p53. MDM2 gene amplifications are found in more than 10% of human tumors. Due to the amplification of the respective chromosomal region, MDM2 is overexpressed in many human tumors such as soft tissue Ideogram of chromosome 12 sarcomas, osteosarcomas, gliomas, indicating the hybridization locations. NSCLC, gastric and breast carcinomas. Well-differentiated liposarcomas (WDLPS), B611O2/T7 B611O2/SP6 5’ 3’ the most common soft tissue tumors in adults, are characterized by the amplifica- MDM2 ~240 kb Liposarcoma tissue section with MDM2 tion of 12q-derived chromosomal material, amplification as indicated by large green clusters. Cen Tel harboring the MDM2 oncogene while 12q15 lipomas show balanced translocations in- SPEC MDM2 Probe map (not to scale). volving 12q13-15. Accordingly, detection of the 12q14-15 amplification is regarded as a valuable tool for the differential diagnosis between well-differentiated liposarcomas and lipomas. Furthermore, detection of the MDM2 amplification might have prognostic relevance in gastrointestinal stromal tumors (GIST), the most common primary mesenchymal tumor of the gastrointestinal tract. References Flanagan A, et al. (2010) Skeletal Radiol 39: 213-24. Kashima T, et al. (2012) Mod Pathol 25: 1384-96. Korcheva VB, et al. (2012) Appl Immunhistochem Mol Morphol 19: 119-25. Larousserie F, et al. (2013) Eur J Radiol 82: 2149-53. Luan SL, et al. (2010) J Pathol 222: 166-79. Momand J, et al. (1992) Cell 69: 1237-45. Oliner JD, et al. (1992) Nature 358: 80-3. Pedeutour F, et al. (1994) Genes Chromosomes Cancer 10: 85-94. Pedeutour F, et al. (2004) Bull Cancer 91: 317-23. Poaty H, et al. (2012) PLoS One 7: e29426. Toledo F & Wahl GM (2006) Nat Rev Cancer 6: 909-23. Tornillo L, et al. (2005) Lab Invest 85: 921-31. Vassilev LT (2007) Trends Mol Med 13: 23-31. Prod. No. Product Label Tests* (Volume) C-3049-100 ZytoDot 2C SPEC MDM2/CEN 12 Probe DIG/DNP 10 (100 μl) C-3049-400 ZytoDot 2C SPEC MDM2/CEN 12 Probe DIG/DNP 40 (400 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

228 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 228 DE016-1-20 ZytoDot ® 2C SPEC FOXO1 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC FOXO1 Break The ZytoDot ® 2C SPEC FOXO1 Break In an interphase nucleus of a normal Apart Probe is designed for the detection Apart Probe is a mixture of a Digoxi- cell lacking a translocation involving the of specific translocations involving the genin-labeled and a Dinitrophenyl-labeled 13q14.11 band, using the ZytoDot ® 2C chromosomal region 13q14.11 harboring probe hybridizing to the 13q14.11 band. CISH Implementation Kit, two red/green the FOXO1 (forkhead box O1, a.k.a. The DNP-labeled probe hybridizes distal fusion signals are expected representing FKHR) gene characteristic for alveolar to the FOXO1 gene breakpoint region two normal (non-rearranged) 13q14.11 rhabdomyosarcoma. at 13q14.11, the DIG-labeled probe loci. A signal pattern consisting of one Among solid tumors of the childhood, hybridizes proximal to the FOXO1 gene red/green fusion signal, one red signal, rhabdomyosarcoma (RMS) is the most breakpoint region. and a separate green signal indicates common soft tissue sarcoma. RMS are one normal 13q14.11 locus and one classified in two main categories: embry- 13q14.11 locus affected by a transloca- onal rhabdomyosarcoma (ERMS) and FOXO1 tion. alveolar rhabdomyosarcoma (ARMS). The alveolar histology is associated with a poorer prognosis. ARMS is characterized

by two tumor-specific reciprocal translo- Ideogram of chromosome 13 cations t(2;13)(q36;q14.1) and t(1;13) indicating the hybridization locations.

(p36.1;q14.1) detectable in more than D13S1162 D13S320 80% of all ARMS. These translocations RH98592 D13S1014 3’ 5’ fuse the FOXO1 locus on 13q14.11 to either PAX3 on chromosome 2 or to FOXO1 PAX7 on chromosome 1. The resulting ~225 kb ~410 kb fusion transcripts encode for the chimeric Cen 13q14.11 Tel proteins PAX3-FOXO1 and PAX7-FOXO1 SPEC FOXO1 Probe map (not to scale). that combine transcriptional domains SPEC FOXO1 Break Apart Probe hybridized from the corresponding wild-type proteins to normal interphase cells as indicated by two red/green fusion signals per nucleus. and thereby acquire oncogenic activity. The translocations and their fusion genes represent highly specific genetic markers useful in the diagnosis of ARMS.

Prod. No. Product Label Tests* (Volume) C-3065-100 ZytoDot 2C SPEC FOXO1 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

229 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 229 DE036-1-20 ZytoDot ® 2C SPEC IGH Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC IGH Break Apart The ZytoDot ® 2C SPEC IGH Break Apart In an interphase nucleus of a normal Probe is designed to detect transloca- Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the tions involving the chromosomal region beled and a Dinitrophenyl-labeled probe 14q32.33 band, using the ZytoDot ® 2C 14q32.33 harboring the IGH gene. hybridizing to the 14q32.33 band. The CISH Implementation Kit, two red/green Rearrangements involving the IGH (im- DNP-labeled probe hybridizes proximal fusion signals are expected representing munoglobulin heavy locus, a.k.a. IGH@) and the DIG-labeled probe hybridizes two normal (non-rearranged) 14q32.33 gene are considered to be cytogenetic distal to the constant regions of the IGH loci. A signal pattern consisting of one hallmarks for non-Hodgkin lympho- locus at 14q32.33. red/green fusion signal, one red signal, ma (NHL). NHLs represent 50% of all and a separate green signal indicates hematological malignancies. IGH gene one normal 14q32.33 locus and one rearrangements have been identified in 14q32.33 locus affected by a transloca- about 50% of NHLs and are associated tion. with specific subtypes of NHLs. Translocation t(11;14)(q13.3;q32.3) can IGH be found in about in 95% of mantle cell Ideogram of chromosome 14 lymphoma (MCL), t(14;18)(q32.3;q21.3) indicating the hybridization locations. in 80% of follicular lymphoma (FL), t(3;14) RH48485 RH69050 (q27;q32.3) in diffuse large B-cell lympho- BV448316 RH66790 ma (DLBCL), and t(8;14)(q24.21;q32.3) 3’ 5’ 3’ 5’ in Burkitt lymphoma. In all of these trans- JAG2 IGH locations an oncogene located near the ~520 kb ~195 kb breakpoint of the translocation partner is Cen 14q32.33 Tel

activated by juxtaposing to IGH regulatory SPEC IGH Probe map (not to scale). sequences. Rearrangements involving 14q32.33 SPEC IGH Break Apart Probe hybridized have unique biological characteristics and to normal interphase cells as indicated correlate with clinical, morphological, by two red/green fusion signals per nucleus. and immunophenotypic features. CISH is a helpful tool for the diagnosis, selecting treatment, and giving prognostic information.

References Bernicot I, et al. (2007) Cytogenet Genome Res 118: 345-52. Hehne S, et al. (2012) Pathol Res Pract 208: 510-7. Kazuhiro N, et al. (1997) Blood 90: 526-34. Lu S, et al. (2004) Cancer Genet and Cytogenet 152: 141-5. Nishida K, et al. (1989) Cancer Res 49: 1275-81. Quintero-Rivera F, et al. (2009) Cancer Genet Cytogenet 190: 33-9.

Prod. No. Product Label Tests* (Volume) C-3071-100 ZytoDot 2C SPEC IGH Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

230 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 230 DE041-1-20 ZytoDot ® 2C SPEC FUS Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC FUS Break Apart The ZytoDot ® 2C SPEC FUS Break Apart In an interphase nucleus lacking a translo- Probe is designed to detect transloca- Probe is a mixture of a Digoxigenin-la- cation involving the 16p11.2 band, using tions involving the chromosomal region beled and a Dinitrophenyl-labeled probe the ZytoDot ® 2C CISH Implementation Kit 16p11.2 harboring the FUS (FUS RNA hybridizing to the 16p11.2 band. The two red/green fusion signals are expected binding protein, a.k.a. TLS, FUS/TLS, DNP-labeled probe hybridizes proximal representing two normal (non-rearranged) hnRNP P2) gene. to the FUS gene, the DIG-labeled probe 16p11.2 loci. A signal pattern consisting The FUS gene encodes an RNA-binding hybridizes distal to that gene. of one red/green fusion signal, one red protein, the C-terminal end of which is signal, and a separate green signal indi- involved in protein and RNA binding and cates one normal 16p11.2 locus and one which appears to be involved in transcrip- FUS 16p11.2 locus affected by a 16p11.2 tional activation with its N-terminal end. translocation. It shares distinct characteristics with EWS and TAF15 which together with FUS are Ideogram of chromosome 16 frequently referred to as the FET family of indicating the hybridization locations. proteins. FUS gene rearrangements have been RH101381 SHGC-106097 RH111999 CHLC.GCT13F06 shown to be involved in both solid tumors 5’ 3’ and leukemias fusing the N-terminal end of FUS FUS to various fusion partners. The most fre- ~175 kb ~715 kb quent translocation involving the FUS gene Tel 16p11.2 Cen

region is t(12;16)(q13.3;p11.2). Occurring SPEC FUS Probe map (not to scale). in over 90% of myxoid liposarcomas, the FUS-DDIT3 fusion protein is regarded as being consequential for the development Myxoid liposarcoma tissue section with translocation of myxoid liposarcomas by acting as an affecting the 16p11.2 locus as indicated abnormal transcription factor and thus by one non-rearranged red/green fusion signal, one red signal, and one separate green signal. deregulating FUS-DDIT3 target genes. Differential diagnosis of liposarcomas and accurate classification, the latter being especially important with regard to appropriate treatment and prognosis, are often problematic. Therefore, detection of FUS rearrangements via in situ Hybridiza- tion analysis is a valuable tool to confirm the histopathological diagnosis of myxoid liposarcoma.

References Andersson M, et al. (2010) BMC Cancer 10: 249-58. Antonescu C, et al. (2000) J Mol Diagn 2: 132-8. Germano G, et al. (2010) Cancer Res 70: 2235-44. Kuroda M, et al. (1995) Am J Pathol 147: 1221-7. Meis-Kindblom JM, et al. (2001) Virchows Arch 439: 141-51. Panagopoulos I, et al. (1994) Cancer Res 54: 6500-3. Panagopoulos I, et al. (1997) Oncogene 15: 1357-62.

Prod. No. Product Label Tests* (Volume) C-3054-100 ZytoDot 2C SPEC FUS Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

231 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 231 DE029-1-20 ZytoDot ® 2C SPEC USP6 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC USP6 Break Apart The ZytoDot ® 2C SPEC USP6 Break Apart In an interphase nucleus of a normal Probe is designed to detect transloca- Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the tions involving the chromosomal region beled and a Dinitrophenyl-labeled probe 17p13.2 band, using the ZytoDot ® 2C 17p13.2 harboring the USP6 (ubiquitin hybridizing to the 17p13.2 band. The CISH Implementation Kit, two red/green specific peptidase 6, a.k.a. Tre-2 or DNP-labeled probe hybridizes proximal to fusion signals are expected representing TRE17) gene. the USP6 breakpoint region at 17p13.2, two normal (non-rearranged) 17p13.2 Translocations affecting USP6 have been the DIG-labeled probe hybridizes distal to loci. A signal pattern consisting of one initially found in primary aneurysmal bone the USP6 breakpoint region. red/green fusion signal, one red signal, cysts (ABC), a benign, but locally aggres- and a separate green signal indicates one sive bone lesion that occurs predominantly USP6 normal 17p13.2 locus and one 17p13.2 during the first two decades of life. USP6 locus affected by a translocation. rearrangements are restricted to spindle cells in primary ABC, indistinguishable from surrounding normal spindle cells. Ideogram of chromosome 17 The resulting fusion genes detected are indicating the hybridization locations. formed by juxtaposition of the USP6 coding sequences to the highly active promot- RH41805 RH54973 PMC17834P1 RH36080 er sequences of several partner genes, as 5’ 3’ e.g. CDH11, COL1A1, OMD, TRAP150, USP6 and ZNF9, leading to the transcriptional ~190 kb ~275 kb upregulation of USP6. No true fusion Tel 17p13.2 Cen genes are formed. SPEC USP6 Probe map (not to scale). More recently, nodular fasciitis (NF), another mesenchymal lesion, has been tested positive for USP6 rearrangements. SPEC USP6 Break Apart Probe hybridized to normal interphase cells as indicated by NF is a subcutaneous pseudosarcomatous two red/green fusion signals per nucleus. myofibroblastic proliferation of unknown pathogenesis that regresses spontaneously when not surgically resected. The translocation results in the fusion of the promoter region of MYH9 located on 22q12.3 to the entire coding sequence of USP6 and subsequently in upregulated USP6 expression. For both lesions, it is assumed that the detection of USP6 rearrangements by CISH might represent a valuable diagnostic tool.

References Erickson-Johnson MR, et al. (2011) Lab Invest 91: 1427-33. Nakamura T, et al. (1988) Oncogene Res 2: 357-70. Oliveira AM, et al. (2004) Cancer Res 64: 1920-3. Oliveira AM, et al. (2005) Oncogene 24: 3419-26.

Prod. No. Product Label Tests* (Volume) C-3077-100 ZytoDot 2C SPEC USP6 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

232 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 232 DE045-1-20 ZytoDot ® SPEC ERBB2 Probe

Background Probe Description Results The ZytoDot ® SPEC ERBB2 Probe is The ZytoDot ® SPEC ERBB2 Probe is a In normal cells, two distinct dot-shaped designed for the detection of ERBB2 gene Digoxigenin-labeled probe specific for the signals per nucleus will be observed. Nu- amplification, frequently observed in solid ERBB2 gene at 17q12, processed by the clei with amplification of the ERBB2 gene malignant neoplasms, in formalin-fixed, unique ZytoVision ® Repeat Subtraction locus or polysomy of chromosome 17 will paraffin-embedded tissue sections or cell Technique resulting in advanced specifi- show multiple dots or large signal clusters. samples. city and less background. The ERBB2 gene (a.k.a. HER2 and NEU) is located in the chromosomal region 17q12 and encodes the cellular growth ERBB2 factor receptor p185. Amplification of the proto-oncogene ERBB2, observed in approximately 20% of Ideogram of chromosome 17 all breast cancer samples, has been cor- indicating the hybridization locations. related with a poor prognosis of the disease. Similar results have been obtained RH112805 RH104281 5’ 3’ for a variety of other malignant neoplasms e.g. ovarian cancer, stomach cancer, and ERBB2 carcinomas of the salivary gland. ~250 kb Cen 17q12 Tel References Normal nuclei each with two ERBB2 signals. Baselga J, et al. (1999) Semin Oncol 26: 78-83. Brockhoff G, et al. (2016) Histopathology 69: 635-46. SPEC ERBB2 Probe map (not to scale). Brunello E, et al. (2012) Histopathology 60: 482-8. Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Coussens L, et al. (1985) Science 230: 1132-9. Ettl T, et al. (2012) Br J Cancer 106: 719-26. Hwang CC, et al. (2011) Histopathology 59: 984-92. Hynes NE & Stern DF (1994) Biochim Biophys Acta 1198: 165-84. Moelans CB, et al. (2011) Crit Rev Oncol Hematol 80: 380-92. Park JB, et al. (1989) Cancer Res 49: 6605-9. Popescu NC, et al. (1989) Genomics 4: 362-6. Sassen A, et al. (2008) Breast Cancer Res 10: R2. Slamon DJ, et al. (1987) Science 235: 177-82. Voutsas IF, et al. (2013) Int J Radiat Biol 89: 319-25. Wolff AC, et al. (2018) J Clin Oncol 14: 437-41.

Breast carcinoma tissue section with ERBB2 amplification.

Prod. No. Product Label Tests* (Volume) C-3001-400 ZytoDot SPEC ERBB2 Probe DIG 40 (400 μl) C-3003-40 ZytoDot SPEC ERBB2 Probe Kit DIG 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Probe, 0.4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

233 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 233 DE001-1-20 ZytoDot ® 2C SPEC ERBB2/CEN 17 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC ERBB2/CEN 17 The ZytoDot ® 2C SPEC ERBB2/CEN 17 Using the ZytoDot ® 2C SPEC ERBB2/CEN Probe is designed for the simultaneous Probe is a mixture of a Digoxigenin-labe- 17 Probe Kit, two green (ERBB2) and two detection of ERBB2 and centromere 17 in led probe specific for the ERBB2 gene at red (CEN 17) signals are expected in a formalin-fixed, paraffin-embedded tissue 17q12 and a Dinitrophenyl-labeled CEN normal interphase nucleus. In a cell with sections or cell samples. 17 probe specific for the alpha satellite amplification of the ERBB2 gene locus, The ERBB2 gene (a.k.a. HER2 and NEU) centromeric region of chromosome 17 multiple copies of the green signal or is located in the chromosomal region (D17Z1). green signal clusters will be observed. 17q12 and encodes the cellular growth factor receptor p185. Amplification of the proto-oncogene ERBB2, observed in CEN 17 (D17Z1) ERBB2 approximately 20% of all breast cancer samples, has been correlated with a poor prognosis of the disease. Similar results Ideogram of chromosome 17 have been obtained for a variety of other indicating the hybridization locations. malignant neoplasms e.g. ovarian cancer, RH112805 RH69066 stomach cancer, and carcinomas of the 5’ 3’ salivary gland. ERBB2 References Baselga J, et al. (1999) Semin Oncol 26: 78-83. ~155 kb Brockhoff G, et al. (2016) Histopathology 69: 635-46. Brunello E, et al. (2012) Histopathology 60: 482-8. Cen 17q12 Tel Brunner K, et al. (2010) Anal Quant Cytol Histol 32: 78-89. Coussens L, et al. (1985) Science 230: 1132-9. SPEC ERBB2 Probe map (not to scale). Ettl T, et al. (2012) Br J Cancer 106: 719-26. Breast cancer tissue section with ERBB2 amplification Hwang CC, et al. (2011) Histopathology 59: 984-92. as indicated by multiple green signals in each nucleus. Hynes NE & Stern DF (1994) Biochim Biophys Acta 1198: 165-84. Moelans CB, et al. (2011) Crit Rev Oncol Hematol 80: 380-92. Park JB, et al. (1989) Cancer Res 49: 6605-9. Popescu NC, et al. (1989) Genomics 4: 362-6. Sassen A, et al. (2008) Breast Cancer Res 10: R2. Slamon DJ, et al. (1987) Science 235: 177-82. Voutsas IF, et al. (2013) Int J Radiat Biol 89: 319-25. Wolff AC, et al. (2018) J Clin Oncol 14: 437-41.

Gastric carcinoma tissue section with strong ERBB2 amplification as indicated by large green clusters.

Prod. No. Product Label Tests* (Volume) C-3032-100 ZytoDot 2C SPEC ERBB2/CEN 17 Probe DIG/DNP 10 (100 μl) C-3032-400 ZytoDot 2C SPEC ERBB2/CEN 17 Probe DIG/DNP 40 (400 μl) C-3022-10 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit DIG/DNP 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Probe, 0.1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3022-40 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit DIG/DNP 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Probe, 0.4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

234 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 234 DE009-1-20 ZytoDot ® 2C SPEC ERBB2/D17S122 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC ERBB2/D17S122 The ZytoDot ® 2C SPEC ERBB2/D17S122 In a normal interphase nucleus, using Probe is designed for the detection of Probe is a mixture of a Digoxigenin-labe- the ZytoDot ® 2C CISH Implementation ERBB2 gene amplification frequently ob- led probe specific for the chromosomal Kit, two green (ERBB2) and two red served in solid malignant neoplasms e.g. region 17q12 harboring the ERBB2 (D17S122) signals are expected. In a cell breast cancer samples. gene and a Dinitrophenyl-labeled SPEC with amplification of the ERBB2 gene locus The ERBB2 gene (a.k.a. HER2 and NEU) D17S122 probe specific for the chromo- or polysomy of chromosome 17, multiple is located in the chromosomal region somal region 17p12. The SPEC D17S122 copies of the green signal or green signal 17q12 and encodes a 185-190 kDa probe is designed to be used for chromo- clusters will be observed. transmembrane glycoprotein, p185, some 17 copy number detection. acting as a cellular growth factor receptor. The p185 protein belongs to the EGFR D17S122 (epidermal growth factor receptor) sub- ERBB2 group of the RTK (receptor tyrosine kinase) superfamily also including ERBB1 (HER1), ERBB3 (HER3), and ERBB4 (HER4). Ideogram of chromosome 17 Amplification of the proto-oncogene indicating the hybridization locations. ERBB2, observed in approximately 20% RH112805 RH69066 of all breast cancer samples, has been 5’ 3’ correlated with a poor prognosis of the ERBB2 disease. ~155 kb Similar results have been obtained for Cen 17q12 Tel Breast carcinoma tissue section with a variety of other malignant neoplasms amplification of the ERBB2 gene as indicated SPEC ERBB2 Probe map (not to scale). by multiple green signals in relation to e.g. ovarian cancer, stomach cancer, and red (D17S122) signals in each nucleus. carcinomas of the salivary gland. D17S1296 D17S122 SHGC-142057 Chromogenic in situ Hybridization targeting the alpha satellite centromeric regions of chromosome 17 may be misleading in ~480 kb Tel Cen some cases due to possible gains or losses 17p12 of this region. For these cases, reflex SPEC D17S122 Probe map (not to scale). testing is recommended using the SPEC ERBB2/D17S122 Probe. References Baselga J, et al. (1999) Semin Oncol 26: 78-83. Coussens L, et al. (1985) Science 230: 1132-9. Hwang CC, et al. (2011) Histopathology 59: 984-92. Hynes NE & Stern DF (1994) Biochim Biophys Acta 1198: 165-84. Moelans CB, et al. (2011) Crit Rev Oncol Hematol 80: 380-92. Park JB, et al. (1989) Cancer Res 49: 6605-9. Popescu NC, et al. (1989) Genomics 4: 362-6. Slamon DJ, et al. (1987) Science 235: 177-82. Voutsas IF, et al. (2013) Int J Radiat Biol 89: 319-25. Wolff AC, et al. (2018) J Clin Oncol 14: 437-41.

Prod. No. Product Label Tests* (Volume) C-3068-100 ZytoDot 2C SPEC ERBB2/D17S122 Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

235 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 235 DE033-1-20 ZytoDot ® 2C SPEC TOP2A/CEN 17 Probe

Background Probe Description Results The ZytoDot ® 2C SPEC TOP2A/CEN The ZytoDot ® 2C SPEC TOP2A/CEN 17 In a normal interphase nucleus, using the 17 Probe is designed for the detection of Probe is a mixture of a Digoxigenin-labe- ZytoDot ® 2C CISH Implementation Kit two TOP2A deletions and gene amplifications led probe specific for the TOP2A gene green and two red signals are expected. in formalin-fixed, paraffin-embedded tissue at 17q21.2 and a Dinitrophenyl-labeled In a cell with amplification of the TOP2A sections or cell samples. CEN 17 probe specific for the alpha satel- gene locus, multiple copies of the green The TOP2A (DNA topoisomerase II alpha) lite centromeric region of chromosome 17 signal or green signal clusters will be gene is located in the chromosomal region (D17Z1). observed. 17q21.2 and encodes for a 170 kDa DNA topoisomerase which controls and alters the topologic state of DNA during TOP2A replication, transcription, and chromosome segregation. TOP2A gene copy number changes are Ideogram of chromosome 17 frequently observed in the majority of indicating the hybridization locations. ERBB2 amplified primary breast tumors as well as in other human malignancies HUMUT5218 RH25598 3’ 5’ without simultaneous ERBB2 amplification e.g. acute lymphoblastic leukemias, gas- TOP2A tric and bladder carcinomas. Recent data ~315 kb Cen 17q21.2 Tel suggests that amplification and deletion SPEC TOP2A Probe map (not to scale). Breast carcinoma tissue section with of the TOP2A gene locus may account TOP2A amplification as indicated by for relative chemosensitivity or resistance multiple green signals per nucleus. to TOP2A inhibitor therapy, respectively. Thus, determination of the TOP2A status may predict benefit from adjuvant anthracyclines in ERBB2 positive breast cancer.

References Arriola E, et al. (2007) Breast Cancer Res Treat 106: 181-9. Brunello E, et al. (2012) Histopathology 60: 482-8. Järvinen TA & Liu ET (2006) Curr Cancer Drug Targets 6: 579-602. Järvinen TA, et al. (2000) Am J Pathol 156: 839-47. Razis E, et al. (2011) Breast Cancer Res Treat 128: 447-56. Tanner M, et al. (2006) J Clin Oncol 24: 2428-36. Tewey KM, et al. (1984) Science 226: 466-8. Tsai-Pflugfelder M, et al. (1988) Proc Natl Acad Sci U S A 85: 7177-81. Wang JC (1996) Annu Rev Biochem 65: 635-92.

Prod. No. Product Label Tests* (Volume) C-3040-400 ZytoDot 2C SPEC TOP2A/CEN 17 Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

236 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 236 DE015-1-20 ZytoDot ® 2C SPEC SS18 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC SS18 Break Apart The ZytoDot ® 2C SPEC SS18 Break In an interphase nucleus lacking a translo- Probe is designed to detect transloca- Apart Probe is a mixture of a Digoxi- cation involving the 18q11.2 band, using tions involving the chromosomal region genin-labeled probe and a Dinitrophe- the ZytoDot ® 2C CISH Implementation Kit 18q11.2 harboring the SS18 (SS18, nyl-labeled probe hybridizing to the two red/green fusion signals are expected nBAF chromatin remodeling complex 18q11.2 band. The DNP-labeled probe representing two normal (non-rearranged) subunit, a.k.a. SYT) gene. hybridizes distal to the SS18 gene and the 18q11.2 loci. A signal pattern consisting Translocations involving the region DIG-labeled probe hybridizes proximal to of one red/green fusion signal, one red 18q11.2 are found in over 90% of that gene. signal, and a separate green signal indi- synovial sarcoma. Among soft tissue cates one normal 18q11.2 locus and one sarcomas, synovial sarcoma is one of 18q11.2 locus affected by an 18q11.2 the most common and classically occurs SS18 translocation. in the extremities of young adults with greater prevalence in males even though, the occurrence of synovial sarcoma has Ideogram of chromosome 18 indicating the hybridization locations. also been described in a wide variety of anatomical locations and in all ages. RH109465 RH112040 RH119469 D18S66 The most frequent translocation involv- 3’ 5’ ing the SS18 gene region is t(X;18) (p11.23;q11.2) juxtaposing the SS18 SS18 gene in 18q11.2 either next to the SSX1 ~150 kb ~365 kb Cen 18q11.2 Tel (synovial sarcoma, translocated to X chro- SPEC SS18 Probe map (not to scale). mosome) or the SSX2 gene, or very rarely to the SSX4 locus located in Xp11.23. Complex translocations involving other SPEC SS18 Break Apart Probe hybridized chromosomes are observed in less than to normal interphase cells as indicated by 10% of synovial sarcomas. In combination two red/green fusion signals per nucleus. with histopathological diagnosis, detection of SS18 rearrangements via in situ Hybrid- ization (ISH) analysis is a valuable tool to confirm the diagnosis of synovial sarcoma.

References Amary MF, et al. (2007) Mod Pathol 20: 482-96. Clark J, et al. (1994) Nat Genet 7: 502-8. Kawai A, et al. (1998) N Engl J Med 338: 153-60. Surace C, et al. (2004) Lab Invest 84: 1185-92. Torres L, et al. (2008) Cancer Genet Cytogenet 187: 45-9.

Synovial sarcoma tissue section with translocation affecting the 18q11.2 locus as indicated by one non-rearranged red/green fusion signal, one red signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) C-3046-100 ZytoDot 2C SPEC SS18 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

237 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 237 DE020-1-20 ZytoDot ® 2C SPEC BCL2 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC BCL2 Break Apart The ZytoDot ® 2C SPEC BCL2 Break Apart In an interphase nucleus of a normal Probe is designed to detect transloca- Probe is a mixture of a Digoxigenin-la- cell lacking a translocation involving the tions involving the chromosomal region beled and a Dinitrophenyl-labeled probe 18q21.33 band, using the ZytoDot ® 2C 18q21.33 harboring the BCL2 gene. The hybridizing to the 18q21.33 band. The CISH Implementation Kit, two red/green BCL2 (BCL2 apoptosis regulator, a.k.a. DNP-labeled probe hybridizes distal to the fusion signals are expected representing PPP1R50) gene encodes a mitochondrial BCL2 gene at 18q21.33, the DIG-labeled two normal (non-rearranged) 18q21.33 membrane protein that regulates apoptosis probe hybridizes proximal to the BLC2 loci. A signal pattern consisting of one and is expressed in B-cells. gene at 18q21.33. red/green fusion signal, one red signal, Translocations involving the BCL2 gene and a separate green signal indicates are commonly identified in B-cell lym- one normal 18q21.33 locus and one phomas. In particular, the translocation 18q21.33 locus affected by a transloca- t(14;18)(q32.3;q21.3) has been identified tion. BCL2 in about 80% of follicular lymphoma (FL), in 20% to 30% of diffuse large B-cell Ideogram of chromosome 18 lymphoma (DLBCL), and rarely in B-cell indicating the hybridization locations. chronic lymphocytic leukemia (B-CLL). D18S91 GDB:435245 In FL this translocation is considered to be BV677679 REN11539 a cytogenetic hallmark. As a result of this 3’ 5’

rearrangement, the BCL2 gene is juxta- BCL2

posed to the IGH (Immunglobulin heavy ~170 kb ~375 kb chain) locus at 14q32.33 which leads Cen 18q21.33 Tel to overexpression of the anti-apoptotic SPEC BCL2 Probe map (not to scale). protein BCL2, and finally to progression to lymphoma. Alternative BCL2 translocations to immu- SPEC BCL2 Break Apart Probe hybridized noglobulin light chain genes as well as to normal interphase cells as indicated by non-IG translocation events have been two red/green fusion signals per nucleus. reported. In DLBCL, BCL2 gene overexpression has been implicated in conferring resistance to chemotherapy and has been associated with poor prognosis. Hence, detection of BCL2 translocations by CISH may be of diagnostic and prognostic relevance. References Da Cunha Santos G, et al. (2011) Cancer Cytopathol 119: 254-62. Dyer MJ, et al. (1994) Blood 83: 3682-8. Gu K, et al. (2008) Arch Pathol Lab Med 132: 1355-61. Hockenbery D, et al. (1990) Nature 348: 334-6. Impera L, et al. (2008) Oncogene 27: 6187-90. López-Guillermo A, et al. (1999) Blood 93: 3081-7. Nelson BP, et al. (2007) Am J Clin Pathol 128: 323-32. Tibiletti MG, et al. (2009) Hum Pathol 40: 645-52. Tomita N, et al. (2009) Haematologica 94: 935-43. Weinberg OK, et al. (2007) J Mol Diagn 9: 530-7.

Prod. No. Product Label Tests* (Volume) C-3073-100 ZytoDot 2C SPEC BCL2 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

238 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 238 DE039-1-20 ZytoDot ® 2C SPEC MALT1 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC MALT1 Break The ZytoDot ® 2C SPEC MALT1 Break In an interphase nucleus of a normal Apart Probe is designed to detect translo- Apart Probe is a mixture of a Di- cell lacking a translocation involving cations involving the chromosomal region goxigenin-labeled and a Dinitrophe- the 18q21.31-q21.32 band, using 18q21.32 harboring the MALT1 gene. nyl-labeled probe hybridizing to the the ZytoDot ® 2C CISH Implementa- The MALT1 (MALT1 paracaspase, a.k.a. 18q21.31-q21.32 band. The DNP-labe- tion Kit, two red/green fusion signals MLT) gene encodes a human paracaspase led probe hybridizes distal to the MALT1 are expected representing two normal and is often rearranged in MALT lympho- gene at 18q21.32, the DIG-labeled probe (non-rearranged) 18q21.31-q21.32 loci. mas accounting for 5-10% of all B-cell hybridizes proximal to the MALT1 gene at A signal pattern consisting of one red/ non-Hodgkin lymphomas (NHL). The 18q21.31-q21.32. green fusion signal, one red signal, and most common translocations affecting the a separate green signal indicates one MALT1 gene are t(11;18)(q22.2;q21.3) normal 18q21.31-q21.32 locus and one and t(14;18)(q32.3;q21.3) occurring in 18q21.31-q21.32 locus affected by a 50% and 15-20% of MALT lymphomas, MALT1 translocation. respectively. These translocations lead to the expression Ideogram of chromosome 18 indicating the hybridization locations. of BIRC3-MALT1 (a.k.a. API2-MALT1) and

IGH-MALT1 fusion proteins, resulting in SHGC-145775 D18S1129 KAM39 D18S529 constitutive activation of the NF-κB signal- 5’ 3’ ing pathway which controls the expression of numerous anti-apoptotic and prolifera- MALT1 tion-promoting genes. ~180 kb ~165 kb Cen 18q21.31-q21.32 Tel The translocation t(11;18)(q22.2;q21.3) is mainly found in pulmonary and SPEC MALT1 Probe map (not to scale). gastric lymphomas, whereas t(14;18) (q32.3;q21.3) occurs more frequently in non-gastrointestinal MALT lymphomas, e.g., of the skin and salivary glands. SPEC MALT1 Break Apart Probe hybridized to normal interphase cells as indicated by The presence of a t(11;18)(q22.2;q21.3) two red/green fusion signals per nucleus. correlates with unresponsiveness to erad- ication of Helicobacter pylori in gastric MALT lymphomas. Hence, detection of MALT1 translocations by CISH may be a supportive tool to identify patients eligible for an anti-H. pylori therapy.

References Afonina IS, et al. (2015) FEBS J 282: 3286-97. Beans M, et al. (2014) PLoS One 9: e103774. Dierlamm J, et al. (1999) Blood 93: 3601-9. Levine EG, et al. (1989) Blood 74: 1796-800. Lucas PC, et al. (2001) J Biol Chem 276: 19012-9. Martinelli G, et al. (2005) J Clin Oncol 23: 1979-83. Pereira MI & Medeiros JA (2014) World J Gastroenterol 20: 684-98. Troppan K, et al. (2015) Gastroenterol Res Pract 2015: 102656.

Prod. No. Product Label Tests* (Volume) C-3072-100 ZytoDot 2C SPEC MALT1 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

239 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 239 DE038-1-20 ZytoDot ® 2C SPEC ERG Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC ERG Break Apart The ZytoDot ® 2C SPEC ERG Break Apart In an interphase nucleus of a normal Probe is designed to detect aberrations Probe is a mixture of a Digoxigenin-labe- cell lacking an aberration involving the involving the ERG gene at 21q22.2 fre- led and a Dinitrophenyl-labeled probe hy- 21q22.13-q22.2 band, using the ZytoDot ® quently found in prostate cancers. bridizing to the long arm of chromosome 2C CISH Implementation Kit, two red/ ERG (ETS transcription factor ERG) 21. The DNP-labeled probe hybridizes green fusion signals are expected repre- rearrangements have been observed in proximal to the ERG gene breakpoint resenting the two normal (non-rearranged) 40-60% of prostate cancers identified via gion at 21q22.13-q22.2, the DIG-labeled 21q22.13-q22.2 loci. prostate-specific antigen (PSA) screening. probe hybridizes distal to the ERG gene A 21q22.13-q22.2 locus affected The most common aberration affecting breakpoint region at 21q22.2. by a 21q22.2 deletion resulting in the ERG is the interstitial deletion of about TMPRSS2-ERG fusion is indicated by the 3 Mb at the chromosomal region 21q22 loss of one green signal. found in 90% of the cases. This deletion ERG A signal pattern consisting of one red/ leads to the fusion of the hormonally green fusion signal, a separate green, Ideogram of chromosome 21 regulated promoter of the TMPRSS2 indicating the hybridization locations. and a separate red signal indicates an (transmembrane serine protease 2) gene ERG translocation without involvement of to the coding region of ERG, resulting in RH118947 RH119925 TMPRSS2 (e.g. SLC45A3-ERG). RH119817 403C6T7 overexpression of the ERG transcription 3’ 5’ factor. However, about 10% of the ERG ERG rearranged prostate cancer cases show ~560 kb ~190 kb alternative fusions, as e.g. SLC45A3-ERG Cen 21q22.13-q22.2 Tel or NDRG1-ERG. SPEC ERG Probe map (not to scale). Several studies detected associations of ERG rearrangements with histomorphologic features as well as characteristic chromosomal copy number changes and gene expression signatures, defining a distinct sub-class of prostate cancers with unfavorable prognosis. Hence, the evaluation of the ERG rearrangement status in Prostate cancer tissue section with translocation tissue or urine samples by CISH might be affecting the 21q22.13-q22.2 locus as indicated of diagnostic and prognostic relevance. by one non-rearranged red/green fusion signal, one red signal, and one separate green signal. EWSR1-ERG gene fusions present in about 10% of patients with Ewing sarcoma may result from complex genomic rearrangements and may therefore not be detected by CISH analysis or may result in a non-classical translocation signal pattern.

References Esgueva R, et al. (2010) Mod Pathol 23: 539-46. Maire G, et al. (2008) Cancer Genet Cytogenet 181: 81-92. Nam RK, et al. (2007) Br J Cancer 97: 1690-5. Perner S, et al. (2006) Cancer Res 66: 8337-41. Pflueger D, et al. (2009) Neoplasia 11: 804-11. Tomlins SA, et al. (2005) Science 310: 644-8.

Prod. No. Product Label Tests* (Volume) C-3058-400 ZytoDot 2C SPEC ERG Break Apart Probe DIG/DNP 40 (400 μl) Related Products C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

240 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 240 DE030-1-20 ZytoDot ® 2C SPEC EWSR1 Break Apart Probe

Background Probe Description Results The ZytoDot ® 2C SPEC EWSR1 Break The ZytoDot ® 2C SPEC EWSR1 Break In an interphase nucleus lacking a trans- Apart Probe is designed to detect translo- Apart Probe is a mixture of a Digoxi- location involving the 22q12.1-q12.2 cations involving the chromosomal region genin-labeled probe and a Dinitrophe- band, using the ZytoDot ® 2C CISH 22q12.2 harboring the EWSR1 (EWS nyl-labeled probe hybridizing to the Implementation Kit two red/green fusion RNA binding protein 1, a.k.a. EWS) 22q12.1-q12.2 band. The DNP-labeled signals are expected representing two gene). probe hybridizes proximal and extends normal (non-rearranged) 22q12.1-q12.2 Translocations involving the chromosomal inward intron 4 of the EWSR1 gene, the loci. A signal pattern consisting of one region 22q12.2 are found in 90-95% of DIG-labeled probe hybridizes distal to that red/green fusion signal, one red signal, patients with Ewing sarcoma or peripheral gene. and a separate green signal indicates primitive neuroectodermal tumors (PNET). one normal 22q12.1-q12.2 locus and Ewing sarcoma is the second most com- one 22q12.1-q12.2 locus affected by a mon, highly malignant bone tumor in chil- 22q12.1-q12.2 translocation. dren and young adults. The most frequent translocation involving the EWSR1 gene Ideogram of chromosome 22 region is t(11;22)(q24.3;q12.2) juxtapos- indicating the hybridization locations.

ing the EWSR1 gene in 22q12.2 next to RH76665 RH122132 the FLI-1 (friend leukemia virus integration D22S56 GDB:342145 5’ 3’ 1) locus in 11q24.3. FLI-1 is a member of the ETS family of transcription factors. Less EWSR1

frequently, EWSR1 can also be fused to ~260 kb ~280 kb ERG, a transcription factor closely related Cen 22q12.1-q12.2 Tel to FLI-1 but located in 21q22.2. SPEC EWSR1 Probe map (not to scale). For prognosis and appropriate treatment it is important to differentiate Ewing sarcoma/PNET from classic neuroblastoma, SPEC EWSR1 Break Apart Probe hybridized Wilms tumor, and rhabdomyosarcoma. to normal interphase cells as indicated by In combination with the histopathological two red/green fusion signals per nucleus. diagnosis, detection of EWSR1 rearrangements by using in situ Hybridization can be used to confirm the diagnosis of Ewing sarcoma/PNET. References Bridge RS, et al. (2006) Mod Pathol 19: 1-8. Delattre O, et al. (1992) Nature 359: 162-5. Lee J, et al. (2005) Cancer Genet Cytogenet 159: 177-80. Romeo S & Dei Tos AP (2010) Virchows Arch 456: 219-34. Sandberg AA & Bridge JA (2000) Cancer Genet Cytogenet 123: 1-26. Zucman J, et al. (1993) EMBO J 12: 4481-7.

Ewing sarcoma tissue section with translocation affecting the 22q12.1-q12.2 locus as indicated by one non-rearranged red/green fusion signal, one red signal, and one separate green signal.

Prod. No. Product Label Tests* (Volume) C-3043-100 ZytoDot 2C SPEC EWSR1 Break Apart Probe DIG/DNP 10 (100 μl) Related Products C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

241 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 241 DE019-1-20 ZytoDot ® Probes for Chromosome Enumeration

Background ZytoDot ® SPEC Probe Maps The ZytoDot ® Chromosome Enumeration SHGC-141884 RH105871 RH98442 D13S1546 Probes are designed for identification and 3’ 5’ 5’ 3’ enumeration of human chromosomes in interphase cells and as an adjunct to stand- WARS2 HAO2 ard karyotyping in metaphases. These ~285 kb ~165 kb Tel 1p12 Cen Cen 13q12.11 Tel probes will produce sharp, bright signals specific for each individual chromosome. SPEC 1p12 Probe map (not to scale). SPEC 13q12 Probe map (not to scale). The ZytoDot ® SPEC 1p12 Probe is The ZytoDot ® SPEC 13q12 Probe is CEN Probe Description designed to hybridize in close proxim- designed to hybridize in close proximity For most chromosomes, direct labeled ity of centromere 1 at 1p12 harboring of centromere 13 at 13q12.11. Since ZytoDot ® CEN™ Probes hybridizing to the WARS2 gene. Since chromosomes chromosomes 13 and 21 share the same highly repetitive human satellite DNA se- 1, 5, and 19 share the same repetitive repetitive sequences, they cannot be differ- quences mainly located at the centromeric sequences, they cannot be differentiated entiated by probes detecting centromere regions of chromosomes are applicable. by probes detecting centromere specific specific repeats. repeats. SPEC Probe Description RH69575 RH121214 SHGC-170072 D21S338 As several chromosomes share the same 3’ 5’ repetitive sequences resulting in cross-hybridization signals, they cannot be differ- AFF3 entiated by centromere specific probes. ~275 kb ~200 kb Cen 2q11.2 Tel Cen 21q22.13-q22.2 Tel Instead these chromosomes can be identified by direct labeled ZytoDot ® SPEC™ SPEC 2q11 Probe map (not to scale). SPEC 21q22 Probe map (not to scale). Probes hybridizing in close proximity to The ZytoDot ® SPEC 2q11 Probe is The ZytoDot ® SPEC 21q22 Probe hybrid- the respective satellite DNA sequences or specific for the AFF3 (AF4/FMR2 family, izes to the so-called Down Syndrome Criti- to other chromosome specific loci. member 3) gene region in 2q11.2. Due to cal Region on 21q22.13-q22.2 commonly cross-hybridizations of chromosome 2 al- duplicated in cases with partial trisomy Results pha satellites to other centromeric regions, 21. Since chromosomes 13 and 21 share In a normal interphase nucleus, two sig- probes specific for 2q11 are frequently the same repetitive sequences, they cannot nals are expected using Chromosome Enu- used for chromosome 2 copy number be differentiated by probes detecting meration Probes specific for autosomes. detection. centromere specific repeats. Using chromosome Y specific probes will result in normal male cells in one signal and in normal female cells in no signal. Using chromosome X specific probes will result in normal male cells in one signal and in normal female cells in two signals per nucleus. Other signal patterns indicate numerical aberrations of the respective chromosome.

242 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 242 DE012-1-20 Normal nuclei each with two CEN 12 signals. CEN X/Y Probe hybridized on normal male CEN X/Y Probe hybridized on normal female interphase cells as indicated by one red (chromosome interphase cells as indicated by two red X) and one green (chromosome Y) signal per nucleus. (chromosome X) signals per nucleus.

Prod. No. Product Alpha/Class. Sat. Chr. Band Label Tests* (Volume) C-3035-400 ZytoDot SPEC 1p12 Probe - 1p12 DIG 40 (400 μl) C-3051-400 ZytoDot SPEC 2q11 Probe - 2q11.2 DIG 40 (400 μl) C-3045-400 ZytoDot CEN 3 Probe D3Z1 3p11.1-q11.1 DIG 40 (400 μl) C-3002-400 ZytoDot CEN 6 Probe D6Z1 6p11.1-q11 DIG 40 (400 μl) C-3008-400 ZytoDot CEN 7 Probe D7Z1 7p11.1-q11.1 DIG 40 (400 μl) C-3016-400 ZytoDot CEN 8 Probe D8Z2 8p11.1-q11.1 DIG 40 (400 μl) C-3014-400 ZytoDot CEN 12 Probe D12Z3 12p11.1-q11 DIG 40 (400 μl) C-3052-400 ZytoDot SPEC 13q12 Probe - 13q12.11 DIG 40 (400 μl) C-3006-400 ZytoDot CEN 17 Probe D17Z1 17p11.1-q11.1 DIG 40 (400 μl) C-3026-400 ZytoDot SPEC 21q22 Probe - 21q22.13-q22.2 DIG 40 (400 μl) C-3025-400 ZytoDot CEN X Probe DXZ1 Xp11.1-q11.1 DIG 40 (400 μl) C-3020-400 ZytoDot CEN Yq12 Probe III DYZ1 Yq12 DIG 40 (400 μl) C-3048-400 ZytoDot 2C CEN X/Y Probe DXZ1/DYZ3 Xp11.1-q11.1/Yp11.1-q11.1 DNP/DIG 40 (400 μl) Related Products C-3018-40 ZytoDot CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

243 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 243 DE012-1-20 Accessories

ZytoDot ® Kits For the detection of Digoxigenin-labeled ZytoDot ® Probes Prod. No. Product Tests C-3005-40 ZytoDot CISH Polymer Detection Kit 40 Incl. Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml C-3018-40 ZytoDot CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; PBS/Tween, good for 2000 ml; Blocking Solution, 4 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

ZytoDot ® 2C Kits For the detection of Digoxigenin/Dinitrophenyl-labeled ZytoDot ® 2C Probes Prod. No. Product Tests C-3028-40 ZytoDot 2C CISH Polymer Detection Kit 40 Incl. 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml C-3044-10 ZytoDot 2C CISH Implementation Kit 10 Incl. Heat Pretreatment Solution EDTA, 150 ml; Pepsin Solution, 1 ml; Wash Buffer SSC, 210 ml; 20x Wash Buffer TBS, 50 ml; Anti-DIG/DNP-Mix, 1 ml; HRP/AP-Polymer-Mix,1 ml; AP-Red Solution A, 0.1 ml; AP-Red Solution B, 4 ml; HRP-Green Solution A, 0.2 ml; HRP-Green Solution B, 4 ml; Nuclear Blue Solution, 4 ml; Mounting Solution (alcoholic), 1 ml C-3044-40 ZytoDot 2C CISH Implementation Kit 40 Incl. Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; Wash Buffer SSC, 560 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-DIG/DNP-Mix, 4 ml; HRP/AP-Polymer-Mix, 4 ml; AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

ZytoDot ® Pretreatment Reagents Prod. No. Product C-3004-40 ZytoDot Pretreatment Kit Incl. Pepsin Solution, 4 ml; Heat Pretreatment Solution EDTA, 500 ml

ES-0001-4 Pepsin Solution, 4 ml

ES-0001-8 Pepsin Solution Set, 2x 4 ml

ES-0001-50 Pepsin Solution, 50 ml

ES-0001-1000 Pepsin Solution, 1000 ml

PT-0002-500 Heat Pretreatment Solution EDTA, 500 ml

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

244 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 244 ADE001-1-20 Accessories

ZytoDot ® Wash Buffers & Ancillary Reagents Prod. No. Product

AB-0001-4 Mouse-anti-DIG, 4 ml

AB-0001-30 Mouse-anti-DIG, 30 ml

AB-0002-4 Anti-Mouse-HRP-Polymer, 4 ml

AB-0013-4 HRP/AP-Polymer-Mix, 4 ml

AB-0014-4 Anti-DIG/DNP-Mix, 4 ml

BS-0001-4 Blocking Solution, 4 ml C-3011-40 ZytoDot Wash Buffer Set Incl. Wash Buffer SSC, 500 ml; PBS/Tween, good for 2000 ml C-3015-100 DAB Solution Set Incl. DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; good for 10 ml DAB Solution C-3038-100 ZytoDot AP-Red Solution Set Incl. AP-Red Solution A, 0.4 ml; AP-Red Solution B, 15 ml; good for 15 ml AP-Red Solution C-3039-100 ZytoDot HRP-Green Solution Set Incl. HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; good for 15 ml HRP-Green Solution

CS-0001-20 Mayer‘s Hematoxylin Solution, 20 ml

CS-0002-20 Nuclear Blue Solution, 20 ml

E-4005-50 Fixogum Rubber Cement, 50 g

E-4005-125 Fixogum Rubber Cement, 125 g

E-4007-2 ERBB2 Control Slide Set, 2 pcs.

MT-0004-4 Mounting Solution (alcoholic), 4 ml

WB-0001-560 Wash Buffer SSC, 560 ml

WB-0004-1000 PBS/Tween, good for 1000 ml

WB-0005-50 20x Wash Buffer TBS, 50 ml

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

245 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 245 ADE001-1-20 Page

Method Introduction - ZytoFast ® 247 - ZytoFast ® PLUS 248

Probes, sorted by Virus Species 249 sorted by mRNAs 249 sorted by Indication 250

Product Data Sheets 251 ff.

Accessories 256 f.

246 246 Achieving Chromogenic in situ Hybridization Results in just 4 Hours!

Introduction ZytoFast ®: Outstandingly fast CISH High Sensitivity and Specificity The ZytoFast ® products are designed Optimized protocols and faster tissue All ZytoFast ® probes are tagged using for outstandingly fast detection and penetration due to short oligonucleotide the unique ZytoFast ® HighTag System determination of lymphocyte clonality by probes of the ZytoFast ® system, make the providing improved signal intensity! High detecting IGK and IGL light chain RNA by ZytoFast ® CISH procedure outstandingly specificity without risk of cross-hybridi- Chromogenic in situ Hybridization (CISH) fast. zations is obtained due to optimized oligo- in formalin-fixed, paraffin-embedded tissue Single color results can be achieved with- nucleotide probes. sections and cell samples. in just 5 hours, hands-on time is about 3 hours! Advantages of CISH • Simultaneous observation of tissue morphology and CISH signals • No risk of false positives due to mis- The ZytoFast ® system uses oligonucle- priming or contamination as with PCR otide probes tagged with Biotin and • Easy method comparable to IHC Digoxigenin 1 which are detected 3 • No costly equipment needed using HRP-conjugated antibodies and • Ability to test archival specimens AP-conjugated streptavidin targeting • High sensitivity and specificity 2 the tags 2. The enzymatic reaction of chromogenic substrates 3, e.g. BCIP/ NBT and AEC, leads to the formation 1 of strong color precipitates that can be visualized by light microscopy.

Protocol Overview

Pepsin Denature Apply Probe Wash Apply Antibody Wash Stain and cover Analyze slides digestion & Hybridize

247 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 247 MFE001-1-20 ZytoFast ® PLUS for Increased Sensitivity!

Introduction ZytoFast ® PLUS – Outstandingly fast and ZytoFast ® PLUS – Flexibility that meets The ZytoFast ® PLUS products are de- sensitive CISH your Needs signed for outstandingly fast and sensitive Depending on the time required for Several ZytoFast ® PLUS CISH Implementa- detection and discrimination of human dewaxing and pretreatment of tissue tion Kits using different enzyme/substrate pathogen viruses, e.g. HPV, EBV, CMV, sections, ZytoFast ® PLUS protocols can be combinations can be combined with any and the determination of lymphocyte clon- performed within approx. 4 hours! separately available Digoxigenin-labeled ality by detecting IGK and IGL light chain Thus, due to optimized protocols, the ZytoFast ® probe to meet your preferences RNA by Chromogenic in situ Hybridiza- ZytoFast ® PLUS method takes only slightly concerning the detection chemistry, tion (CISH) in formalin-fixed, paraffin-em- more time compared to ZytoFast ® proto- counterstaining, and embedding. Each bedded tissue sections and cell samples. cols while being much more sensitive! ZytoFast ® PLUS CISH Implementation Kit The signal intensity of ZytoFast ® probes includes a detailed protocol, all necessary is increased even more when using the reagents as well as positive and negative ZytoFast ® PLUS Implementation Kits. control probes for versatile use in DNA as well as RNA in situ hybridizations. 4

The ZytoFast ® PLUS system uses Digoxigenin-labeled probes 1 which are detected using primary antibodies 2. These antibodies are detected by polymerized en- 3 zyme-conjugated secondary antibodies 3. The enzymatic reaction of chromogenic substrates 4, e.g. NBT/BCIP or DAB, leads to the formation of strong color precipitates that can be visualized by light microscopy. 2

Protocol Overview

Pepsin Denature Apply 1st Anti- Apply 2nd Pretreatment Apply Probe Wash Wash Wash Stain and cover Analyze slides digestion & Hybridize body Antibody

248 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 248 MFE002-1-20 Virus Index

Virus Product Name Label Product No. Quantity Page Index

HPV ZytoFast HPV type 6/11 Probe DIG T-1055-400 400 µl 251 ZytoFast HPV type 16/18 Probe DIG T-1056-400 400 µl 251 ZytoFast HPV type 31/33 Probe DIG T-1057-400 400 µl 251 Zyto HPV High-Risk (HR) Types Probe Fast DIG T-1140-400 400 µl 251 (specific for HPV type 16/18/31/33/35/39/45/51/52/56/58/59/66/68/82) Zyto HPV Screening Probe Fast DIG T-1144-400 400 µl 251 (specific for HPV type 6/11/16/18/31/33/35/39/45/51/52/56/58/59/66/68/82)

EBV ZytoFast EBV Probe DIG T-1114-400 400 µl 252 (a.k.a. HHV-4)

CMV ZytoFast CMV Probe DIG T-1113-400 400 µl 253

mRNA Index

mRNA Product Name Label Product No. Quantity Page Index

Ig-kappa ZytoFast human lg-kappa Probe DIG T-1115-400 400 µl 254 f. ZytoFast human lg-kappa/lg-lambda Probe DIG/Biotin T-1017-400 400 µl 254 f. ZytoFast human lg-kappa/lg-lambda CISH Kit DIG/Biotin T-1005-40 40 tests 254 f. ZytoFast human lg-kappa/lg-lambda Permanent CISH Kit DIG/Biotin T-1105-40 40 tests 254 f.

Ig-lambda ZytoFast human lg-lambda Probe DIG T-1116-400 400 µl 254 f. ZytoFast human lg-kappa/lg-lambda Probe DIG/Biotin T-1017-400 400 µl 254 f. ZytoFast human lg-kappa/lg-lambda CISH Kit DIG/Biotin T-1005-40 40 tests 254 f. ZytoFast human lg-kappa/lg-lambda Permanent CISH Kit DIG/Biotin T-1105-40 40 tests 254 f.

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

249 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 249 IFE001-1-20 Indication Index Indication Product Name Label Product No. Quantity Page

Solid Tumors Cervical Cancer ZytoFast HPV type 6/11 Probe DIG T-1055-400 400 µl 251 ZytoFast HPV type 16/18 Probe DIG T-1056-400 400 µl 251 ZytoFast HPV type 31/33 Probe DIG T-1057-400 400 µl 251 Zyto HPV High-Risk (HR) Types Probe Fast DIG T-1140-400 400 µl 251 (specific for HPV type 16/18/31/33/35/39/45/51/52/56/58/59/66/68/82) Zyto HPV Screening Probe Fast DIG T-1144-400 400 µl 251 (specific for HPV type 6/11/16/18/31/33/35/39/45/51/52/56/58/59/66/68/82)

Hematology Specific Probes Lymphoma ZytoFast EBV Probe DIG T-1114-400 400 µl 252 ZytoFast human Ig-kappa Probe DIG T-1115-400 400 µl 254 f. ZytoFast human Ig-lambda Probe DIG T-1116-400 400 µl 254 f. ZytoFast human Ig-kappa/Ig-lambda Probe DIG/Biotin T-1017-400 400 µl 254 f. ZytoFast human Ig-kappa/Ig-lambda CISH Kit DIG/Biotin T-1005-40 40 tests 254 f. ZytoFast human lg-kappa/lg-lambda Permanent CISH Kit DIG/Biotin T-1105-40 40 tests 254 f.

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

250 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 250 IFS001-1-20 ZytoFast ® HPV-CISH System

Background Probe Description Results The ZytoFast ® HPV-CISH System is de- ZytoFast ® HPV specific probes are A positive reactivity for HPV DNA in epi- signed for the detection and discrimination directed against DNA sequences which thelial cells is indicated by a distinctly of human papillomavirus (HPV) DNA in encode the HPV proteins E6, E7 and/or stained nucleus. Due to the detection of paraffin-embedded tissue sections or cell L1. The probes consist of HPV-type-specific HPV DNA as well as E6, E7, and/or L1 samples. oligonucleotides, Digoxigenin-labeled by RNAs, depending on the infection stage, At least 50 percent of sexually active men using the unique ZytoFast ® HighTag cytoplasmic staining might be observed and women acquire some form of genital System providing improved signal intensi- additionally. Depending on the detection HPV infection at some point in their lives. ty. In addition to the detection of HPV at chemistry that is used, colored precipi- Most of the approx. 30 identified genital the DNA level, HPV probes will also allow tates, which can be clearly distinguished HPV types, predominantly types 6 and 11, detection of E6, E7, and/or L1 RNAs, from the background, will be dark vio- are called “low-risk” types, and may cause which are expressed during some stages let-blue when using NBT/BCIP as substrate, mild Pap test abnormalities or genital warts. of infection. dark brown when using DAB, or strong Until now, approximately 10–15 HPV red when using Permanent Red. types are associated with lesions that can progress to cancer. Among those are the HPV types 16/18/31/33/35/39/45/ 51/52/56/58/59/66/68/82. These cancer-associated HPV types are designated as high-risk HPV (hr-HPV) types. The infection with the HPV hr-types can lead to development of cancer of the cervix, vulva, vagina, anus, or penis. The majority of malignant cervical carcinomas (approx. 70%) occur as a result of infec- Schematic representation of the HPV genome with E and L open reading frames. Genomic tions with HPV types 16 or 18. regions targeted by ZytoFast ® HPV specific References oligonucleotides are indicated in blue. Cothran MM & White JP (2002) Health Care Women Int 23: 306-19. HPV infected cervix tissue hybridized Cubie HA & Norval M (1988) J Virol Methods 20: 239-49. with the ZytoFast ® HPV type 6/11 Probe, Faulkner-Jones BE, et al. (1993) J Virol Methods 41: 277-96. detected with the ZytoFast ® PLUS CISH Francis IM, et al. (2013) Sultan Qaboos Univ Med J 13: 527-33. Grundmeier N, et al. (2011) Dermatology 223: 293-300. Implementation Kit AP-Permanent Red. Kaspersen MD, et al. (2011) PLoS One 6: e18095. Mirasoli M, et al. (2009) Anal Bioanal Chem 394: 981-7. Montag M, et al. (2011) Arch Gynecol Obstet 284: 999-1005. Poljak M & Kocjan BJ (2010) Expert Rev Anti Infect Ther 8: 1139-62. Reinholz M, et al. (2013) Arch Dermatol Res 305: 723-32. Walboomers JMM, et al. (1999) J Pathol 189: 12-9.

Digoxigenin-labeled Prod. No. Product Tests* (Volume) T-1055-400 ZytoFast HPV type 6/11 Probe 40 (400 μl) T-1056-400 ZytoFast HPV type 16/18 Probe 40 (400 μl) T-1057-400 ZytoFast HPV type 31/33 Probe 40 (400 μl) T-1140-400 ZytoFast HPV High-Risk (HR) Types Probe (specific for HPV type 16/18/31/33/35/39/45/51/52/56/58/59/66/68/82) 40 (400 μl) T-1144-400 ZytoFast HPV Screening Probe (specific for HPV type 6/11/16/18/31/33/35/39/45/51/52/56/58/59/66/68/82) 40 (400 μl) Related Products T-1061-40 ZytoFast PLUS CISH Implementation Kit AP-NBT/BCIP 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; NBT/BCIP Solution, 4ml; Nuclear Red Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1063-40 ZytoFast PLUS CISH Implementation Kit HRP-DAB 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1151-40 ZytoFast PLUS CISH Implementation Kit AP-Permanent Red 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; Permanent Red Solution A, 0.25 ml; Permanent Red Solution B, 15 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

251 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 251 CE002-1-20 ZytoFast ® EBV-CISH System

Background Probe Description Results The ZytoFast ® EBV-CISH System is The ZytoFast ® EBV Probe is directed A positive reactivity for Epstein-Barr-virus designed for the detection of Epstein-Barr against EBER-1 and EBER-2 RNA (EBV) EBER RNA in the target cells is indi- virus (EBV) EBER RNA in paraffin-embed- sequences that were found to be tran- cated by a distinctly stained nucleus. De- ded tissue sections or cell samples. scribed in every latently infected cell. Due pending on the detection chemistry that is EBV (a.k.a. human herpesvirus-4, HHV-4) to the large number (up to 107) of copies used, colored precipitates, which can be is a member of the gamma-herpesvirus per cell, these RNAs are the most abun- clearly distinguished from the background, group and one of the most common virus- dant transcripts in latently EBV-infected will be dark violet-blue when using NBT/ es in humans. cells. BCIP as substrate, dark brown when using Transmission of EBV requires close, The probe consists of EBV-specific oligo- DAB, or strong red when using Permanent intimate contact with a person excreting nucleotides, Digoxigenin-labeled by using Red. the virus in its saliva. EBV has two major the unique ZytoFast ® High Tag System target tissues in vivo, B lymphocytes and providing improved signal intensity. squamous pharyngeal epithelium. Infec- tion of B lymphocytes with EBV results in persistant latent infection, immortalization of the cells, and perpetual proliferation. EBV, the first virus to be identified as an oncovirus, is the etiological agent of Schematic representation of the EBV genome with the EBER-1 and EBER-2 encoding region indicated in blue. U1-U5 indicate unique nucleotide sequences, hatched infectious mononucleosis and has been EBV infected tonsil tissue hybridized with boxes represent terminal and internal repeats. implicated in the pathogenesis of an ZytoFast ® EBV Probe, detected with ZytoFast ® increasing number of human malignancies PLUS CISH Implementation Kit HRP-DAB. such as Burkitt lymphoma, nasopharyn- geal carcinoma, and polyclonal lymphomas in immunocompromised individuals. CISH-based diagnosis of EBV infection has the advantage over other methods in that it permits unequivocal localization of EBV genomes in cells and thereby obviates the risk of false positive results due to laboratory or clinical contamination.

References Agaimy A & Wünsch PH (2006) Pathol Res Pract 202: 541-8. Fuchs S, et al. (2012) J Immunol 188: 1523-33. Gaiser T, et al. (2012) Diagn Pathol 7: 38. Greifenegger N, et al. (1998) J Virol 72: 9323-8. Khan G, et al. (1992) J Clin Pathol 45: 616-20. Kim DN, et al. (2013) J Gen Virol 94: 497-506. Murphy JK, et al. (1990) J Clin Pathol 43: 220-3. Rosa MD, et al. (1981) Mol Cell Biol 1: 785-96. Sides MD, et al. (2013) Virol J 10: 152. Thorley-Lawson DA (2001) Nat Rev Immunol 1: 75-82.

Digoxigenin-labeled Prod. No. Product Tests* (Volume) T-1114-400 ZytoFast EBV Probe 40 (400 μl) Related Products T-1061-40 ZytoFast PLUS CISH Implementation Kit AP-NBT/BCIP 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; NBT/BCIP Solution, 4ml; Nuclear Red Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1063-40 ZytoFast PLUS CISH Implementation Kit HRP-DAB 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1151-40 ZytoFast PLUS CISH Implementation Kit AP-Permanent Red 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; Permanent Red Solution A, 0.25 ml; Permanent Red Solution B, 15 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

252 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 252 CE001-1-20 ZytoFast ® CMV-CISH System

Background Probe Description Results The ZytoFast ® CMV-CISH System is de- The ZytoFast ® CMV Probe is directed Due to the detection of CMV DNA as well signed for the detection of cytomegalovi- against the sequence of the b2.7 gene, as of the abundantly transcribed β2.7 rus (CMV) DNA in paraffin-embedded tis- the most abundantly transcribed early RNA, a positive reactivity for cytomegalo- sue sections or cell samples. CMV (a.k.a. CMV gene. The probe consists of CMV- virus (CMV) in the target cells is indicated human herpesvirus-5, HHV-5) is a member specific oligonucleotides, Digoxigenin- by a cytoplasmic and/or nuclear staining of the beta-herpesvirus group and may be labeled by using the unique ZytoFast ® pattern. Depending on the detection chem- found in 40-100% of people. CMV can be HighTag System providing improved istry that is used, colored precipitates, transmitted sexually as well as via breast signal intensity. In addition to the detection which can be clearly distinguished from milk, transplanted organs, and rarely from of CMV at the DNA level, the CMV Probe the background, will be dark violet-blue blood transfusions. will also allow detection of the b2.7 RNA, when using NBT/BCIP as substrate, dark Following primary CMV infection in the which is expressed during all stages of brown when using DAB, or strong red normal host, the virus remains in a latent infection. when using Permanent Red. state and can be found in multiple body sites as it is, unlike other herpesviruses, not restricted to certain body areas. Among immunosuppressed patients, such as Schematic representation of the CMV genome with recipients of solid organ or haematopoi- the b2.7 encoding region indicated in blue. UL and US indicate unique nucleotide sequences, hatched etic stem cell allografts, CMV infections boxes represent terminal and internal repeats. are common causes of morbidity and mortality. In histology, the hallmark of CMV infection is the finding of intranuclear inclusions consistent with the virus. CISH-based

diagnosis of CMV infection has the advan- CISH analysis of paraffin-embedded adrenal tage over other methods in that it permits gland tissue using the ZytoFast ® CMV Probe, detected with ZytoFast ® PLUS CISH unequivocal localization of CMV genomes Implementation Kit AP-NBT/BCIP. in cells and thereby obviates the risk of false positive results due to laboratory or clinical contamination.

References Cobbs CS, et al. (2002) Cancer Res 62: 3347-50. Chen HP, et al. (2012) J Clin Virol 54: 240-4. Greenaway PJ & Wilkinson GWG (1987) Virus Res 7: 17-31. Spector SA (1990) Semin Hematol 27 (2 Suppl 1): 11-6; 28-9. Wu TC, et al. (1992) Am J Pathol 141: 1247-54.

Digoxigenin-labeled Prod. No. Product Tests* (Volume) T-1113-400 ZytoFast CMV Probe 40 (400 μl) Related Products T-1061-40 ZytoFast PLUS CISH Implementation Kit AP-NBT/BCIP 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; NBT/BCIP Solution, 4ml; Nuclear Red Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1063-40 ZytoFast PLUS CISH Implementation Kit HRP-DAB 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1151-40 ZytoFast PLUS CISH Implementation Kit AP-Permanent Red 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; Permanent Red Solution A, 0.25 ml; Permanent Red Solution B, 15 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

253 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 253 CE003-1-20 ZytoFast ® Ig-kappa/Ig-lambda-CISH System

Background Probe Description Results The ZytoFast ® Ig-kappa/Ig-lambda-CISH ZytoFast ® human Ig-kappa Probe is direct- A positive reactivity in the target cells is System is designed for the detection of im- ed against mRNA sequences encoding k indicated by cytoplasmic staining. De- munglobulin kappa locus (Ig-k, IGK) and/ light chain constant regions of human pending on the detection chemistry that is or immunglobulin lamda locus (Ig-l, IGL) immunoglobulins. used, colored precipitates, which can be light chain mRNA in paraffin-embedded The ZytoFast ® human Ig-lambda Probe clearly distinguished from the background, tissue sections or cell samples. is directed against mRNA sequences will be dark violet-blue when using NBT/ B-cells (a.k.a. B lymphocytes) develop from encoding l light chain constant regions of BCIP as substrate, strong red when using lymphoid stem cells in the bone marrow. human immunoglobulins. AEC, dark brown when using DAB, green Each clone of B-cells expresses a unique The ZytoFast ® human Ig-kappa/Ig-lambda when using HRP-Green, or strong red antibody molecule, composed of 2 identi- Probe is a probe mixture consisting of a when using Permanent Red. cal heavy and 2 identical light chains, the Digoxigenin-labeled IGK mRNA specific latter either of k or l type. Determination probe and a Biotin-labeled IGL mRNA Using the ZytoFast ® human Ig-kappa of kappa-to-lambda ratio is useful to dis- specific probe. All probes are tagged by Probe, B-cells expressing antibodies with tinguish between neoplastic and reactive using the unique ZytoFast ® HighTag Sys- κ light chains will result in cytoplasmic lymphoid proliferations. Polyclonal expres- tem providing improved signal intensity. staining whereas IGL expressing B-cells sion of k or l light chains is considered to are not stained. reflect a reactive hyperplasia in contrast to the monoclonal expression in malignant Using the ZytoFast ® human Ig-lambda lymphoma, the most common hematologic Probe, B-cells expressing antibodies with malignancy encountered in the Western λ light chains will result in cytoplasmic world. Whereas detection of IGK and staining whereas IGK expressing B-cells IGL by immunohistochemistry often results Basic immunoglobulin structure indicating are not stained. in excessive background staining, in situ the heavy chains (black), l (blue) and k (red) lights chains. The light chain constant regions (C) Hybridization has the advantage of a whose encoding mRNA sequences are targeted by Using the ZytoFast ® human Ig-kappa/ ZytoFast ® lg-lambda and lg-kappa probes are virtually background-free signal, allowing indicated in dark blue and red respectively, the Ig-lambda CISH Kit, B-cells expressing a safe and simple analysis of the clonality variable regions (V) in light blue and red. antibodies with κ light chains will result in of a given lymphocyte population. a red cytoplasmic staining and simultane-

References ously IGL expressing B-cells will result in a Erber WN, et al. (1993) Pathology 25: 63-7. Ke L, et al. (2011) Int J Clin Exp Pathol 4: 190-6. dark violet-blue cytoplasmic staining. McElroy MK, et al. (2011) Hum Pathol 42: 1813-8. McNicol AM & Farquharson MA (1997) J Pathol 182: 250-61. Pringle JH, et al. (1990) J Pathol 162: 197-207. Using the ZytoFast ® human Ig-kappa/ Ig-lambda Permanent CISH Kit, B-cells expressing antibodies with κ light chains will result in a green cytoplasmic staining and simultaneously IGL expressing B-cells CISH analysis of a paraffin-embedded will result in permanent red cytoplasmic tonsil tissue using the ZytoFast ® human Ig-kappa/Ig-lambda Permanent CISH Kit. staining.

CISH analysis of a paraffin-embedded Tonsil tissue with B-cells expressing Ig-kappa Tonsil tissue with B-cells expressing Ig-kappa bone marrow biopsy specimen using the hybridized with ZytoFast ® human Ig-kappa Probe, hybridized with ZytoFast ® human Ig-kappa ZytoFast ® human Ig-kappa/Ig-lambda CISH Kit. detected with ZytoFast ® PLUS CISH Probe, detected with ZytoFast ® PLUS CISH Implementation Kit AP-Permanent Red. Implementation Kit HRP-DAB.

254 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 254 CE004-1-20 ZytoFast ® Ig-kappa/Ig-lambda Probes Digoxigenin-labeled Prod. No. Product Tests* (Volume) T-1115-400 ZytoFast human lg-kappa Probe 40 (400 μl) T-1116-400 ZytoFast human lg-lambda Probe 40 (400 μl) Related Products T-1061-40 ZytoFast PLUS CISH Implementation Kit AP-NBT/BCIP 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; NBT/BCIP Solution, 4ml; Nuclear Red Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1063-40 ZytoFast PLUS CISH Implementation Kit HRP-DAB 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1151-40 ZytoFast PLUS CISH Implementation Kit AP-Permanent Red 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; Permanent Red Solution A, 0.25 ml; Permanent Red Solution B, 15 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

ZytoFast ® Ig-kappa/Ig-lambda Probe Biotin/Digoxigenin-labeled Prod. No. Product Tests* (Volume) T-1017-400 ZytoFast human lg-kappa/lg-lambda Probe 40 (400 μl) Related Products T-1005-40 ZytoFast human lg-kappa/lg-lambda CISH Kit 40 Incl. Ig-kappa/lg-lambda Probe (DIG/Biotin-labeled), 0.4 ml; 28S rRNA (+) Control Probe (DIG-labeled), 0.1 ml; RNA (-) Control Probe (DIG-labeled), 0.1 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-Biotin/DIG-Mix, 4 ml; AEC Solution, 4 ml; NBT/BCIP Solution, 4 ml; Nuclear Green Solution, 20 ml; Mounting Solution (aqueous), 4 ml T-1105-40 ZytoFast human Ig-kappa/Ig-lambda Permanent CISH Kit 40 Incl. Ig-kappa/lg-lambda Probe (DIG/Biotin-labeled), 0.4 ml; 28S rRNA (+) Control Probe (DIG-labeled), 0.1 ml; RNA (-) Control Probe (DIG-labeled), 0.1 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 2x 50 ml; Anti-Biotin/DIG-Mix, 4 ml; HRP-Green-Solution A, 0.8 ml; HRP-Green-Solution B, 15 ml; Permanent Red Solution A, 0.25 ml; Permanent Red Solution B, 15 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

255 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 255 CE004-1-20 Accessories

Peroxidase (HRP) Alkaline Phosphatase (AP)

DAB Permanent Red NBT/BCIP

T-1063 T-1151 T-1061

ZytoFast ® PLUS Implementation Kits For the detection of Digoxigenin-labeled ZytoFast ® Probes Prod. No. Product Tests T-1061-40 ZytoFast PLUS CISH Implementation Kit AP-NBT/BCIP 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; NBT/BCIP Solution, 4ml; Nuclear Red Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1063-40 ZytoFast PLUS CISH Implementation Kit HRP-DAB 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x 50 ml; Mouse-anti-DIG, 4 ml; Anti-Mouse-HRP-Polymer, 4 ml; DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; Nuclear Blue Solution, 20 ml; Mounting Solution (alcoholic), 4 ml T-1151-40 ZytoFast PLUS CISH Implementation Kit AP-Permanent Red 40 Incl. DNA (+) Control Probe, 0.1 ml; DNA (-) Control Probe, 0.1 ml; 28S rRNA (+) Control Probe, 0.1 ml; RNA (-) Control Probe, 0.1 ml; Heat Pretreatment Solution EDTA, 500 ml; Pepsin Solution, 4 ml; 20x Wash Buffer TBS, 4x50 ml; Rabbit-anti-DIG, 4 ml; Anti-Rabbit-AP-Polymer, 4 ml; Permanent Red Solution A, 0.25 ml; Permanent Red Solution B, 15 ml; Mayer’s Hematoxylin Solution, 20 ml; Mounting Solution (alcoholic), 4 ml

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

256 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 256 AFE001-1-20 Accessories

ZytoFast ® Pretreatment Reagents Prod. No. Product

ES-0001-4 Pepsin Solution, 4 ml

ES-0001-8 Pepsin Solution Set, 2x 4 ml

ES-0001-50 Pepsin Solution, 50 ml

ES-0001-1000 Pepsin Solution, 1000 ml

PT-0002-500 Heat Pretreatment Solution EDTA, 500 ml

ZytoFast ® Wash Buffers & Ancillary Reagents Prod. No. Product

AB-0001-4 Mouse-anti-DIG, 4 ml

AB-0001-30 Mouse-anti-DIG, 30 ml

AB-0002-4 Anti-Mouse-HRP-Polymer, 4 ml

AB-0011-4 Rabbit-anti-DIG, 4 ml

AB-0012-4 Anti-Rabbit-AP-Polymer, 4 ml

AB-0015-4 Anti-Biotin/DIG-Mix, 4 ml C-3015-100 DAB Solution Set Incl. DAB Solution A, 0.3 ml; DAB Solution B, 10 ml; good for 10 ml DAB Solution C-3039-100 ZytoDot HRP-Green Solution Set Incl. HRP-Green Solution A, 0.8 ml; HRP-Green Solution B, 15 ml; good for 15 ml HRP-Green Solution

CS-0001-20 Mayer‘s Hematoxylin Solution, 20 ml

CS-0002-20 Nuclear Blue Solution, 20 ml

CS-0003-20 Nuclear Red Solution, 20 ml

CS-0004-20 Nuclear Green Solution, 20 ml

E-4005-50 Fixogum Rubber Cement, 50 g

E-4005-125 Fixogum Rubber Cement, 125 g

MT-0004-4 Mounting Solution (alcoholic), 4 ml

SB-0004-4 NBT/BCIP Solution, 4 ml

SB-0005-4 AEC Solution, 4 ml

WB-0005-50 20x Wash Buffer TBS, 50 ml

ZytoFast ® Control Probes Digoxigenin-labeled

Prod. No. Product Tests* (Volume) T-1053-400 ZytoFast DNA (+) Control Probe 40 (400 μl) T-1054-400 ZytoFast DNA (–) Control Probe 40 (400 μl) T-1120-400 ZytoFast 28S rRNA (+) Control Probe 40 (400 μl) T-1119-400 ZytoFast RNA (–) Control Probe 40 (400 μl)

* Using 10 µl probe solution per test. only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

257 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 257 FE073-1-19 VisionArray Arrays for DNA analysis Page

Method Introduction - VisionArray ® 259

Product Data Sheets 260 ff.

Accessories 263 ff.

258 258 VisionArray Arrays for DNA analysis

VisionArray ® Chip - Fast and Reliable Detection of DNA Sequences!

Introduction Advantages of VisionArray ® Sample Collection The VisionArray ® products are designed • High sensitivity and specificity For the detection of DNA sequences with for the qualitative detection of specific • Quick & easy 1 hour protocol the VisionArray ® system, the following raw DNA sequences by DNA/DNA hybridi- • Automated evaluation using a material can be used for DNA extraction; zation on immobilized catcher molecules VisionArray ® Analyzer Software – depending on the VisionArray ® Chip used: which are arranged on a glass chip. simple visualization & quick analysis • Formalin-fixed, paraffin-embedded All capture sequences and positive in just a few minutes (FFPE) tissue or cell samples controls are set up on the VisionArray ® • Liquid based cytology specimens Chips as duplicates. (e.g. ThinPrep®, swab/brush specimen, sputum)

Step 1: Amplification and Labeling in a PCR The DNA is extracted from, e.g., FFPE samples and is used as a template for PCR. Biotinylated primers are used to amplify and label different sections of the target sequences. The human HLA-DQA1 gene is also amplified and serves as a PCR positive control and as a genomic control. Step 2: Hybridization on the Glass Chip

After amplification, the biotinylated sequences hybridize to complementary DNA capture sequences on the glass chip.

Step 3: Detection and Visualization Specifically bound and biotinylated sequences are visualized by secondary marking with a streptavidin-peroxidase conjugate and a staining with tetramethyl- benzidine. After color development, evaluation is performed using a VisionArray ® Analyzer Software.

259 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 259 MVAE001-1-20 VisionArray Arrays for DNA analysis

VisionArray ® HPV Chip 1.0

Introduction The VisionArray ® HPV Chip 1.0 is intended to be used for the qualitative detection and genotyping of PCR amplificates of 41 clinically relevant human papillomavirus (HPV) genotypes that have been produced with the help of the VisionArray ® HPV Primer Kit 2.0 and the VisionArray ® Detection Kit. HPV has been conclusively identified as the major risk factor for cervical cancer. It is the third most common cancer in women worldwide, with an estimated number of 530,000 new cases and 280,000 deaths each year. Over the last years, the relevance of HPV in the history of oropharyngeal cancers has become more and more important which is indicated by a dramatically risen number of cancers of the oral cavity and pharynx linked to HPV.

At present, there are more than 150 different HPV types described. Depending on their risk to induce cancer, they are divided into Low Risk (LR), Probably High Risk, and High Risk (HR) types.

Chip Description The VisionArray ® HPV Chip 1.0 is designed to detect 41 clinically relevant HPV genotypes. All capture sequences and the positive control are set up on the Chip as duplicates and the guide dots as triplicates. The signals are visible on the Chip as dark blue areas. The automated evaluation of the results is performed by a VisionArray ® Analyzer Software.

High Risk HPV Types detected by the VisionArray ® HPV Chip GD + 6 11 16 18 26 GD 16 18 31 33 35 39 31 33 34 35 39 40 45 51 52 56 58 59 42 43 44 45 51 52 53 54 55 56

57 58 59 61 62 66 67 68a 68b 69 Probably High Risk HPV Types detected by the VisionArray ® HPV Chip

82 82 70 72 73 81 IS39 MM4 83 84 90 91 26 34 53 66 67 68a

+ 35 34 33 31 26 18 16 11 6 82 82 68b 69 70 73 IS39 MM4 54 53 52 51 45 44 43 42 40 39

® 68a 67 66 62 61 59 58 57 56 55 Low Risk HPV Types detected by the VisionArray HPV Chip

81 73 72 70 69 68b 6 11 40 42 43 44 54 55 57

82 82 GD 91 90 84 83 MM4 IS39 61 62 72 81 83 84 90 91

Low Risk References High Risk Colombo N, et al. (2012) Ann Oncol 23 Suppl 7: vii27-32. Crow JM, et al. (2012) Nature 488: S2-S3. Probably High Risk Guide Dots (GD)/Positive Control (+) IARC (2012) Biological Agents. IARC Monogr Eval Carcinog Risks Hum, 100B: 1-441. Poljak M, et al. (2016) J Clin Virol 76 Suppl 1: S3-S13.

Prod. No. Product Tests VA-0001-10 VisionArray HPV Chip 1.0 Incl. 10 pieces 10 VA-0001-50 VisionArray HPV Chip 1.0 Incl. 5x 10 pieces 50

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

260 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 260 VAE001-1-20 VisionArray Arrays for DNA analysis

VisionArray ® HPV High Risk Chip 1.0

Introduction The VisionArray ® HPV High Risk Chip 1.0 is intended to be used for the qualitative detection and genotyping of PCR amplificates of 24 clinically relevant human papillomavirus (HPV) genotypes that have been produced with the help of the VisionArray HPV Primer Kit 2.0 and the VisionArray Detection Kit. HPV has been conclusively identified as the major risk factor for cervical cancer. It is the third most common cancer in women worldwide, with an estimated number of 530,000 new cases and 280,000 deaths each year. Over the last years, the relevance of HPV in the history of oropharyngeal cancers has become more and more important which is indicated by a dramatically risen number of cancers of the oral cavity and pharynx linked to HPV.

At present, there are more than 150 different HPV types described. Several HPV types were classified as High Risk and Probably High Risk types based on their association with cervical cancer.

Chip Description The VisionArray ® HPV High Risk Chip 1.0 is designed to detect 24 clinically relevant HPV genotypes. All capture sequences and the positive control are set up on the Chip as duplicates and the guide dots as triplicates. The signals are visible on the Chip as dark blue areas. The automated evaluation of the results is performed by a VisionArray ® Analyzer Software.

High Risk HPV Types detected by the HPV High Risk Chip GD GD 16 18 31 33 35 39 + 45 51 52 56 58 59 16 18 26 31 39 35 34 33

45 51 52 53 66 59 58 56 Probably High Risk HPV Types detected by the HPV High Risk Chip

82 82 26 34 53 66 67 68a 67 68a 68b 69 MM4 IS39 73 70

82 82 39 35 34 33 16 18 26 31 68b 69 70 73 IS39 MM4 66 59 58 56 45 51 52 53 References Colombo N, et al. (2012) Ann Oncol 23 Suppl 7: vii27-32. 82 82 Crow JM, et al. (2012) Nature 488: S2-S3. MM4 IS39 73 70 67 68a 68b 69 IARC (2012) Biological Agents. IARC Monogr Eval Carcinog Risks Hum, 100B: 1-441. Poljak M, et al. (2016) J Clin Virol 76 Suppl 1: S3-S13. +

High Risk Guide Dots (GD)/Positive Control (+) Probably High Risk

Prod. No. Product Tests VA-0002-10 VisionArray HPV High Risk Chip 1.0 Incl. 10 pieces 10 VA-0002-50 VisionArray HPV High Risk Chip 1.0 Incl. 5x 10 pieces 50

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

261 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 261 VAE002-1-20 VisionArray Arrays for DNA analysis

VisionArray ® MYCO Chip 1.0

Introduction The VisionArray ® MYCO Chip 1.0 is intended to be used with a VisionArray ® Analysis Package for the qualitative detection and identification of PCR amplificates of the generaMycobacterium, Mycobacteroides, Mycolicibacillus, Mycolicibacter, and Mycolicibac- terium as well as several clinically relevant mycobacterial species that have been produced with the help of the VisionArray ® MYCO Primer Kit 1.0 or the VisionArray ® MYCO PreCise Master Mix. The mycobacterial genera comprise more than 140 species, which, for the purpose of diagnosis and treatment, have been grouped into three categories: M. tuberculosis complex (MTC), M. leprae, and non-tuberculous mycobacteria (NTM). The majority of the Mycobacterium species belongs to the NTM group and can be found in different environments. Many of these bac- teria cause life-threatening infections in humans and in recent years, the mortality and morbidity associated with NTMs has increased especially in immunocompromised patients worldwide. Treatment of NTMs is specific to each species and therefore a clear distinction between the present species is of extreme importance. Reliable and rapid molecular diagnostics are the basis of an adequate therapy that is given by the VisionArray ® MYCO Chip 1.0.

Chip Description The VisionArray ® MYCO Chip 1.0 is designed to detect several clinically relevant mycobacterial species. All capture sequences and the positive control are set up on the Chip as duplicates and the guide dots as triplicates. The signals are visible on the Chip as dark blue areas. The automated evaluation of the results is performed by a VisionArray ® Analyzer Software.

M. tuberculosis (MTC) complex GD GD 2 M. tuberculosis complex (ITS Region)

3 M. tuberculosis complex (IS6110 Region) + 1 2 3 4 5 Nontuberculous Mycobacteria (NTM) 6 7 8 9 10 11 4 M. abscessus 11 M. malmoense 12 13 14 15 16 17 M. avium / 5 12 M. marinum / M. ulcerans 5 4 3 2 1 + M. intracellulare complex M. scrofulaceum / 6 M. chelonae 13 11 10 9 8 7 6 M. parascrofulaceum

17 16 15 14 13 12 7 M. fortuitum 14 M. simiae

8 M. genavense 15 M. smegmatis

GD 9 M. haemophilum 16 M. szulgai

10 M. kansasii 17 M. xenopi GD Guide Dot + Positive Control 1 MYCO spec.

References Griffith DE, et al. (2007) Am J Respir Crit Core Med 175: 367-416. Gupta RS, et al. (2018) Front Microbiol 9: 67. Oren A & Carrity GM (2019) Int J Syst Evol Microbiol 69: 597-9. Perez-Martinez I, et al. (2013) BMC Res Notes 6: 531. Simons S, et al. (2011) Emerg Infect Dis 17: 343-9. Tortoli E (2009) Clin Microbiol Infect 15: 906-10.

Prod. No. Product Tests VA-0003-10 VisionArray MYCO Chip 1.0 Incl. 10 pieces 10 VA-0003-50 VisionArray MYCO Chip 1.0 Incl. 5x 10 pieces 50

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

262 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 262 VAE003-1-20 VisionArray Arrays for DNA analysis

VisionArray ® DNA Extraction, PCR, and Detection

VisionArray ® Detection Kit For hybridization and detection of PCR products on VisionArray ® Chips

Prod. No. Product Tests VK-0003-50 VisionArray Detection Kit 50 Incl. Hybridization Solution, 1 ml; Detection Solution, 5 ml; Blue Spot Solution, 5 ml; 100x Wash Buffer, 250 ml

VisionArray ® DNA Extraction Kits For isolation of genomic DNA from FFPE as well as liquid based cytology specimens

Prod. No. Product Tests VI-0001-50 VisionArray FFPE DNA Extraction Kit 50 Incl. Paraffin Dissolver; Tissue Lysis Buffer; Decrosslink Buffer; DNA Wash Buffer; Proteinase K; Proteinase K Buffer; Elution Buffer; Columns; Collection Tubes VI-0002-50 VisionArray Cytology DNA Extraction Kit 50 Incl. Pre-Lysis Buffer; Cell Lysis Buffer; DNA Wash Buffer; Proteinase K; Proteinase K Buffer; Elution Buffer; Columns; Collection Tubes

VisionArray ® PCR Reagents For contamination-free amplification and biotinylation of target sequences with a high quality heat stable Taq polymerase

Prod. No. Product Tests VP-0001-50 VisionArray HPV Primer Kit 2.0 50 Incl. HPV Primer Mix 2.0; dNTP/dUTP Solution VP-0002-50 VisionArray MYCO Primer Kit 1.0 50 Incl. MYCO Primer Mix 1.0; dNTP/dUTP Solution VE-0001-100 VisionArray PreCise Taq DNA Polymerase 100

Incl. VisionArray PreCise Taq DNA Polymerase; PreCise Reaction Buffer, 10x; PreCise MgCl2, 25 mM VE-0002-100 VisionArray Uracil-DNA Glycosylase 100 ES-0007-50 VisionArray HPV PreCise Master Mix 50

Containing HPV Primer Mix 2.0; dNTP/dUTP Solution; VisionArray PreCise Taq DNA Polymerase; PCR-Buffer; MgCl2; VisionArray Uracil-DNA Glycosylase ES-0008-50 VisionArray MYCO PreCise Master Mix 50

Containing MYCO Primer Mix 1.0; dNTP/dUTP Solution; VisionArray PreCise Taq DNA Polymerase; PCR-Buffer; MgCl2; VisionArray Uracil-DNA Glycosylase

Slide Centrifuge Prod. No. Product E-4051-1 Mini Slide Centrifuge Incl. 2 place slide rotor; two slide holders

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

263 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 263 AVAE001-1-20 VisionArray Arrays for DNA analysis

VisionArray ® Analysis Package SingleScan For visualization and quick analysis of the VisionArray ® Chips data

3 5 Legend 1 Scanner 8100 2 2 Hand Scanner 3 Slide Holder 6 4 Laptop  5 USB-Hub 6 External Hard Drive 4 7 Computer Mouse

Prod. No. Product E-4060-1 VisionArray Analysis Package SingleScan Incl. Scanner 8100; Slide Holder SingleScan; Hand Scanner; PC with preinstalled VisionArray Analyzer Software SingleScan; USB-Hub; External Hard Drive; Computer Mouse

VisionArray ® Analyzer Software SingleScan • Simple visualization and quick analysis of the VisionArray ® Chip data • Analysis of a Chip and the report of the results can be achieved in just a few minutes • Program navigation is easy and intuitive for the user • Scans are stored including all sample and Chip data in an integrated database on the enclosed external hard drive

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

264 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 264 AVAE002-1-20 VisionArray Arrays for DNA analysis

VisionArray ® Analysis Package MultiScan For visualization and quick analysis of up to 6 VisionArray ® Chips simultaneously 3 5

Legend 4 1 Scanner V600 Photo 2 Laptop 3 USB-Hub 4 External Hard Drive  5 Computer Mouse 2

Prod. No. Product E-4070-1 VisionArray Analysis Package MultiScan Incl. Scanner V600 Photo; Slide Holder MultiScan; PC with preinstalled VisionArray Analyzer Software MultiScan; USB-Hub; External Hard Drive; Computer Mouse

VisionArray ® Analyzer Software MultiScan • Simple visualization and quick analysis of up to 6 VisionArray ® Chips simultaneously • All available VisionArray ® Chips can be combined using the Scanner V600 Photo and are automatically detected by the software offering maximum flexibility • Analysis of the Chips and the report of the results can be achieved in just a few minutes • Scans are stored including all sample and Chip data in an integrated database on the enclosed external hard drive

only available in certain countries. All other countries research use only! Please contact your local dealer for more information.

265 42 life sciences GmbH & Co. KG · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 265 AVAE003-1-20 General Information

Product Use All products manufactured and/or distributed by ZytoVision GmbH should be used in accordance with the labeled intended use of the given product. Products labeled as CE/IVD are in vitro diagnostic medical devices according to the European Directive 98/79/EC. Products labeled as “for research use only” should be used for research applications, not for diagnostic procedures. Trademarks ZytoDot ®, ZytoFast ®, ZytoLight ®, ZytoMation ®, FlexISH ® and ZytoVision® are registered trademarks of ZytoVision GmbH. VisionArray ® is a registered trademark of 42 life sciences GmbH & Co. KG, Bremerhaven, Germany. CEN™, SPEC™, ZyBlack™ , ZyBlue™, ZyGold™, ZyGreen™, ZyOrange™, ZyRed™, ZytoDot ® 2C™, TriCheck™ , DuraTect™ and DistinguISH™ are trademarks of ZytoVision GmbH. TexasRed ® is a registered trademark of Molecular Probes, Inc., Eugene, OR. BOND™ is a trademark of Leica Biosystems and its affiliates. CE Marking & ISO Certificates All probes in this catalog are manufactured by ZytoVision GmbH, Bremerhaven, Germany. ZytoLight ®, ZytoMation ®, and FlexISH ® probes are direct labeled using the unique ZytoLight ® Direct Label System II (ZytoVision GmbH) providing improved signal intensity. Advanced specificity of single copyZyto Light ®, ZytoMation ®, FlexISH ®, ZytoDot ®, and ZytoDot ® 2C™ probes is obtained by the unique ZytoVision® Repeat Subtraction Technique (ZytoVision GmbH). ZytoFast ® probes are tagged using the unique ZytoFast ® HighTag System (ZytoVision) providing improved signal intensity. Product development and manufacturing of all products by ZytoVision GmbH is carried out according to ISO 9001and ISO 13485 regulations, for which ZytoVision GmbH holds certificates. These certificates were issued and are annually monitored by mdc medical device certification GmbH, Germany. mdc was, as one of the first German entities, notified in 1994 by the German Ministry of Health to the European Commission for conformity assessment procedures under the European Directive 98/79/EC for in vitro diagnostic devices. Current certificates can be downloaded at www.zytovision.com.

266 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 266 Index

Prod. No. Product Page Prod. No. Product Page

AB-0001-4/-30 Mouse-anti-DIG 245, 257 C-3055-100/-400 ZytoDot 2C SPEC ALK Break Apart Probe 210 AB-0002-4 Anti-Mouse-HRP-Polymer 245, 257 C-3056-400 ZytoDot 2C SPEC FGFR2/CEN 10 Probe 223 AB-0011-4 Rabbit-anti-DIG 257 C-3057-400 ZytoDot 2C SPEC MET/CEN 7 Probe 216 AB-0012-4 Anti-Rabbit-AP-Polymer 257 C-3058-400 ZytoDot 2C SPEC ERG Break Apart Probe 240 AB-0013-4 HRP/AP-Polymer-Mix 245 C-3059-400 ZytoDot 2C SPEC EML4 Break Apart Probe 211 AB-0014-4 Anti-DIG/DNP-Mix 245 C-3062-400 ZytoDot 2C SPEC CDK4/CEN 12 Probe 226 AB-0015-4 Anti-Biotin/DIG-Mix 257 C-3063-100/-400 ZytoDot 2C SPEC ROS1 Break Apart Probe 213 BS-0001-4 Blocking Solution 245 C-3064-100/-400 ZytoDot 2C SPEC RET Break Apart Probe 221 BS-0002-8 ZyBlack™ Quenching Solution 26, 187 C-3065-100 ZytoDot 2C SPEC FOXO1 Break Apart Probe 229 C-3001-400 ZytoDot SPEC ERBB2 Probe 233 C-3066-400 ZytoDot 2C SPEC MYC Break Apart Probe 219 C-3002-400 ZytoDot CEN 6 Probe 242 f. C-3067-400 ZytoDot 2C SPEC CDKN2A/CEN 9 Probe 220 C-3003-40 ZytoDot SPEC ERBB2 Probe Kit 233 C-3068-100 ZytoDot 2C SPEC ERBB2/D17S122 Probe 235 C-3004-40 ZytoDot Pretreatment Kit 244 C-3071-100 ZytoDot 2C SPEC IGH Break Apart Probe 230 C-3005-40 ZytoDot CISH Polymer Detection Kit 244 C-3072-100 ZytoDot 2C SPEC MALT1 Break Apart Probe 239 C-3006-400 ZytoDot CEN 17 Probe 242 f. C-3073-100 ZytoDot 2C SPEC BCL2 Break Apart Probe 238 C-3007-400 ZytoDot SPEC EGFR Probe 214 C-3074-100 ZytoDot 2C SPEC BCL6 Break Apart Probe 212 C-3008-400 ZytoDot CEN 7 Probe 242 f. C-3075-100 ZytoDot 2C SPEC CCND1 Break Apart Probe 224 C-3011-40 ZytoDot Wash Buffer Set 245 C-3076-10/-40 ZytoDot 2C Glioma 1p/19q Probe Set 206 C-3012-400 ZytoDot SPEC MDM2 Probe 227 C-3077-100 ZytoDot 2C SPEC USP6 Break Apart Probe 232 C-3013-400 ZytoDot SPEC MYC Probe 218 CS-0001-20 Mayer's Hematoxylin Solution 245, 257 C-3014-400 ZytoDot CEN 12 Probe 242 f. CS-0002-20 Nuclear Blue Solution 245, 257 C-3015-100 DAB Solution Set 245, 257 CS-0003-20 Nuclear Red Solution 257 C-3016-400 ZytoDot CEN 8 Probe 242 f. CS-0004-20 Nuclear Green Solution 257 C-3018-40 ZytoDot CISH Implementation Kit 244 E-4005-50/-125 Fixogum Rubber Cement 185, 245, 257 C-3020-400 ZytoDot CEN Yq12 Probe 242 f. E-4007-2 ERBB2 Control Slide Set 245 C-3022-10/-40 ZytoDot 2C SPEC ERBB2/CEN 17 Probe Kit 234 E-4010-1 DAPI/ZyGreen™/ZyOrange™ Triple Bandpass Filter Set 188 C-3025-400 ZytoDot CEN X Probe 242 f. E-4012-1 ZyGreen™ Single Bandpass Filter Set v2 188 C-3026-400 ZytoDot SPEC 21q22 Probe 242 f. E-4013-1 ZyOrange™ Single Bandpass Filter Set v2 188 C-3028-40 ZytoDot 2C CISH Polymer Detection Kit 244 E-4016-1 ZyGreen™/ZyOrange™ Dual Bandpass Filter Set v2 188 C-3029-400 ZytoDot SPEC MYCN Probe 209 E-4017-1 ZyRed™ Single Bandpass Filter Set v2 188 C-3032-100/-400 ZytoDot 2C SPEC ERBB2/CEN 17 Probe 234 E-4026-1 ZyBlue™ Single Bandpass Filter Set v2 188 C-3033-100/-400 ZytoDot 2C SPEC EGFR/CEN 7 Probe 215 E-4027-1 ZyGold™ Single Bandpass Filter Set v2 188 C-3035-400 ZytoDot SPEC 1p12 Probe 242 f. E-4028-1 ZyBlue™/ZyGreen™/ZyOrange™ Triple Bandpass Filter Set 188 C-3036-100/-400 ZytoDot 2C SPEC 1p36/1q25 Probe 207 E-4030-1 DAPI Single Bandpass Filter Set v2 188 C-3037-100/-400 ZytoDot 2C SPEC 19q13/19p13 Probe 208 E-4051-1 Mini Slide Centrifuge 263 C-3038-100 ZytoDot AP-Red Solution Set 245 E-4060-1 VisionArray Analysis Package SingleScan 264 C-3039-100 ZytoDot HRP-Green Solution Set 245, 257 E-4070-1 VisionArray Analysis Package MultiScan 265 C-3040-400 ZytoDot 2C SPEC TOP2A/CEN 17 Probe 236 E-4111-1 ZEISS Fluorescence Filter Holder "FL EC P&C" 188 C-3043-100 ZytoDot 2C SPEC EWSR1 Break Apart Probe 241 E-4113-1 ZEISS Fluorescence Filter Holder "FL" 188 C-3044-10/-40 ZytoDot 2C CISH Implementation Kit 244 E-4121-1 OLYMPUS Fluorescence Filter Holder "U-MF 2" 188 C-3045-400 ZytoDot CEN 3 Probe 242 f. E-4122-1 OLYMPUS Fluorescence Filter Holder "U-FF" 188 C-3046-100 ZytoDot 2C SPEC SS18 Break Apart Probe 237 E-4131-1 LEICA Fluorescence Filter Holder "DM K" 188 C-3047-100 ZytoDot 2C SPEC DDIT3 Break Apart Probe 225 E-4141-1 NIKON Fluorescence Filter Holder "C-FL" 188 C-3048-400 ZytoDot 2C CEN X/Y Probe 242 f. ES-0001-4/-50/-1000 Pepsin Solution 185, 244, 257 C-3049-100/-400 ZytoDot 2C SPEC MDM2/CEN 12 Probe 228 ES-0001-8 Pepsin Solution Set 185, 244, 257 C-3050-400 ZytoDot 2C SPEC FGFR1/CEN 8 Probe 217 ES-0002-4/-50 Cytology Pepsin Solution 185 C-3051-400 ZytoDot SPEC 2q11 Probe 242 f. ES-0007-50 VisionArray HPV PreCise Master Mix 263 C-3052-400 ZytoDot SPEC 13q12 Probe 242 f. ES-0008-50 VisionArray MYCO PreCise Master Mix 263 C-3053-400 ZytoDot 2C SPEC PTEN/CEN 10 Probe 222 MT-0004-4 Mounting Solution (alcoholic) 245, 257 C-3054-100 ZytoDot 2C SPEC FUS Break Apart Probe 231 MT-0007-0.8 DAPI/DuraTect™-Solution 186

267 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 267 IND001-1-20 Index

Prod. No. Product Page Prod. No. Product Page

MT-0008-0.8 DAPI/DuraTect™-Solution (ultra) 186 Z-2008-200 ZytoLight CEN X Probe 170 f. PT-0001-1000 Heat Pretreatment Solution Citric 185 Z-2010-200 ZytoLight CEN Yq12 Probe 170 f. PT-0002-500 Heat Pretreatment Solution EDTA 244, 257 Z-2013-50/-200 ZytoLight SPEC MDM2/CEN 12 Dual Color Probe 124 PT-0006-100 Formaldehyde Dilution Buffer Set 185 Z-2014-50/-200 ZytoLight SPEC MAML2 Dual Color Break Apart Probe 108 SB-0004-4 NBT/BCIP Solution 257 Z-2015-50/-200 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe 143 SB-0005-4 AEC Solution 257 Z-2016-50/-200 ZytoLight CEN X/Yq12 Dual Color Probe 170 f. T-1005-40 ZytoFast human lg-kappa/lg-lambda CISH Kit 254 f. Z-2018-50/-200 ZytoLight SPEC FOXO1/PAX3 Dual Color Single Fusion Probe 126 T-1017-400 ZytoFast human lg-kappa/lg-lambda Probe 254 f. Z-2019-50/-200 ZytoLight SPEC FOXO1/PAX7 Dual Color Single Fusion Probe 128 T-1053-400 ZytoFast DNA (+) Control Probe 257 Z-2020-5/-20 ZytoLight SPEC ERBB2/CEN 17 Dual Color Probe Kit 143 T-1054-400 ZytoFast DNA (-) Control Probe 257 Z-2028-5/-20 ZytoLight FISH-Tissue Implementation Kit 184 T-1055-400 ZytoFast HPV type 6/11 Probe 251 Z-2033-50/-200 ZytoLight SPEC EGFR/CEN 7 Dual Color Probe 74 T-1056-400 ZytoFast HPV type 16/18 Probe 251 Z-2049-200 ZytoLight SPEC 2q11 Probe 170 f. T-1057-400 ZytoFast HPV type 31/33 Probe 251 Z-2050-200 ZytoLight CEN 12 Probe 170 f. T-1061-40 ZytoFast PLUS CISH Implementation Kit AP-NBT/BCIP 256 Z-2056-200 ZytoLight SPEC ERBB3/CEN 12 Dual Color Probe 121 T-1063-40 ZytoFast PLUS CISH Implementation Kit HRP-DAB 256 Z-2057-200 ZytoLight SPEC ERBB4/2q11 Dual Color Probe 41 T-1105-40 ZytoFast human lg-kappa/lg-lambda Permanent CISH Kit 254 f. Z-2062-200 ZytoLight SPEC FHIT/CEN 3 Dual Color Probe 45 T-1113-400 ZytoFast CMV Probe 253 Z-2063-50/-200 ZytoLight SPEC CDKN2A/CEN 9 Dual Color Probe 89 T-1114-400 ZytoFast EBV Probe 252 Z-2067-200 ZytoLight CEN 9 Probe 170 f. T-1115-400 ZytoFast human lg-kappa Probe 254 f. Z-2069-50/-200 ZytoLight SPEC ESR1/CEN 6 Dual Color Probe 72 T-1116-400 ZytoFast human lg-lambda Probe 254 f. Z-2071-50/-200 ZytoLight SPEC CCND1/CEN 11 Dual Color Probe 106 T-1119-400 ZytoFast RNA (-) Control Probe 257 Z-2072-50/-200 ZytoLight SPEC FGFR1/CEN 8 Dual Color Probe 82 T-1120-400 ZytoFast 28S rRNA (+) Control Probe 257 Z-2074-50/-200 ZytoLight SPEC MYCN/2q11 Dual Color Probe 35 T-1140-400 ZytoFast HPV High-Risk (HR) Types Probe 251 Z-2075-50/-200 ZytoLight SPEC 1p36/1q25 Dual Color Probe 27 T-1144-400 ZytoFast HPV Screening Probe 251 Z-2076-50/-200 ZytoLight SPEC 19q13/19p13 Dual Color Probe 28 T-1151-40 ZytoFast PLUS CISH Implementation Kit AP-Permanent Red 256 Z-2077-50/-200 ZytoLight CEN 17/SPEC ERBB2 Dual Color Probe 144 VA-0001-10/-50 VisionArray HPV Chip 1.0 260 Z-2078-50/-200 ZytoLight SPEC PTEN/CEN 10 Dual Color Probe 98 VA-0002-10/-50 VisionArray HPV High Risk Chip 1.0 261 Z-2079-200 ZytoLight CEN 10 Probe 170 f. VA-0003-10/-50 VisionArray MYCO Chip 1.0 262 Z-2080-200 ZytoLight SPEC MDM4/1p12 Dual Color Probe 34 VE-0001-100 VisionArray PreCise Taq DNA Polymerase 263 Z-2081-50/-200 ZytoLight SPEC CDKN2A/CEN 3/7/17 Quadruple Color Probe 90 VE-0002-100 VisionArray Uracil-DNA Glycosylase 263 Z-2082-200 ZytoLight SPEC FGFR3/4p11 Dual Color Probe 53 VI-0001-50 VisionArray FFPE DNA Extraction Kit 263 Z-2083-200 ZytoLight SPEC 4p11 Probe 170 f. VI-0002-50 VisionArray Cytology DNA Extraction Kit 263 Z-2084-200 ZytoLight SPEC VHL/CEN 3 Dual Color Probe 42 VK-0003-50 VisionArray Detection Kit 263 Z-2085-200 ZytoLight SPEC 13q12 Probe 170 f. VP-0001-50 VisionArray HPV Primer Kit 2.0 263 Z-2086-200 ZytoLight SPEC 21q22 Probe 170 f. VP-0002-50 VisionArray MYCO Primer Kit 1.0 263 Z-2087-50/-200 ZytoLight SPEC MET/CEN 7 Dual Color Probe 76 WB-0001-560 Wash Buffer SSC 184, 245 Z-2090-50/-200 ZytoLight SPEC MYC Dual Color Break Apart Probe 84 WB-0002-50 25x Wash Buffer A 184 Z-2091-50/-200 ZytoLight SPEC TERT/5q31 Dual Color Probe 57 WB-0003-50 20x SSC Solution 184 Z-2092-50/-200 ZytoLight SPEC MYC/CEN 8 Dual Color Probe 85 WB-0004-1000 PBS/Tween 245 Z-2093-50/-200 ZytoLight SPEC ERBB2/TOP2A/CEN 17 Triple Color Probe 146 WB-0005-50 20x Wash Buffer TBS 184, 245, 257 Z-2095-50/-200 ZytoLight SPEC 13/CEN 18/SPEC 21 Triple Color Probe 170 f. WB-0007-500 Cytology Stringency Wash Buffer SSC 184 Z-2096-50/-200 ZytoLight SPEC EWSR1 Dual Color Break Apart Probe 165 WB-0008-500 Cytology Wash Buffer SSC 184 Z-2097-50/-200 ZytoLight SPEC SS18 Dual Color Break Apart Probe 149 WB-0010-500 5x FlexISH Wash Buffer 184 Z-2099-20 ZytoLight FISH-Cytology Implementation Kit 184 Z-2001-200 ZytoLight CEN 3 Probe 170 f. Z-2100-50/-200 ZytoLight SPEC DDIT3 Dual Color Break Apart Probe 122 Z-2002-200 ZytoLight CEN 6 Probe 170 f. Z-2101-200 ZytoLight SPEC 1p12 Probe 170 f. Z-2003-200 ZytoLight CEN 7 Probe 170 f. Z-2102-200 ZytoLight SPEC VHL/1p12/CEN 7/17 Quadruple Color Probe 43 Z-2004-50/-200 ZytoLight CEN 8 Probe 170 f. Z-2103-50/-200 ZytoLight SPEC CDK4/CEN 12 Dual Color Probe 123 Z-2005-200 ZytoLight CEN 11 Probe 170 f. Z-2104-5/-20 ZytoLight Aneuploidy Panel X/Y and 13/18/21 173 Z-2006-200 ZytoLight CEN 17 Probe 170 f. Z-2105-50/-200 ZytoLight SPEC MYC/IGH Dual Color Dual Fusion Probe 86 Z-2007-200 ZytoLight CEN 18 Probe 170 f. Z-2107-50/-200 ZytoLight SPEC EGR1/5p15 Dual Color Probe 59

268 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 268 IND001-1-20 Index

Prod. No. Product Page Prod. No. Product Page

Z-2108-50/-200 ZytoLight SPEC CCND1 Dual Color Break Apart Probe 105 Z-2171-200 ZytoLight SPEC BCL2L1/CEN 20 Dual Color Probe 157 Z-2109-50/-200 ZytoLight SPEC TFE3 Dual Color Break Apart Probe 169 Z-2173-200 ZytoLight SPEC MCL1/1p12 Dual Color Probe 30 Z-2110-50/-200 ZytoLight SPEC IGH Dual Color Break Apart Probe 130 Z-2174-50 ZytoLight SPEC BCL2/CEN 18 Dual Color Probe 152 Z-2111-50/-200 ZytoLight SPEC BCR/ABL1 Dual Color Dual Fusion Probe 94 Z-2175-50 ZytoLight SPEC YWHAE Dual Color Break Apart Probe 142 Z-2112-50/-200 ZytoLight SPEC RUNX1/RUNX1T1 Dual Color Dual Fusion Probe 83 Z-2176-50/-200 ZytoLight SPEC ETV6 Dual Color Break Apart Probe 118 Z-2113-50/-200 ZytoLight SPEC PML/RARA Dual Color Dual Fusion Probe 132 Z-2177-50/-200 ZytoLight SPEC BCL6 Dual Color Break Apart Probe 51 Z-2114-50/-200 ZytoLight SPEC BCL2/IGH Dual Color Dual Fusion Probe 153 Z-2178-50 ZytoLight SPEC SMARCB1/22q12 Dual Color Probe 164 Z-2115-200 ZytoLight SPEC KRAS/CEN 12 Dual Color Probe 120 Z-2179-50/-200 ZytoLight SPEC CD274,PDCD1LG2/CEN 9 Dual Color Probe 87 Z-2116-50/-200 ZytoLight SPEC COL1A1/PDGFB Dual Color Dual Fusion Probe 148 Z-2180-200 ZytoLight SPEC 21/CEN X/Yq12 Triple Color Probe 170 f. Z-2117-50/-200 ZytoLight SPEC ALK/EML4 TriCheck™ Probe 36 Z-2181-200 ZytoLight SPEC NRG1 Dual Color Break Apart Probe 80 Z-2118-200 ZytoLight SPEC CCND1 Break Apart/2q11/CEN 6 Quadruple Color Probe 44 Z-2182-5/-20 FlexISH-Tissue Implementation Kit 184 Z-2119-50/-200 ZytoLight SPEC PDGFB Dual Color Break Apart Probe 167 Z-2183-50 ZytoLight SPEC EWSR1/FLI1 TriCheck™ Probe 166 Z-2120-200 ZytoLight CEN X/Y Dual Color Probe 170 f. Z-2184-50 ZytoLight SPEC SS18/SSX1 TriCheck™ Probe 150 Z-2121-200 ZytoLight SPEC COL1A1 Dual Color Break Apart Probe 147 Z-2185-50 ZytoLight SPEC FOXO1/PAX3 TriCheck™ Probe 127 Z-2122-200 ZytoLight SPEC FGFR2/CEN 10 Dual Color Probe 100 Z-2189-200 ZytoLight SPEC BRAF Dual Color Break Apart Probe 77 Z-2123-200 ZytoLight CEN Y (DYZ3) Probe 170 f. Z-2190-50/-200 ZytoLight SPEC ERBB2/D17S122 Dual Color Probe 145 Z-2124-50/-200 ZytoLight SPEC ALK Dual Color Break Apart Probe 37 Z-2191-200 ZytoLight SPEC BRAF/CEN 7 Dual Color Probe 78 Z-2125-50/-200 ZytoLight SPEC CCND1/IGH Dual Color Dual Fusion Probe 107 Z-2192-50/-200 ZytoLight SPEC BCL2 Dual Color Break Apart Probe 151 Z-2127-200 ZytoLight SPEC SOX2/CEN 3 Dual Color Probe 50 Z-2193-50/-200 ZytoLight SPEC KMT2A Dual Color Break Apart Probe 116 Z-2130-50 ZytoLight SPEC FUS Dual Color Break Apart Probe 135 Z-2194-200 ZytoLight SPEC NRG1/CD74 TriCheck™ Probe 79 Z-2131-50 ZytoLight SPEC KIF5B Dual Color Break Apart Probe 96 Z-2195-200 ZytoLight SPEC VEGFA/CEN 6 Dual Color Probe 67 Z-2132-50 ZytoLight SPEC JAZF1 Dual Color Break Apart Probe 73 Z-2196-50/-200 ZytoLight SPEC MALT1 Dual Color Break Apart Probe 154 Z-2135-200 ZytoLight SPEC ERG/TMPRSS2 TriCheck™ Probe 160 Z-2197-50 ZytoLight SPEC PDGFRB Dual Color Break Apart Probe 63 Z-2136-50 ZytoLight SPEC EML4 Dual Color Break Apart Probe 39 Z-2198-50 ZytoLight SPEC TP53/17q22 Dual Color Probe 139 Z-2137-50 ZytoLight SPEC CARS Dual Color Break Apart Probe 101 Z-2199-50 ZytoLight SPEC ABL1 Dual Color Break Apart Probe 93 Z-2138-200 ZytoLight SPEC ERG Dual Color Break Apart Probe 159 Z-2200-50 ZytoLight SPEC ABL2 Dual Color Break Apart Probe 33 Z-2139-50 ZytoLight SPEC FOXO1 Dual Color Break Apart Probe 125 Z-2201-50 ZytoLight SPEC CRLF2 Dual Color Break Apart Probe 168 Z-2140-200 ZytoLight SPEC PIK3CA/CEN 3 Dual Color Probe 49 Z-2202-50 ZytoLight SPEC CSF1R Dual Color Break Apart Probe 61 Z-2142-50 ZytoLight SPEC WT1 Dual Color Break Apart Probe 103 Z-2203-50/-200 FlexISH ALK/ROS1 DistinguISH™ Probe 179 Z-2143-50/-200 ZytoLight SPEC MYB Dual Color Break Apart Probe 70 Z-2205-50/-200 ZytoLight SPEC NTRK2 Dual Color Break Apart Probe 91 Z-2144-50/-200 ZytoLight SPEC ROS1 Dual Color Break Apart Probe 68 Z-2206-50/-200 ZytoLight SPEC NTRK3 Dual Color Break Apart Probe 133 Z-2145-50 ZytoLight SPEC NR4A3 Dual Color Break Apart Probe 92 Z-2207-50 ZytoLight SPEC CBFB Dual Color Break Apart Probe 136 Z-2146-50/-200 ZytoLight SPEC BIRC3/MALT1 Dual Color Dual Fusion Probe 109 Z-2208-200 ZytoLight SPEC NUTM1 Dual Color Break Apart Probe 131 Z-2148-50/-200 ZytoLight SPEC RET Dual Color Break Apart Probe 97 Z-2209-50 ZytoLight SPEC PDGFRA/FIP1L1 TriCheck™ Probe 55 Z-2151-50 ZytoLight SPEC USP6 Dual Color Break Apart Probe 141 Z-2210-50 ZytoLight SPEC IRF4,DUSP22 Dual Color Break Apart Probe 64 Z-2152-50/-200 ZytoLight SPEC RREB1/MYB/CEN 6 Triple Color Probe 65 Z-2211-50 ZytoLight SPEC EGR1/D5S23,D5S721 Dual Color Probe 60 Z-2153-50/-200 ZytoLight SPEC TP53/CEN 17 Dual Color Probe 140 Z-2212-50 ZytoLight SPEC WWTR1 Dual Color Break Apart Probe 47 Z-2157-50/-200 ZytoLight SPEC ETV6/RUNX1 Dual Color Dual Fusion Probe 119 Z-2213-50 ZytoLight SPEC PTPRT/20q11 Dual Color Probe 158 Z-2159-50/-200 ZytoLight SPEC TP53/ATM Dual Color Probe 112 Z-2214-50 ZytoLight SPEC CUX1/EZH2/CEN 7 Triple Color Probe 75 Z-2160-50/-200 ZytoLight SPEC D13S319/13q34/CEN 12 Triple Color Probe 113 Z-2215-50 ZytoLight SPEC PHF1 Dual Color Break Apart Probe 66 Z-2161-200 ZytoLight SPEC ALK/2q11 Dual Color Probe 38 Z-2216-50 ZytoLight SPEC 11q gain/loss Triple Color Probe 115 Z-2162-200 ZytoLight SPEC ROS1/CEN 6 Dual Color Probe 69 Z-2265-50 ZytoLight SPEC NUP214 Dual Color Break Apart Probe 95 Z-2163-200 ZytoLight SPEC 18/CEN X/Y Triple Color Probe 170 f. Z-2266-50 ZytoLight SPEC NUP98 Dual Color Break Apart Probe 102 Z-2164-200 ZytoLight SPEC 13/21 Dual Color Probe 170 f. Z-2267-50 ZytoLight SPEC CREBBP Dual Color Break Apart Probe 134 Z-2165-50/-200 ZytoLight SPEC RB1/13q12 Dual Color Probe 129 Z-2268-50 ZytoLight SPEC CSF1R/D5S23,D5S721 Dual Color Probe 62 Z-2166-50/-200 FlexISH ERBB2/CEN 17 Dual Color Probe 183 Z-2269-50/-200 FlexISH RET/KIF5B TriCheck™ Probe 182 Z-2167-50/-200 ZytoLight SPEC NTRK1 Dual Color Break Apart Probe 32 Z-2270-50 ZytoLight SPEC MAF/IGH Dual Color Dual Fusion Probe 137 Z-2168-50/-200 ZytoLight SPEC FGFR1 Dual Color Break Apart Probe 81 Z-2271-50 ZytoLight SPEC MAFB/IGH Dual Color Dual Fusion Probe 138 Z-2169-200 ZytoLight SPEC FGFR2 Dual Color Break Apart Probe 99 Z-2272-20 ZytoLight Glioma 1p/19q Probe Set 26 Z-2170-50/-200 ZytoLight SPEC FGFR3 Dual Color Break Apart Probe 52 Z-2273-50 ZytoLight SPEC TERT Dual Color Break Apart Probe 56

269 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 269 IND001-1-20 Index

Prod. No. Product Page

Z-2274-50 ZytoLight SPEC C19MC/19p13 Dual Color Probe 156 Z-2275-50 ZytoLight SPEC ZNF384 Dual Color Break Apart Probe 117 Z-2276-50 ZytoLight SPEC CKS1B/CDKN2C Dual Color Probe 29 Z-2277-50 ZytoLight SPEC MEF2D/BCL9 TriCheck™ Probe 31 Z-2278-200 ZytoLight SPEC RICTOR/5q31.1 Dual Color Probe 58 Z-2279-20 ZytoLight Aneuploidy Panel 18/X/Y and 13/21 172 Z-2280-50 ZytoLight SPEC D13S319/13q34 Dual Color Probe 114 Z-2281-50 ZytoLight SPEC MYB/CEN 6 Dual Color Probe 71 Z-2282-50 ZytoLight SPEC FGFR3/IGH Dual Color Dual Fusion Probe 54 Z-2283-50/-200 FlexISH BCL2/BCL6 DistinguISH™ Probe 180 Z-2284-200 ZytoLight SPEC TERC/CEN 3 Dual Color Probe 48 Z-2285-50 ZytoLight SPEC CIC Dual Color Break Apart Probe 155 Z-2286-50 ZytoLight SPEC IGL Dual Color Break Apart Probe 163 Z-2287-50 ZytoLight SPEC GATA2/MECOM Dual Color Dual Fusion Probe 46 Z-2288-50 ZytoLight SPEC IGK Dual Color Break Apart Probe 40 Z-2289-50 ZytoLight SPEC DiGeorge Triple Color Probe 162 Z-2291-50 ZytoLight SPEC SPI1 Dual Color Break Apart Probe 104 Z-2292-5.1ML ZytoMation ERBB2/CEN 17 Dual Color FISH Probe 194 Z-2293-50 FlexISH MYC/IGH TriCheck™ Probe 181 Z-2294-50 ZytoLight SPEC JAK2 Dual Color Break Apart Probe 88 Z-2296-50 ZytoLight SPEC ATM/CEN 12 Dual Color Probe 111 Z-2297-50 ZytoLight SPEC ATM/CEN 11 Dual Color Probe 110 Z-2298-5.1ML ZytoMation ROS1 Dual Color Break Apart FISH Probe 193 Z-2299-50 ZytoLight SPEC DiGeorge/Phelan McDermid Dual Color Probe 161

270 ZytoVision GmbH · Fischkai 1 · 27572 Bremerhaven · Germany · www.zytovision.com 270 IND001-1-20 Prod. # M-5005 © January 2020 ZytoVision GmbH ZytoVision GmbH · Fischkai 1 27572 Bremerhaven · Germany Phone: +49 (0)471/4832-300 Fax: +49 (0)471/4832-509 [email protected] www.zytovision.com

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