DATA SHEET ViralSEQ Vesivirus Detection Assay ViralSEQ Vesivirus Detection Assay Integrated sample preparation and real-time PCR assay for the detection of vesivirus in cell culture samples
Table 1. Exclusion panel. The ViralSEQ Vesivirus Detection Assay has been demonstrated to be specific for vesivirus and does not detect RNA purified from other organisms.
RNA RNA Sample quantity Sample quantity Bacillus 2.5 ng Chicken 5 ng Clostridium 2.5 ng Dog 5 ng • Detection of known vesivirus strains E. coli 2.5 ng Human 5 ng, 1 µg • Rapid time-to-results of less than 5 hours Lactococcus 2.5 ng MDCK 5 ng ™ ™ • Applied Biosystems TaqMan assay–based real-time PCR Shigella 2.5 ng Rat 5 ng • Demonstrated sensitivity and specificity Yeast 5 ng CHO 500 ng • Proprietary optimized sample preparation for high
RNA recovery Vesivirus targets FAM FAM Viral contamination in mammalian product manufacturing presents a serious risk to the manufacturing process, the manufacturing facility, and the integrity of the product. Discriminatory RT-PCR positive control Vesivirus 2117 is an RNA virus that has reemerged as a FAM FAM potential threat to mammalian cell culture production. VIC VIC To meet the challenge of rapid, specific, and sensitive Internal positive control detection of vesivirus, we have developed the first NED commercially available integrated kit for isolation and detection of vesivirus from mammalian cell culture samples. The Applied Biosystems™ ViralSEQ™ Vesivirus Figure 1. Multiplex assay and control design. The unique design of the ViralSEQ Vesivirus Detection Assay includes two viral targets and Detection Assay is a PCR-based test that enables users discriminatory positive controls. This allows detection of known vesivirus to quickly isolate and detect vesivirus 2117 and all other genomes. The discriminatory positive control helps monitor the real- known strains with high sensitivity and specificity. The test time PCR reaction and cross-contamination of samples to reduce false positive results. utilizes Applied Biosystems™ PrepSEQ™ sample preparation magnetic beads to isolate viral RNA from a wide range of sample types, combined with TaqMan assay–based technology for sensitive and specific detection of the viral RNA. The kit also includes a proprietary positive control template that helps eliminate the possibility of false positive results arising from inadvertent contamination of test samples with the positive control. 40 10
35
1 30 t C Delta Rn 25 0.1
20
0.01 15 1 3 5 7 9 11 13 15 17 19 21 23 25 27 29 31 33 35 37 39 10−6 10−7 10−8 NTC Cycle
TCID50 280 28 2.8 Figure 3. High sensitivity and broad dynamic range. The ViralSEQ Vesivirus Detection Assay can detect from 106 copies down to 10 copies of in vitro–transcribed vesivirus RNA per reaction. −6 10 dilution (TCID50 = 280)
−7 10 dilution (TCID50 = 28)
−8 10 dilution (TCID50 = 2.8) 40 40 y = 1.4663Ln(x) 39.433 R2 = 0.9987 35 35
30 30 t t C C 25 25
20 20 qPCR efficiency 98 15 15 104 105 106 1 10 102 103 104 105 106 107 Number of cells Copy number Figure 2. Applied Biosystems™ PrepSEQ™ 1-2-3 Nucleic Acid Figure 4. High PCR efficiency. Using the viral RNA template, the Extraction Kit. Optimized for use with the ViralSEQ Vesivirus Detection ViralSEQ Vesivirus Detection Assay demonstrated high real-time PCR
Assay, the sample preparation procedure allows detection of TCID50 = 2.8 efficiency and a broad linear detection range. in 100 µL of cell culture and serum. In this example, 10–6 to 10 –8 dilutions of feline calicivirus (used as a surrogate for vesivirus) were spiked into 100 µL of fetal bovine serum (top panel) and cell culture (bottom panel). The serial dilutions of the virus correspond to TCID50 of 280, 28, and 2.8.
Ordering information
Product Quantity Cat. No.
ViralSEQ Vesivirus Detection Assay Contains real-time PCR assay, buffer and enzyme mix, positive control, 100 rxns 4448398C and negative control PrepSEQ 1-2-3 Nucleic Acid Extraction Kit 100 rxns 4452222 Contains buffers, magnetic beads, and enzymes
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For Research Use Only. Not for use in diagnostic procedures. © 2016 Thermo Fisher Scientific Inc. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. TaqMan is a trademark of Roche Molecular Systems, Inc., used under permission and license. COL11491 0516