Flash manuelle Interchim and Berthold Technologies collaboration
Interchim, a provider of consummables for life sciences, and Berthold Technologies, a leader in microplate instrumentation technology, have entered into a collaboration agreement to offer complete instrumentation and reagent solutions.
BERTHOLD TECHNOLOGIES provides with the Mithras and TriStar extremely versatile multimode readers for all comprehensive technologies used in today´s laboratory.
Additionally dedicated microplate readers for luminescence, fluorescence and absorbance can be offered for all common microplate formats. Petri dishes and Teraski plates can be measured with respective adapters. Powerful software allows kinetics, scanning, repeated mode, dual ratio measurements etc.
For higher troughput the instruments can be run with the Stacker LB 931. Robot access enables integration into robotic HTS systems.
Mithras LB 940 TriStar LB 941
For instrument informations, please contact :
Berthold France SAS Parc Technologique des Bruyères 8, route des Bruyères 78770 Thoiry France
Phone : (+33) 1 34 94 79 00 Fax : (+33) 1 34 94 79 01 E-mail : [email protected] 4 Summary Coelenterazines Luciferase HumanizedGaussiaLuciferase, GaussiaLuciferase FireflyandRenillaLuciferases FireflyandRenillaLuciferases RenillaLuciferase RenillaLuciferase constructs:siFLuc,pRNA ExpressionVectors FireflyLuciferase,recombinant,Mammalian :pCMVLuxA FireflyLuciferase1-Step FireflyLuciferase Reporter Gene Assays. NADHandNAD/NADH NAD/NADH,NADP/NADPH ADP ATP ATP, Calcein Livecellstaining Propidiummonoazide(PMA) Live/DeadMammalian Viability Dualstainingtodetectliveanddeadcells Fura-2 Viability &Cytotoxicity Ratiometriccalciumdyes Fluo-3 Singleλ aCella–AchE Calcium AcetylcholinesteraseDetection Cellulase CellulaseDetection Lysyl Oxidase Collagenandelastinstudy Lipase LipaseassaysandLipidstudy Protease MUP Acidphosphataseassays AlkalinePhosphatase Alkalinephosphataseassays forglycosidases samplerKit,Othercolorimetricsubstrates β-Galactosidasefluorescentsubstrates Enzymes Detection. Substratesforglycosidasesreportersystems(β-galactosidase,β-glucuronidase,β-glucosidase) D-Luciferins 400a H(Benzyl-Coelenterazine),Coelenterazine Coelenterazine,native,Coelenterazine ACella™ - Summary ADP, Phosphate&Pyrophosphate Assays.
Assay kit,0.1to100pmol, exc. Plus Assay Kit,RedFluorescence, Assay Kits AM, Fluo-8NW, Fluo-8H AM, Fura-PE3 acids Assay Kit,Unboundfreefatty / AM CellCounting&Viability em. Assay Kit,RedFluorescence TOX BioluminescenceCytotoxicity Ca ...... Acetylcholinesterase Assay Kit,RedFluorescence 2+ ...... indicators plasmid(pRLuc) MullereiLuciferase,recombinant,andpUC19 Assay Kit,Renilla ...... Assays. AM, Indo-1 Assay Kit,Blue,Green,RedFluorescence,Luminometric Assay Kits Assay Kit,2hreading,Stable Luc/Neoformonitoringtranscriptionalactivity Assay Kits Assay Kit Assay Kit,Live/DeadBacterialViability/Cytotoxicity kit,Ethidiummonoazide,bromide(EMA), ATP ...... AM, Fluo-8L AM, Indo-PE3 Phosphate
Assay
Assay Kit,SteadyGlow, BrightGlow, UniversalFluorimetric Kinase Assay Kit,CFDA Assays Assay
AM; Rhod-4™,Rhod-4NWCalcium Assay (GAPDH) Assay Kit,Pyrophosphate AM Assay kit SE,UptiBlueCellViability .com.com Assay Kit,3-5hreading, votre source d’innovations Assay Kit, &pCMVLuxB,FireflyluciferasesiRNA
Assay Kit i, Assay Kit,WST-8 CellProliferationandCytotoxicityAssayKit, Assay Kit,
28 23 17 13 12 10 41 36 32 29 27 22 21 21 20 19 18 17 16 15 24 11 7 7
Summary
Apoptosis Assays...... 43 Caspases study 43 Caspases Fluorometric HTS Assay Kits, Fluorimetric and Colorimetric Assay Kits, Caspase-6 and GranzymeB based apoptosis/toxicity Assays DNA Damage & Condensation study 46 Universal Chemiluminescent PARP Assay Kit, HT Chemiluminescent PARP/Apoptosis Assay, HT Chemiluminescent PARG Assay Kit, FlowTACS™ Apoptosis Detection Kits, Hoechst 33342 Mitochondrial events 49 JC-10 Mitochondria Membrane Potential Assay Kit, Safranine O, Glutathione Detection Kit, Monochlorobimane, Live Cell Glutathione Transferase Activity Kit Reactive Oxygen Species (ROS) 51
H2DCFDA, Carboxy-H2DCFDA, H2DCFDA-SE, Coelenterazine (native) , MCLA, Dihydroethidium (hydroethidine), dihydrorhodamine-1,2,3 (DHR 123), Dihydrocalcein AM, trans-1-(2'-Methoxyvinyl)pyrene, Hydroxyphenyl fluorescein (HPF) Assay Kit, Hypochlorite Detection Kit, APF (for hROS detection), ADHP Hydrogen peroxide Assay Kit, Myeloperoxidase Detection Kit, Catalase Detection Kit, Cis-Parinaric Acid (CPA), SOD Assay Kit with colorimetric substrate WST-1, Malachite green isothiocyanate Aldehyde oxidation study 58 Monoamine Oxidase A&B Detection Kit, SSAO Detection Kit Nitric Oxide (NO) assay 59 DAF-2 diacetate, DAF-FM, 2,3-Diaminonaphthalene, NBD Methylhydrazine for nitrite assay
DNA, Proteins and glucides Biochemistry Assays...... 60
Fluorescent DNA Assay 60 Summary EvaGreen - DNA quantification in solution Protein assays 61 Colorimetric Protein assays – BC Assay, Coo Assay, Fluorescent Protein assays – LavaPep Biochemistry tests for biological samples 61
Immunological Assays...... 62 ELISA – Luminescent detection (ECL) 62 UptiLight ELISA HRP Chemiluminescent Substrate, Luminometric Alkaline Phosphatase Assay Kit Other Colorimetric & Fluorimetric Enzymatic Substrates for Microplates assays 64 Uptima TMB ELISA chromogenic substrate, Substrates for horseradish peroxidase, Alkaline Phosphatase Fluorescent secondary reagents for ELISA (II Abs, Avidin) 66 Streptavidin conjugates, Secondary Antibodies conjugates ImmunoAssay - Buffers & Saturating Agents 68 Standard Buffers, Ready to use Buffers, SeaBlock saturating agent (non mammalian serum), Others Antibody Labeling Kits 69 Microspin biotinylation labeling kits-NH2 and –SH, FluoProbes® Protein Labeling Kits
Accessory tools...... 71 FPlyte Microplates 71 Bottom-free wells microplates - IMAplate Technology (Intelligent MultiFunctional Analysis) 72 Antifade Kit for Microplate 75
Legals - Trademarks...... 75
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 5 6 Reporter Gene Assays Our luciferaseassaykitsprovidealonglastingsignal(steadyglow)bypreventingtheformationofadenyl-oxyluciferinatenzymesurface. Gaussia princeps Renilla reniformis Photynus pyralis Species Interchim Firefly Luciferase Comparison ofdifferentspeciesluciferases luciferases, wealsooffer withthenewGaussialuciferasesforimprovedsignalintensity: reporter and tissues. as The Firefly However, oxyluciferin withemissionoflightcenteredon562nm(figure1). half-life through removinginhibitoryoxyluciferinfromtheenzymesurface.Butduration(10-15min)isstilltooshortforbatchprocessscreening. Technical tip
a 96 wellsmicroplatein340-800nmrange, with8-channels *Apollo LB912 assays . Includes avarietyoftechnologieswithsamplesinjectorsandrobotintegrationmodule. *Mithras LB940MultiModeReader luminescence reagentsandkitswerevalidatedwithBerthold Interchim Microplate Readers ...... Reading *Twinkle LB970Fluorometer exists alsoinaClinicalversion(LB961). A *Centro XSLB960Luminometer
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■ Firefly Luciferase 1-Step Assay Kit, 2 h reading
» Linear range – Assay linear over seven orders of magnitude » Limit of detection – less than 1 fg of luciferase per sample » No disposal problems or hazards are associated with the use of this luciferase assay kit » Reproducibility – CV less than 5%
Luciferase 1-Step assay system is a homogeneous high sensitivity firefly lu- ciferase reporter gene assay kit with a half-life of 2 hours for the quantification of firefly luciferase expression in mammalian cells. This kit is specially designed for batch processing systems using microplates such as 96-well plates. In ad- dition, Interchim Luciferase 1-Step assay kit offers higher sensitivity and wider dynamic range for detecting luciferase activity within mammalian cells, consis- tent reproducibility and cost effectiveness along with the added convenience of a one step assay. Reporter Gene Assays Description P/N : Qty Firefly Luciferase 1-Step Assay Kit FP-BX0320 100 ml (1000 tests in 96-well plate) FP-BX0321 100 ml (1000 tests in 96-well plate) Sensibility study on the Mithras luminometer from Berthold Technologies : Different quantities of Firefly luciferase on the range of 0.0001-1 µg (0.005-50 µg/ml) have been assayed.
■ Firefly Luciferase Stable Assay Kit, 3-5 h reading
» Simple : single step assay » High sensitivity : higher sensitivity than others steady substrates » Suitable for HTS - batch processing
This kit is a homogeneous high sensitivity firefly luciferase reporter gene assay kit with a half-life of 3-5 hours for the quantification of firefly luciferase expression in mammalian cells. It is specifically designed for batch processing systems using high-density microplates such as 384- and 1536-well plates, in high throughput environments.
Description P/N : Qty Firefly Luciferase Stable Assay Kit, 3-5 h reading FP-BU6870 10 ml FP-BU6871 100 ml FP-BU6872 1000 ml Kit contents (10/100/1000 ml) : 1 vial (2.5 /25 /250 mg) of D-Luciferin 1 bottle (10 /100/ 1000 mL) Firefly Assay Buffer 10 ml are sufficient for 100, 400 and 3,300 assays in 96-well, 384-well and 1536-well microplates.
■ Firefly Luciferase, recombinant, from Photinus pyralis
Luciferase can be used to detect trace amounts of ATP (signalling biological contamination). Less than or equal to one femtomole of ATP can be detected using 0.2 μg of luciferase. Recombinant Firefly luciferase can also be used to prepare standard curve of reporter genes for the study of gene expression.
Description P/N : Qty Firefly Luciferase, recombinant, from Photinus pyralis FP-D1826B 1 mg
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 7 8 Reporter Gene Assays » Please contactusforotherspromotersavailable: using BamHIandHindIIIforsiRNA It pRNA-U6.1/Neo/siFLuc pRNA-U6.1/Neo/siFLuc Description The targetsequencematchespGL3-control,andisdesignedtosilenceFireflyluciferaseexpressedbyaco-transfectedpGL3-controlvector. In ourcontrolexperiments,thissiRNA siFLuc aresiRNA ■ FireflyluciferasesiRNA luciferase, and1.6ugofpRNA-H1.1/Neo/siLuc » and1.6ugofpRNA-U6.1/Neo/siLuc » expression fortwoindependentvectorconstructs,uponthedualexpression. mammalian insertion the RNA These luciferase this as selection A oxygen of pCMVLuxA pCMVLuxA Description The A ■ MammalianExpressionVectors :pCMVLuxA Firefly Luciferase is heterodimer
singlereportertomonitorexpressionoftwoclonedgenes uses new catalyzed 42K pGL H1-siLuc :HEK293cellstransfected U6-siLuc :HEK293cellstransfected well LuxA bacterial light-emitting
splice luciferase vectors U6 :HEK293cellstransfectedwith and and as
of and or MammalianLuxBExpressionVector MammalianLuxA subunits, promoter and an other donor LuxB a cells. 37K encoded by luciferase amplification, long-chain ampicillin also can the marker constructsdesignedtoknockdownFireflyluciferase. genes respectively. reaction gene and These be include luxA for bacterial (positive controlinmammaliantransfection) by acceptor used isolated siRNA can is to gene
the resistance and aldehyde systems a ExpressionVector of be siRNA the a be luxA as the luciferase luxB CMV
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contains ampicillin a bacterium exists to expression. also developed visible reaction regulatory two NotI BG9670 P/N : DO8140 DO8130 P/N : .com.com monitor in for be molecular separate as votre source d’innovations mammalian sites encoding siRNA and light mammalian used resistance) an Vibrio harveyi with regulation based elements to the In at alpha-beta
construct as allow addition, FMNH2, 490 vectors. weights SD/SA- the a upon cells siRNA nm. two transfection. the 10 µg Qty for of in &pCMVLuxB for
vector firefly 20 µg 20 µg Qty pRL-TK U6: HEK293cellstransfectedwithpGL3-control(0.16ug)and (0.16 ug), and 1.6 ug of pRNA-U6.1/Neo empty vector. Firefly Luciferase
■ pRNA Luc/Neo for monitoring transcriptional activity
The pRNA-Luc/Neo includes a modified coding region for firefly (Photinus pyralis) luciferase that has been optimized for monitoring transcriptional activity in transfected eukaryotic cells. The purpose of this reporter vector is to screen for efficient siRNA for the target gene using Luc activity as a reporter gene. The principle is that when a siRNA silence the target gene by degrading mRNA, the Luc will not be expressed either, because the mRNA for both the gene and Luc as a whole molecule is degraded. The assay of this genetic reporter is rapid, sensitive and quantitative. Reporter Gene Assays
Description P/N : Qty pRNA-Luciferase-Neomycine DT3120 10 µg Related product β-Amyloid (1-40) HT8360 0.5 mg HT8361 1 mg
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 9 10 Reporter Gene Assays pUC19 pRLuc Description The Renillaproteinexpresseswellinbacteria. These proteinusescoelenterazineandderivativesassubstrate. ■ RenillaMullereiLuciferase,pUC19plasmid (pRLuc) activity, luciferase. translation. Renilla Luciferase Renilla MullereiLuciferase,95%purity(morethan8x10 Description ■ RenillaMullereiLuciferase,recombinant See "Coelenterazines",asstandaloneproducts Related substrates : Assay Enhancer Kit contents:Coelenterazine,RenillaLuciferaseLysis Buffer, Renilla Luciferase Description Renilla » Reporter geneusedasnormalizingtransfectioncontrol ■ RenillaLuciferase LuciferaseRenilla luciferase been The catalyzes assays gene » Fireflyluciferase and tissues. Coelenterazine alsoemitslightfromenzyme-independentoxidation(autoluminescence),enhancedbysuperoxideanionandperoxynitriteincells cell membranepermeabilityandquantumefficiency. These assay. luciferase reliable, linearresultswithminimalautoluminescencebackgroundandsuperiorsensitivity. using1ngtoµgDNA expressionandlightoutputfortransfection Sensitivity andLinearity:Linearcorrelationbetweenluciferasegene Conveniently packagedsubstrate enzyme regulation synthesized, coelenterazine or Renilla luciferase and has with as coelenterazine However, native may Through theuseofaspeciallydesignedcoelenterazinederivativeandbuffer formulation,theRenillaLuciferase does a been different normalizing luciferase, Assay Kit and function Coelenterazine has not reporterconstruct widely now function over analogs been require properties as commercially a oxidation used a transfection a dozen used in genetic monomeric all post-translational vitro in Assay Kit is function in as multiplex of the terms by and reporter a sizes permittingyoutoruna coelenterazine
natural control reporter oxygen available in as 36 of transcriptional vivo. substrates emission immediately 000 substrate for modification 14 gene to Recently, Qps/mg) Firefly from Dalton produce analogs for wavelength, FP-BS8180 P/N : FP-BE7931 FP-BE7930 P/N : for for Interchim. .com.com luciferase following studying reporter protein, Renilla Renilla Renilla votre source d’innovations for have light. its Assay Buffer, RenillaLuciferase ofassays variable number P/N : FP-BX6710 1000 tests 150 tests Qty 25 µg Qty Qty 1 mg
Assay Kityields ofa Firefly & Renilla Luciferase
■ Firefly and Renilla Luciferases Assay Kit
» Sensitivity and Linearity : Linear correlation between reporter gene expression and light output for transfection using 1 ng to 1 µg DNA of either Firefly or Renilla luciferase reporter construct. » Low Autoluminescence & High Sensitivity : Reduced autoluminescence background for Renilla luciferase assay and consequently increased sensitivity. » Convenient : One kit for both luciferase assays.
Firefly and Renilla luciferases are widely used as reporter genes for studying gene regulation and function, and for pharmaceutical screening. Renilla Luciferase is often used in conjunction with Firefly Luciferase as a normalizing transfection control or for multiplex transcriptional reporter assays. As with many enzymes, Firefly luciferase and Renilla luciferase follow Michaelis-Menten kinetics and thus maximum light output is not achieved until substrates (above the Km) and co-factor are present in large Reporter Gene Assays excess. When assayed under these conditions, light emitted from the reaction is directly proportional to the number of luciferase enzyme molecules. Our Firefly & Renilla luciferase assay kit is designed for detection and quantification of Firefly and Renilla luciferase reporter enzymes from cultured cells in a simple, efficient and linear fashion.
Description P/N : Qty Firefly and Renilla Luciferases Assay Kit FP-BE7810 10 ml FP-BE7811 100 ml Kit contents per 10 ml : 2 x 1 mg D-Luciferin, 100 uL 100X Coelenterazine, 10 mL 5X Passive Lysis Buffer, 10 mL Firefly Luciferase Assay Buffer, 10 mL Renilla Luciferase Assay Buffer, 10 mL Renilla Luciferase Assay Enhancer.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 11 12 Reporter Gene Assays » hard-to-transfect cellsandHTSapplications. coelenterazine. » » lysingthecell samegroupoftransfectedcellswithout theneedfor activitywithout enzyme » » » UptiFectin-On transfectionreagent Anti-Gaussia Luciferase,rabbittiter>1:10000 Related products: pCMV-KDEL-Gluc-1 forintracellularexpression pCMV-KDEL-Basic-1 forintracellularexpression resistance andNeomycinresistance. This positivecontrolvectorisveryusefulinevaluatingtheefficiencyoftransgeneexpressionusingGaussialuciferaseasareporter. pCMV-Gluc-1 positivecontrolwithsecretionsignal analysis andwillexpresssecretedGaussialuciferase. (increase emissionupto45minutes). Kit contains:pre-dissolvedcoelenterazine (100X concentration)andanassaybuffer withstabilizers Gaussia Luciferase Description - The Gaussia The ■ GaussiaLuciferase A pGluc-basic-1 promoter-lesswithsecretionsignal Description Gaussia ■ HumanizedGaussiaLuciferase LuciferaseGaussia - Mixwellandreadinluminometer h humanized The for highthroughputapplications promoterlessvectorwithaMCSsiteupstreamofthehumanizedGaussialuciferasecodingsequence(withsecretionsignal). Add 50µlofGARto20µl Greater brightness:Gaussia pH resistance:survivinga followinga15 :upto60°Candapprox.20%recovery Extremely stabletoelevatedtemperature Avoid celllysis:Gaussia detergents(NP-40, Resistance todetergents:1-5%non-ionic +NP-40 with7Mguanidinechlorideor8Murea Ability torecoveractivityaftertreatment Gaussia
luciferase, Assay Reagent(GAR)ispreparedfreshlybydilutingthecoelenterazinestock withtheassaybuffer. luciferase Gaussia Assay kit a novel assay
luciferase reporter Gaussia luciferase pH rangeof3-11 kit Therefore itcanbeeasilypropagatedin stabilizes Renillareniformis muchas750-foldbrighterthannative isas expressedinmammaliancells luciferase luciferasesamplefrommicrotiterorculturewellsamples for Assay kit is gene sincetimecourseexperiments canbeperformedsamplingthe lysingthecells.Considerabletimeissaved toonlyassaysupernatantsfor media.Itisthereforenecessary intothe withsecretionsignalissecreted secreted the expression, The transfectedcellscanbereusedformultiplesampling. flash into signal the is FP-BY7161 FP-BY7160 P/N :
.com the
.com culture emitted votre source d’innovations smallest Triton X-100, of coelenterazine(10μM)isextremelyhigh:1.24x10 The specificactivityofthisluciferaseinthepresencehighconcentrations light. Ithasanemissionspectralpeakat480nm. oxidative decarboxylationofcoelenterazinetoproducecoelenteramideand Gaussia second permilligram) media E. coliandcanbeusedtoestablishstablecelllinesexpressingGaussialuciferase. by and the and brightest luciferaseusescoelenterazineanditsderivativestocatalysethe Gaussia Triton X-114, CHAPSO),cholate,deoxycholateetc only known
the luciferase 50 ml(1000tests) 5 ml(100tests) Qty media luciferase. thus needs minuteincubationat99°C making Recommended to be it The assayisperformedasfollowing: possible assayed CK5060 CJ3430 BU2560 BU2570 BS8160 BU2550 P/N : This vectorisdesignedforpromoter This vectorhasboth for to by studying 16 use Qps/mg(Quantaper the it as addition weak 0.5 ml 250 µl 25 µg 25 µg 25 µg 25 µg Qty a reporter Ampicillin luciferase promoters, of native gene Coelenterazines
Coelenterazine – coelenterate luciferin – is the substrate for a number of marine bioluminescent enzymes, including those from marine organisms Renilla, Gaussia, Pleuromamma (luciferases) Aequorea (aequorin) and Obelia (obelin). In some of these reactions it is utilized as a simple substrate being catalytically turned over in the bioluminescent reaction catalyzed by luciferases), while in others, such as aequorin or obelin, it is incorporated as part of the photoprotein.
■ Coelenterazine, native
The native coelenterazine, the luminophore of the native aequorin complex, is the standard substrate used in many applications using luciferase reporter assays. Bioluminescent detection of calcium concentration is highly sensitive in a broad concentration range (0.1μM to >100μM) 1-4. Monitoring of reporter genes (phot gene and luc gene) using coelenterazine is also a major application. Other uses of coelenterazine include bioluminescence resonance Reporter Gene Assays energy transfert (BRET)5,10 and chemiluminescent detection of superoxide anion and peroxynitrite in cells or tissues 6-9. Coelenterazine native is recommended when a fast regeneration is important.
References 1) Meth. Cell Biol. 40, 305(1994); This result prompts to prepare FRESH Coelenterazine working solution and then to let it sit for 2) Meth. Enzymmol. 172, 164, (1989); 15-20 minutes at room temperature before use in order to achieve best accurate and high sensitivity. 3) J. BioChem. 105, 473(1989); 4) J. Chem. Soc. Chem. Commun. 21, 1566(1986) 5) Meth. Enzymol. 57, 271(1978); 6) Tetrahedron Lett. 31, 2963(1973); 7) Nature 256, 236(1975); 8) Anal. BioChem. 219, 169(1994); 9) proc. Natl. Acad. Sci. U SA 96, 151(1999); 10) Molecular Pharmacology, 70:1802-1811 (2006)
Description P/N : Qty Coelenterazine, native UP972331 50 µg Highest purity FP-97233B 250 µg UP972333 1 mg UP972334 10 mg
Please contact us for bulk quote at [email protected] Also available : Coelenterazine native for in vivo applications #FP-BV0730.
■ Coelenterazine 400a Protein interactions study in BRET with GFP acceptor
Colenterazine 400a, also known as DeepBlue™ C, is a coelenterazine derivative that serves as a substrate for Renilla luciferase (Rluc) and generates an emission peak centered around 400 nm. It is the preferred Rluc substrate for BRET studies because it has minimal interference with the emission of the GFP acceptor (GFP vectors are presented in the Bioscience Innovation catalog).
Description P/N : Qty Coelenterazine 400a UPBB8391 50 µg FP-BB839B 250 µg UPBB8392 1 mg
See other coelenterazines in the Bioscience Innovation catalog.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 13 14 Reporter Gene Assays Coelenterazine n Coelenterazine ip Coelenterazine i Coelenterazine hcp Coelenterazine h Coelenterazine fcp Coelenterazine f Coelenterazine cp Coelenterazine Native # maximum emission. saturating § * Coelenterazine See completedescriptionsintheBioscienceInnovationcatalog. Table ofLuminescentPropertiesCoelenterazineProductswith ■ Othercoelenterazinesavailable 6-phenyl-OH andposition2oftheimidazopyrazinone core. CycloPentyl directly (YFP) allowsreal-timedetectionofprotein-proteininteractionsinvivo. The Dictyostelium discoideum This Ca BAPTA, Related product: Coelenterazine H Description Coelenterazine For calciumassayinvitroorproteininteractionsstudyBRET ■ CoelenterazineH(Benzyl-Coelenterazine) Coelenterazines Also available:CoelenterazineHforinvivo#FP-BV0680 in only true is max. aequorin, All datasfromBioChem.J.261,913(1989)(otherdatacanbefoundinO.Shimoraura Cell Calcium14,373(1993) Luminescence substituant Ca bioluminescence : 2+ ~ +2
AM chelatorthatdelaysthepeakandincreasesdurationoflightemissionfrom proportional i.e. 466 Ca2+ (CP), a groups protein in nm n vitro. in measured capacity 2-propionyl cells H ; half-time R1, works
that that to resonance R2 is This at FP-39819A FP-R4712A FP-R3080A FP-08353A FP-R3078A FP-R4711A FP-43876A FP-R3079A FP-97233A P/N : the (2P), emits the chemotaxisandinplantwoundhealing. and max have better total total Napthyl cell R3, Ca emission light light of 2+ been in energy with
membrane-permeable, 0.6 concentration. positions (Naph), in emission - transfected wavelength. the Calcium 1.2 transfer methyl Naph I I H H F F OH OH # R1 presence sec.) 2, of (2) 6 aequorin and (Met). activated as (BRET) Half-rise Coelentrazine-H with OH OH OH OH OH OH OH OH OH # R2 compared 8, of Coelenterazine are in (6) calcium, apoaequorin saturating time FP-486104 FP-486103 UPR30784 UPR30783 FP-R3078B UPR30782 P/N : method, very hydrogen
.com.com photoproteins Phe 2P Phe CP Phe CP Phe CP Phe # R3 votre source d’innovations is with to (8) sensitive, the Apoaequorin* from the Ca2+. between e (H), YFP delay has exhibits
native cDNA. apoaequorin hydroxyl acceptor a Intensity 467 441 476 444 464 452 473 442 465 Emission (nm) λ max. elapsed -CH2CHbridge (Aequorin, specific, Coelenterazine. Renilla an In
aequorin (OH), of to this approximate luminescence get luciferase produced intracellular Phenyl system, obelin) 20 x1mg 25 mg 10 mg 1 mg 250 µg 50 µg Qty between 50% 0.26 0.54 0.70 0.67 0.82 0.57 0.80 0.95 1.00 capacité § Relative Luminescence (Phe), of Has the the compared and coelenterazine in in 16-fold been cells. luminophore a variant used The greater to to luminescence of native is measure GFP, is luminescence required useful Coelenterazine; the 0.01 47 0.03 190 10 135 18 20 1.00 Intensity § Relative intracellular yellow for for intensity measuring the intensity fluorescent regeneration appears Ca however 5 1 8 0.15-0.3 0.4-0.8 0.4-0.8 0.4-0.8 0.15-0.3 0.4-0.8 Time(s) § Half-rise 2+ signals in signals (emission changes protein to this be of
D-Luciferins
The popular reporter gene, luc gene, encodes the Firefly luciferase. The ATP-dependent oxidation of the substrate D-luciferin by oxygen produces an emission centered around 562 nm. The light output is proportional to luciferase concentration when both D-luciferin and ATP exist in large excess.
Interchim supplies D-luciferin in various forms : free acid, potassium salt and sodium salt and derivatives with acetoxymethyl (AM), methyl ether and DMNPE. The potassium and sodium salt forms are the most popular because they are readily water-soluble. The potassium salt is also the form used in live animal assay. Interchim’s D-Luciferins are strictly controlled via several chemical analyses and also via a final enzyme assay to ensure consistency.
Description P/N : Qty D-Luciferin, free acid, >99,0% FP-27060A 25 mg FP-27060B 100 mg FP-27060C 250 mg FP-27060D 1 g
D-Luciferin, K+ salt, >99,0% FP-M1224A 25 mg Potassium salt is the recommand salt form for in vivo uses. FP-M1224B 50 mg FP-M1224C 500 mg FP-M1224D 1 g Reporter Gene Assays
D-Luciferin, Na2+ salt, >99,0% FP-726045 10 mg FP-72604A 25 mg FP-72604B 50 mg FP-72604C 1 g
D-Luciferin AM, cell permeant FP-M1909A 5 mg The cell-permeant D-luciferin AM ester enters easily into live cells, and is well retained once it is cleaved by intracellular esterases to D-luciferin.
D-Luciferin ethyl ether FP-CF4421 10 mg Cell permeant analog with 30% higher signal intensity
DMNPE-caged D-Luciferin FP-21639A 5 mg DMNPE-caged D-luciferin is a D-luciferin ester derivative which can cross cell membranes efficiently. Once inside the cells, the ester is continuously hydrolyzed to a supply of D-luciferin. Alternatively a burst of D-luciferin is generated by UV photolysis. References : Luque-Ortega J.R. et al. - In Vivo Monitoring of Intracellular ATP Levels in Leishmania donovani Promastigotes as a Rapid Method To Screen Drugs Targeting Bioenergetic Metabolism, Antimicrobial Agents and Chemotherapy, p. 1121-1125, Vol. 45, No. 4 (2001)
D-Luciferin 6-methyl ether, Na salt FP-M1418A 10 mg D-Luciferin methyl ether has been proposed to be a substrate for microsomal dealkylase/cytochrome P450. Demethylation of the substrate generates D-luciferin, and thus can be detected via bioluminescence with extremely high sensitivity. Reference : Denburg J. et al. Substrate-binding properties of firefly luciferase I. Luciferin-binding site. Archs Biochem. Biophys. 134, 381-394 (1969).
D-Luciferin-6-0-β-D-galactopyranoside FP-CQ6410 5 mg β-Galactosidase substrate Reference : Yang,Y. et al., Homogeneous enzyme immunoassay modified for application to luminescence-based biosensors, Anal. Biochem 33: 102-107 (2005)
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 15 16 Reporter Gene Assays development. lacZ X-GAL X-GLU (5-Bromo-4-Chloro-3-Indolyl-B-D-Glucopyranoside,MW:408.6) Fluo.Std. #524739,10mg #95432A,10mg Subst. #52476A,10mg Fluo.Std. Resorufin Subst. Res-Gal #193659,10mg Fluo.Std. Fluorescein Subst. FDG Contains : β-Galactosidase substratessamplerKit o-NPG (o-Nitrophenyl-β-D-galactopyranoside,MW:301.3;λ Description ■ Othercolorimetricsubstratesforglycosidases Description This ■ β-GalactosidasefluorescentsubstratessamplerKit kit β-Galactosidase (5-Bromo-4-Chloro-3-Indolyl-β-D-Galactopyranoside,MW:408.6) FUGl #M11419, 10mg TFMU-Gal systems Substrates for glucosidases reporter consists FU #434769,10mg TFMU of samples activity -galactosidase, β-glucuronidase, β-glucosidase) (β-galactosidase, of at several a variety of our of wavelengths. most popular abs (cleaved) :420nm) This galactosidase FP-BM8400 .com.com P/N : kit votre source d’innovations is perfect substrates for those UP40534M UP193325 UP556683 P/N : and occasions 1 kit Qty their reference where 1 g 100 mg 5 g Qty inquire. other derivatives(inhibitors,activators,...).Please Each substrateisavailableseparately, andalsomany the fluorophores preferred wavelength allowing multiplexed of detection analysis is under of Alkaline phosphatase assays
Phosphatases are a specific class of enzymes that catalyze the removal of phosphate groups from proteins, whereas kinases enzymatically add a phosphate to a protein. Phosphatases are important in signal transduction because they regulate the proteins they are attached to. To reverse the regulatory effect, the phosphate has to be removed. Cell proliferation and differentiation are regulated by phosphatases.
Phosphatases serve also as enzyme markers, allowing to quantify phosphatase activity in different types of cells. Alkaline phosphatase is finally a highly sensitive enzyme for ELISA, immuno-histochemical, Northern, Southern and Western blot applications.
■ Fluorimetric Assay kits
The Alkaline Phosphatase Assay Kits use fluorogenic phosphatase substrate, to quantify alkaline phosphatase activity in solutions, in cell extracts, in live cells as well as on solid surfaces (such as PVDF membranes). The kits provide all the essential components with our optimized "mix and read" assay protocol that is compatible with HTS liquid handling instruments.
■ Alkaline Phosphatase Assay Kit, Alkaline phosphatase dose response on 96-well Blue Fluorescence black plate with 30 minutes incubation time (n=3).
Substrate : MUP Enzymes Detection
λex./em. : 360 / 449 nm Sensitivity : 0.3 pg of alkaline phosphatase
Description P/N : Qty Fluorimetric Alkaline Phosphatase JQ6730 500 tests Assay Kit, Blue fluorescence
■ Alkaline Phosphatase Assay Kit, Alkaline phosphatase dose response on 96-well Green Fluorescence black plate with 30 minutes incubation time (n=3).
Substrate : FDP.
λex./em. : 490 / 514 nm Sensitivity : 0.1 pg of alkaline phosphatase
Description P/N : Qty Fluorimetric Alkaline Phosphatase JQ6740 500 tests Assay Kit, Green fluorescence
■ Alkaline Phosphatase Assay Kit, Alkaline phosphatase dose response on 96-well black plate with Red Fluorescence 30 minutes incubation time (n=3).
Substrate : Phospholite™ Red
λex./em. : 570 / 590 nm Sensitivity : 0.2 pg of alkaline phosphatase Description P/N : Qty Fluorimetric Alkaline Phosphatase JQ6750 500 tests Assay Kit, Red fluorescence
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 17 18 Enzymes Detection phosphatases this phosphatase λ Substrate :proprietaryluminescentsubstrate ■ Luminometric Luminometric phosphatase assaysAlkaline λ MUP Description Although ■ MUP Acide phosphatase assays See Colorimetric Luminometric Description the MW :300g.mol activity This Sensitivity : solid continuous assays.Itcanalsobeusedfortheassaysthatrequireacidic pHsuchasacidphosphatases. phosphatase. that phosphatase ex./em. em. :560nm AP enzymatic pH Plus™,sodiumsalt :360/450nm produces Alkaline surfaces colorimetricassayssubstratesandkitsinImmunoassayreagentssection. in limitation MUP solutions, Plus 0.01 pgofalkalinephosphatase Alkaline Phosphatase -1 substrate, substrate Phosphatase
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exhibits with in fluorescence on acid JQ6760 P/N : FP-JQ6710 P/N : .com.com solution phosphatases. votre source d’innovations maximum Assay Kit it is at not pH well fluorescence value FluoProbes suited of for >10. 200 assays Qty Alkaline phosphatasedoseresponseon96-wellblackplate 25 mg Qty above living is Thus pleased cell pH with 30minutesincubationtime(n=3). it or 7.0, is continuous to also thus offer difficult MUP MUP assays
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■ Protease Assays Kits, Green & Red Fluorescence
» Optimized Performance : Optimal conditions for the detection of generic protease activity » High Speed : Minimal hands-on time » Assured Reliability : Detailed protocol and references are provided
The Protease Assay Kits are widely used for detection of generic protease activities. The kits use a casein derivative that is heavily labeled with green or red fluorescence, resulting in almost total quenching of the conjugate's fluorescence. Protease-catalyzed hydrolysis relieves this quenching conjugate, yielding brightly fluorescent dye-labeled peptides. The increase in fluorescence intensity is directly proportional to protease activity. The protease assay kits do not require any separation steps and can be used to continuously measure the kinetics of a variety of exopeptidases and endopeptidases.
The kits contains : • Fluorescent labeled casein with high ratio of dye/protein (pH-insensitive) • Trypsin (as positive control) • Assay buffer • A detailed protocol Enzymes Detection
Description P/N : Qty Protease Assay Kit, Green Fluorescence (488/520 nm) BK962A 500 tests* Protease Assay Kit, Red Fluorescence (546/575 nm) BK963A 500 tests*
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 19 20 Enzymes Detection Binding fluorescence The ratio isabouttwicethatfor ADIFAB unbound freefattyacids. the while triglycerides, of groups ester primary the for reagents, used ADIFAB2 ADIFAB Description The ■ Unboundfreefattyacids Fluorescent Lipase Description Kouker G.andJaegerK.E.,"Specificsensitiveplateassayforbacteriallipases". Howard G.T., etal."Sensitiveplateassayforscreeninganddetectionofbacterialpolyurethanaseactivity". References : substrate from manner. Lipases Fast andeasymeasurementoflipaseactivityinvitro,cellpreparationsorvivo ■ Lipase Lipase assays andLipidstudy (Product fluorescent contained numerous assaysandcontrolexperiments,alsocontainsreferencestandardsadetailedprotocolforuse. obtained released and activity measurements are easily fatty nonaqueous in system Leu72 FFAu affect cellularprocesses aswellthoseinvolvingpurifiedenzymes. ADIFAB (FFAu) levels. (Product 1,2-dioleoyl-3-pyrenyldecanoyl-rac-glycerol monitored using activity, ADIFAB2 the behaviorofenzymesthateitherproduceoruseFFA. of vivo. lipase FFA assay fluorescence acids either to some probes to can
The affinities assay ubiquitous levels. and are has # Ala Most either activity, # is is fluorescence from can FP-37853A, in typically the FP-M14031, be a lipases fatty a kit ADIFAB2 phase a mutant emission high have unbound family used the wider be lipases triacylglycerols Assay Kit C-1 For in contains the for Assay Kit fatty of vitro, used acid affinity among
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at labeling plates. other or l 53(1):211-3 (1987) (72 drugs ADIFAB2 fatty 375 emission that assays can hand, nm). alter This acid that the be of of of in 96 well 32(3):211-4 (2001) Copyright FFA ADIFAB micro-injectedintofatcells– Intracellular FFAu levelsmeasured with plate) biosciences Collagen and elastin study
■ Lysyl Oxidase Assay Kit, Red Fluorescence
Sensitivity : 40 ng of lysyl oxidase in solution
Lysyl oxidase is an extracellular enzyme that catalyzes formation of aldehydes from lysine residues in collagen and elastin precursors. These aldehydes are highly reactive, and undergo spontaneous chemical reactions with other lysyl oxidase-derived aldehyde residues, or with unmodified lysine residues. This results in cross- linking collagen and elastin which is essential for stabilization of collagen fibrils and for the integrity and elasticity of mature elastin. Lysyl oxidase has been identified as a possible tumor suppressor. Lysyl oxidase activity in biological samples is traditionally and most reliably assessed by tritium release end-point assays using radiolabeled collagen or elastin substrates involving laborious vacuum distillation of the released tritiated water. This kit offers a sensitive fluorescent assay for lysyl oxidase activity that utilizes Enzymes Detection 1,5-diaminopentane as substrate, and released hydrogen peroxide is detected using our HRP substrate in HRP-coupled reactions. This method allows the detection of sub ng/mL lysyl oxidase and is Lysyl oxidase dose response on 96-well black plate with 30 minutes incubation time (n=3). The insert shows the low levels of lysyl oxidase detection. much more sensitive than the currently available fluorimetric assay for this enzyme activity. This method eliminates the interference that occurs in some biological samples and can be readily used to detect lysyl oxidase activity in cell culture experiments. Description P/N : Qty Lysyl Oxidase Assay Kit, red fluorescence JQ7270 500 assays Cellulase Detection ■ Cellulase Assay Kit, Red Fluorescence
Substrate : Resorufin Cellobioside Reaction volume : 100 µl
λex./em. : 571/585 nm
Cellulases are a family of enzymes that include ß-Glucosidases, endoglucanases, and exoglucanases. These enzymes cleave the ß-1,4-D-glycosidic bonds that link the glucose units comprising cellulose. In addition to being produced by plants, cellulase activity is found in many fungi and bacteria, including some plant pathogens. Most animal cells are not known to produce cellulase; cellulolytic activity is often carried out in animals by symbionts. However, recent evidence does suggest cellulase production in some animals, such as insects and arthopods. The study of cellulase activity has many applications in plant Suspension of flowering buds from two mature Arabidopsis thaliana plants in triplicate (50 μL) on a 96-well clear, flat bottom plate read at 3-minute intervals for 120 minutes. molecular biology, agriculture, and manufacturing. Cellulase is also becoming important in the development of alternative fuel sources, as glucose obtained from cellulose hydrolysis is easily fermented into ethanol. Activity of most cellulases can be monitored using our long wavelength fluorescent substrate, Resorufin Cellobioside, contained in the kit. Upon cleavage, the fluorescent compound, Resorufin, is released and activity measurements are easily obtained in a microtiter plate based assay format. Description P/N : Qty Fluorescent Cellulase Assay Kit DO8110 200 assays Kit contains : Substrate Reagent, Reference Standard, Reaction Buffer, Stop Buffer, DMSO votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 21 22 Enzymes Detection » Bioluminescence assayforMonitoring ■ aCella– » » » transmission. Acetylcholinesterase junction, of a inhibition both as enzymatic current assaymethods. Acethylcholinesterase Detection Control tomeasuremaximumluminescence Kit Contents: aCella –AchE Description Reaction III: Reaction II:Coupledenzymereaction+ Reaction I: Assay Principle field increasing Alzheimer’s FAST Versatile monitoring;drug :Nervegas,pesticide Ultra Sensitiveassaytomonitor Homogenous :One-step,nowashassay environmental of 30seconds-5minutes :Resultsin intensive of endoplasmic activity AChE Acetylcholinesterase, Detectionreagent,acetylcholineandcoupledenzymereaction, Assay AChE +Inhibitor interest The ATP treatments Acetylcholinesterase by (remaining)+Luciferase/Luciferin as enzyme scientific detection both (AChE) drugs in reticulum, various nerve (e.g., is investigation. have of bound is AChE gases one disease tacrine been etc). AChE activity to of AcetylCholinEsterase inhibitors and cellular Acute hampered the (tetrahydroaminoacridine)) ATP processes AChE an most important toxicity membranes and inhibitors important by screeningapplications development and Assay the to No class treatment humans difficulty Acetate andCholine. have CA6652 CA6651 CA6650 .com.com P/N : of Activity enzymes of votre source d’innovations excitable also pesticides of and and strategies. and modulators been Reaction doesnotproceed. involved complexity animals tissue are used has LIGHT the
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Calcium Assays
Intracellular Ca++ levels have become important indicators for the activation state of ion channels and G-protein coupled receptors as well as for the phases of apoptosis and cell injury. Though the respective kinetics and the absolute amounts of the Calcium levels are different for each of these physiological processes there are common ways for monitoring them. Luminescent labels like Aequorin as well as fluorescent ones are versatile and widely used solutions for microplate assays. Fura 2 and Indo-1 provide ratiometric readout thereby reducing effects caused by leaking or bleached dyes or varying assay conditions.
Product name λ \λ max. (a) λ \λ max. (a) MW (g·mol-1) exc em exc em Kd Ca2+ (nM) Applications Free Ca2+(nm) High Ca2+ (nm) Fluo-3 AM(b) 1129 503 / weak 505 / 526 390 . Most classical calcium indicator . No Wash step needed Fluo-8 AM 1000 490 / weak 490 / 514 389 . 4 times brighter than Fluo-3 . Loading at room temperature Fluo-8H AM 1100 490 / weak 490 / 514 232 . High Ca2+ concentration indicator Fluo-8L AM 1100 490 / weak 490 / 514 1 860 . Low Ca2+ concentration indicator . Red calcium indicator Rhod-4 AM 530 / weak 530 / 555 525 . 4 times brighter and 10 times larger windows assay than Rhod-2 . Loading at room temperature
. Leakage resistant form of Fura-2 Calcium Assays
Fura-PE3 AM 1258 335 / 495 380 / 495 250 . Ratio of reads with 2 different λex. . Avoids interference due to dye distribution and photobleaching . Leakage resistant form of Indo-1
Indo-PE3 AM 1266 338 / 480 338 / 410 260 . Ratio of reads with 2 different λem. . Avoids interference due to dye distribution and photobleaching
(a) after hydrolysis (b) AM ester are membrane-permeant and thus increases greatly cell loading that can be performed by simple incubation of the cells or tissue preparation in a buffer containing the AM ester. Pluronic® F-127, a mild non-ionic detergent, can facilitate AM esters loading. The AM esters themselves do not bind to Ca2+. However, once they have entered the cells, they are rapidly hydrolyzed by intracellular esterases into the parent Ca2+ compounds, thus becoming fully fluorescent upon binding to Ca2+. Many other ion indicators are available in our Interchim’ range of dye. Please contact us for specific application needs.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 23 24 Calcium Assays The nextgenerationcalciumindicatorforautomatedscreening(HTS)applications ■ Fluo-8 Ionomycin, Ca λ Fluo-3 Description Ionomycin, Ca Probenecid, watersoluble Probenecid, Cellculturetested,tosuppress effluxofdyes Related products: Description » » Standard Ca ■ Fluo-3 (490/514 The » » » » » Kd =390nm λ Fluo-8 Fluo-8 NoWash Calcium Fluo-8 NoWash Calcium complex withcalcium)makeFluo-8NWanidealindicatorformeasurement ofcellularcalcium. which dye groups When Single λ exc. exc. /λ /λ Rapid dyeloading :at Increased signalintensity Performed in96or384-wellmicrotiter-plate Convenient androbust:Nowashstepneeded. Appropriate apparentCa Low compartmentalization Large dynamicrange Fluo-3 Fluo-8 that em. em. AM AM greatly =505/523nm :(Hydr.,Ca cells of stays nm), Fluo-8 NW stimulated 2+ 2+ 2+ increase ionophore ionophore concentrationindicator AM -NoWash AM inside high (No 2+ exc./em. NW ) :505/523nm;Kd(Ca Wash) sensitivity, cells are the Assay Kit,1%FBSMedium Assay Kit,Mediumremoval with fluorescence cleaved can and agonists, 2+
binding affinity cross and its Ca by fluorescence >100 cell the esterases, 2+ 2+ FP-78932A P/N : FP-78932D FP-M2036A FP-78932C FP-78932B FP-R1245A RT of ) =390nM;MW:1000 membrane. receptor Fluo-8 indicators (ratherthan37°CrequiredforFluo-4 times 1 2 is NW. fluorescence resulting signals greatly Fluo-4 Once The characteristics FP-53989B FP-53989A FP-288653 FP-288652 CP7501 CJ2551 CJ2550 CJ2561 CJ2560 P/N : enhanced in inside .com.com the a 1 mg Qty 50 mg 1 ml(1mMsolutioninDMSO) 20 x50µg 10 x100µg 1 mgFluoProbesPureGrade votre source d’innovations enhancement release negatively the cell, upon of of the intracellular charged binding its lipophilic (when long AM) wavelength fluorescent to 5 mg 1 mg 10 x150mg 10 x150mg 5 x50µg 100 plates 10 plates 100 plates 10 plates Qty it blocking calcium, calcium. forms a
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has binding. 2 1 at Ca : : fluorescence ATP with Alternatively, It to 488 2+ an is
indicator, doseresponsesinHEK-293cells minimize 0.5-to Fluo-3 important absorption nm by 1% proves one even intensity argon-ion background FBS to can spectrum remove if to to it grow be avoid increase is laser more the the fluorescence, the 2+ compatible medium sources,
growth generally indicators. cells susceptible in response in removal medium growth and most with and to a Calcium Assays 25 ] is 2+ [Ca standard : indicator measurement the cytosolic to by of detected indicators suited 2+ hardly studies introduced better were Ca are kinetic i.e.), for buffering linearity and (fibroblast exc./em. additional advantages range cells the has and dynamic 1 mg Qty 10 x 50 µg 20 x 50 µg 1 mg Qty 10 x 50 µg excitable in Single λ Single indicators
release, Their 2+ and or Ca influx of indicators. affinity
vacuoles) 2+ low Ca sites . FP-CP7550 P/N : FP-CP7551 FP-35374C FP-CP7530 P/N : FP-CP7531 with 2+ near affinity exist (mitochondria, high that used concentration organelles ion
changes 2+ )=232 nM ; MW : 1100 )=1.86 µM 2+ 2+ commonly Ca some in free more with concentration present Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com cytosolic : 369, 329/510 nm) em. of concentration indicator concentration concentration indicator /λ ): 490/514 nm ; Kd(Ca ): 490/514 nm ; Kd(Ca AM 2+ transient AM 2+ 2+ 2+ compared
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: (Hydr.,Ca : (Hydr.,Ca Ca measurement em. em. /λ /λ high-localised exc. exc. minimised. too high affinity for Ca Fluo-4 and Rhod-2 have dyes Fluo-3, transients High Related products Mag-Fura-2 Description ■ Fluo-8L Low cytosolic Ca Description Fluo-8H λ ■ Fluo-8H Ca High cytosolic of The Fluo-8L λ votre source d’innovations 26 Calcium Assays λ Rhod-4™ Rhod-4™ NWCalcium AM fluorescent Rhod-4™ NWCalcium Rhod-2. while to cellular calcium responses. small very has and hydrolysis, esterase Rhod-2 The brighestredfluorescentcalciumindicator ■ Rhod-4™ Single λ Description Rhod-4™ Cells are with and Rhod-4. more sensitive thanRhod-2 times 10 is that response calcium cellular mobilization, fluorescence-based (GPCRs) receptors G-protein-coupled monitoring for method assay optimized convenient of membrane. of stimulated fluorescence Rhod-4™ screening. The adapted toautomation. negatively exc. cellular intracellular improve /λ is pre-loaded em. >250 calcium) Rhod-4 expressing only :(Hydr.,Ca maintaining is In The AM
and calcium. are moderately times is with charged most CHO Once 96-well calcium cell and is characteristics the NW make calcium cleaved calcium, greatly with screening 2+ fluorescence and exc./em. commonly a brightest a n clim response calcium and loading ) :530/555nm;Kd(Ca inside preferred Calcium This Rhod-4™ AM - our GPCR fluorescent the or Assay Kit,1%FBSGrowthMedium Assay Kit,MediumRemoval indicators. Rhod-4™ assay HEK AM. fluorescent enhanced channels. by 384-well proprietary which Rhod-4 the spectral compounds, non-specific cells red of for No Wash (NW) used of method Assay cell, has interest increases greatly its an calcium Ca Rhod-4™ dye However, detecting NW upon microtiter-plate The in been the long among wavelength ideal Rhod-4 live that in rvds homogeneous a provides Kit Calcium 2+ assay binding 2+ lipophilic increase that drug
) =525nm the wavelength, developed cell cells indicator indicator (when stays indicators AM the Rhod-2 the NW receptor signals esterase, discovery can upon Assay to has red intracellular which inside the of calcium. it blocking format be available for forms through fluorescence high signals Kit performed incubated well Figure were washedwith2times200µlHHBS,thenimagedunderfluorescentmicroscopeusing measurement can for resulting cells, provides When and complexes screening. sensitivity, in CQ6092 CQ6091 CQ6090 CQ6061 CQ6082 CQ6081 CQ6080 P/N : CQ6064 CQ6063 CQ6062 groups .com.com cross 2. for a calcium calcium release and 96 Rhod-4 votre source d’innovations easily with cells HTS in in wells cell an its of of 100 a a AM black µl vs of 100 cium Figure or removed, added EC
5uM Rhod-2 wall/clear 5 59 µl ofrhod-4NWisabout0.8uM. µM Assay Rhod-4 per by 1. AM Rhod-2 Carbachol and NOVOstar well bottom kit in AM 100 plates 10 plates 1 plate 5 x50µg 100 plates 10 plates 1 plate Qty 1 mg 20 x50µg 10 x50µg the and U2OS. in or AM a cells costar Rhod-2 Rhod-2 96-well Dose at (BMG U2OS were 37°C, plate. Response AM. AM black LabTech) cells incubated 5% in HEK-293 The wall/clear HHBS were CO growth in 2 to
seeded with incubator at HEK-293 achieve cells 37°C, bottom medium 100 were overnight µl 5% the for cells costar of seeded CO was 1 final the hour. measured 2
at incubator plate. removed, Rhod-4 indicated 40 overnight Carbachol 000 The NW with for cells and growth concentrations. at calcium 1 Tritc channel. Rhod-4 (25 hour. 40 per the 000 medium µl/well) 100 cells The assay NW cells µl were cells per Cal- was was The per kit, Ratiometric calcium dyes
■ Fura-PE3 AM & Fura-2 AM Leakage resistant form of Fura-2, a popular Setrometric Ca2+ indicator
Fura-2 AM may be useful in microplate studies, where cell lines with different properties are compared or where screening treatments lead to differences in the number of cells or dye loading. Some of the limitations in the use of Fura-2 appear to be overcome by the use of glass bottom microplates (See page 78).
Reference : Robinson JA et al., Ratiometric and non-ratiometric Ca2+ indicators for the assessment of intracellular free Ca2+ in a breast cancer cell line using a fluorescence microplate reader, J Biochem Biophys Methods. 2004 Mar 31;58(3):227-37.
Fura-PE3 is an improved version of Fura-2, that reduces cell leakage and thus increases dye loading accuracy. Description P/N : Qty Fura-2 AM FP-42776A 1 mg FP-42776C 20 x 50 µg Representative experiment of angiotensin II FP-85312A 1 ml (1 mM) (ANG II)-evoked changes in the ratio of fura 2 fluoresence (340/380 nm) in adherent neonatal rat cardiomyocytes Fura-PE3 AM FP-AM603A 500 µg
(NRC). Primary cultures of NRC loaded with Fura 2 Calcium Assays were stimulated with increasing concentrations of ANG II (10 pM-1 µM) either in the absence (control ; A) or ■ Indo-1 AM presence of AA-861 (10 µM ; B) or MK-571 (100 nM ; C).
Indo-1 AM has a shift in the emission from 485 nm to 405 nm in the presence of calcium. Indo-1 AM can be used with a single argon-ion laser for excitation and to monitor two different emissions.
Description P/N : Qty Indo-1 AM FP-427755 500 mg Related product FP-42775A 20 x 50 µg Description P/N : Qty FP-98180A 1 ml (1 mM) Calcium Calibration kit FP-21527A 1 kit ® Related products : Pluronic F-127 FP-37361A 1 g ® 4-bromo A-23187, Ca2+ ionophore for UV light-excited dyes FP-372221 1 mg Pluronic F-127, 20% FP-69806A 1 ml A-23187 (calcimycin or Calcium Ionophore III) solution (in DMSO) FP-28362B 5 mg to equilibrate intracellular and extracellular [Ca2+]
Representative tracings from indo 1-loaded myocytes show simultaneous Ca2+ transients (top panels) and cell length (bottom panels). Myocytes were stimulated at 0.5 Hz after 6 h of no treatment (control) (A), LPS (10 ng/ml) (B), or LPS with ANG II (100 nM) (C).
■ Indo-PE3 AM Leakage resistant form of Indo-1
Description P/N : Qty Indo-PE3 AM FP-AM602A 500 µg
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 27 28 Viability & Cytotoxicity Assays or application At Several » » » » » Cell countingisrequired methods forgeneraltospecificcellassays. Interchim It maybeusefultoquantitateonlyviablecells,orfastproliferatingcells. Cell counting, &proliferation viability 51 Probe Selection guide Propidium Trypan blue Iodide, MTT Hoechst XTT UptiBlue WST Calcein-AM GAPDH CFSE Luciferin Syst. LDH AnnexinV BRDU -3H Thymidine
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+++ Proliferating - ++ +++ +++ +++ +++ +++ ++ + ++ + +++ + +++ +++ study. *Mithras LB940MultiModeReader *NightOWL and kitswerevalidatedwithinstruments. Interchim MicroPlate readers&Imagingsystems works. Technical tip Annexin V-FP488 todiscriminatedeadcellsfromalivecells. Cons :hazardous(radioelements). Recommended forcytotoxicityassays. Features/Advantages -Drawbacks Used incombinaison ofgreenfluorescencedyelike. Do notstateonviability. Cheap, buttimeconsuming,notscalable. toXTT NontoxicIncreasedsolubilityandperformancefromMTT Popularmethod.Sensitive,Scalable. More rapidthanMTT/XTT Cheap, Scalable,Nontoxic.Donotstateonviability. easier touseFluorimetry/SuperiorsensitivityMTT adherent cells.SensitivitysimilartoMTT/XTT, but No solubilizationstep(unlikeMTT). No solubilizationstep(unlike May altersomecellfunctions. in vivocelltracing.Donotworkforbacteria. wide varietyoftechniques,including:microplateassays, Complement-Mediated Cytolsis. Measurement Do notstateonviability. Useful whenothermethoddonotworkproperly. Recommended forcytotoxicityassaysSerumInterference. Cons :signaldependsoneachcellline, on temperature Pros :sensitivity/linearity. Useful for Cons :hazardous(radioelements).
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■ Live/Dead Mammalian Viability Assay Kit Two-color fluorescent staining of live (green) and dead cells (red)
» Dual Detection : Detect both live and dead cells simultaneously. » Simple & Fast : Require only a 30-min dye loading time and then measure without washing. » Economical : Perform viability and cytotoxicity assays at the same time. » Versatile : Analyze with flow cytometers, fluorescence microscopes or fluorescence plate readers.
The Viability/Cytotoxicity Assay Kit for Live / Dead Cells provides a two-color fluorescence staining on both live and dead cells using two probes that measure two recognized parameters of cell viability — intracellular esterase activity and plasma membrane integrity [Papadopoulos, 1994]. The kit is suitable for use with fluorescence microscopes, fluorescence multiwell plate scanners and flow cytometers and other fluorescence detection systems. The assay principles are general and applicable to most eukaryotic cell types, including adherent cells [Vaughan, 1995] and certain tissues [Poole, 1993], but not to bacteria or yeast. This fluorescence-based method of assessing cell viability can be used in place of trypan blue exclusion, Viability & Cytotoxicity Assays 51Cr release and similar methods for determining cell viability and cytotoxicity. It is generally faster, less expensive, safer and a more sensitive indicator of cytotoxic events than alternative methods. EthD-III shares the same property with EthD-I used in Live/Dead Viability/Cytotoxicity Assay Kit #486301 and is 40% brighter at intensity compared to EthD-I. Validity of the Live/Dead Viability/Cytotoxicity assay for animal cell applications has been established by several publications.
Live cells are distinguished by the presence of ubiquitous intracellular esterase activity, determined by the enzymatic conversion of the virtually nonfluorescent cell-permeant calcein AM to the intensely fluorescent calcein. The polyanionic dye calcein is well retained within live cells, producing an intense uniform green fluorescence in live cells (ex/em ~495 nm/~515 nm). EthD-III enters cells with damaged membranes and undergoes a 40-fold enhancement of fluorescence upon binding to nucleic acids, thereby producing a bright red fluorescence in dead cells (ex/em ~495 nm/~635 nm). EthD-III is excluded by the intact plasma membrane of live cells. The determination of cell viability depends on these physical and biochemical properties of cells. Cytotoxic events that do not affect these cell properties may not be accurately assessed using this method. Background fluorescence levels are inherently low with this assay technique because the dyes are virtually nonfluorescent before interacting with cells.
If cells are first fixed, and then stained, the Live/Dead Bacterial Viability/Cytotoxicity kit can also be considered. To replace the dye Calcein AM that will only stain the live cells, the DMAO; a DNA-binding dye, will stain both intact and damaged cell membranes.
References : J Immunol Methods, 177, 101 (1994). J Cell Sci, 106, 685 (1993). J Neurosci, 15, 5389 (1995). Description P/N : Qty Live/Dead Mammalian Viability Assay Kit FP-BF4710 1000 tests in microplate reader Related products : DMAO, nuclei stain for live cells, 2 mM soln in DMSO FP-CA8150 1 ml Ethidium Bromide III, 1 mM solution FP-BP9341 200 µl
MTT (λabs.(cleaved) : 650 nm (550-600 nm)) FP-65939A 1 g Live/Dead Yeast Viability Assay Kit 486301 1 kit based on calcein-AM and PI. Hela Cells incubated with assay solution.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 29 30 Viability & Cytotoxicity Assays may may yields A and applicabletomostbacteriatypes. appropriate This kitcanalsobeconsideredifcellslikemammaliancells,arefirstfixed,andthenstained. difference ofliveanddead bacteriacountsisobservedbetweenthisassayandgrowthassays. membranes (red) and Viability/Cytotoxicity popularsettings using fluorescencemicroscopes withflowcytometersand » » andmeasurewithoutwashing. » inacell deadbacteriacells » Two colorfluorescencestainingonbothlivebacteria(green)anddead(red) ■ Live/DeadBacterialViability/Cytotoxicity kit cells live stainingtoDual detect anddead based onWST-8 formazandye.Readat450nm(450-490nm) Live/Dead Related products: Live/Dead BacterialViability/Cytotoxicity Description
common and propidiumiodide. Versatile : time. atthesame assays cytotoxicity Economic :Performviabilityand Simple &Fast:15mindyeloading population simultaneously. Dual Detection:Detectliveand damaged be be bacteria results unable able Yeast Viability criterion mixture is to that Analysis compatiable using stained cell to recover correlate reproduce membranes. for for fluorescein two of Assay fluorescent Assay Kit bacterial DMAO and probes, well Kit reproduce in and for nutrient with viability EtD-III DMAO red. Bacteria EtD-III, growth The
is medium, — is and a the such kit bacteria assays red-fluorescent Live EtD-III. is ability suitable bacteria and & in Dead of DMAO with yet liquid 486301 FP-BU1040 P/N : a for bacterium these may intact Cell use is or .com.com nucleic a be solid Staining votre source d’innovations may with green-fluorescent cell scored to media. acid fluorescence membranes be 1 Kit 1000testsinmicroplatereader Qty reproduce scored Kit dye as Under provides "dead" that as in is microscopes stains certain "alive". suitable nucleic stained in two-color this only conditions, Therefore, acid assay. nutrient fluorescent dead dye and fluorescence Conversely, media bacteria flow that however, these stains cytometers. green, that situations with staining is both some bacteria whereas referred damaged live The bacteria need on having and assay to bacteria both as to cell dead with be growth live damaged principles membranes. considered bacteria with intact (green) assays. damaged membranes membranes are with and if With general This a intact dead vast cell an kit Dual staining to detect live and dead cells
■ Ethidium monoazide, bromide (EMA) Selectively and covalently labels membrane-damaged or metabolically compromised cells in the presence of live cells
Ethidium monoazide bromide is a red fluorescent nucleic acid stain with a photoaffinity label. The dye, after photolysis, binds covalently to nucleic acids.1
After photocrosslinking to DNA, the wavelengths (λexc. / λem. = 504 / 600 nm) are compatible with a simultaneous observation of another green indicator. The dye has been used to "footprint" drug binding sites on DNA2 to modify plasmid DNA,3,4 and to determine hemopoietic cell phenotype, function and position in the cell cycle.5 A particularly useful application of the dye is to selectively and covalently label dead cells in the presence of live cells. Since ethidium monoazide bromide is relatively impermeant to live cells, it selectively labels DNA in dead cells in a Viability & Cytotoxicity Assays mixed population of live and dead cells. Photolysis following the dye application renders the dead cell DNA covalently labeled with the dye. One can then wash and fix the cell preparation and exam it by microscopy fluorescence plate reader or flow cytometry. The major advantage of this method is that researchers can avoid extensive manipulation of live pathogenic organisms.6 At the difference of propidium iodide, the ethidium monoazide binds covalently, and, when applied to cells before fixation, provides an indication of what fraction of the unfixed population Two photon fluorescent image of live PTK2 cells vitally stained with EMA : were membrane-damaged or metabolically compromised. A late prometaphase cell illustrating the high selectivity of the stain for the chromosomes. References : 1)J. Mol. Biol. 92, 319(1975) 2) Euro. J. Biochem. 182, 437(1989) 3)J. Biol. Chem. 257, 13205(1982) 4) J. Biol. Chem. 259, 11090(1984) 5) Cytometry 11, 610(1990) 6)Cytometry, 12, 133(1991) 7) PNAS, 97 , no. 17, p. 9504-9507 (2000)
Description P/N : Qty Ethidium monoazide, bromide (EMA) FP-48256A 5 mg
λex./λem.(DNA bound) : 504/600 nm ; MW : 420.3
■ Propidium monoazide (PMA) Selectively and covalently labels dead cells in the presence of live cells
PMATM is a derivative of EMA, but it has significantly higher DNA binding affinity and is cell impermeant. As EMA, after photolysis, the dye is converted to a fluorescent DNA stain covalently bound to DNA.
Description P/N : Qty Propidium monoazide (PMA) FP-BZ9340 1 mg
λex./λem.(DNA bound) : 510/610 nm ; MW : 512
References : Nocker, A. et al., Comparison of propidium monoazide with ethidium monoazide for differentiation of live vs. dead bacteria by selective removal of DNA from dead cells. J. Microbio Meth. 67(2), 310-320 (2006). Nocker A. et al., Use of Propidium Monoazide for Live/Dead Distinction in Microbial Ecology, Applied and Environmental Microbiology, p. 5111-5117, Vol. 73, No. 16 (2007). Pan Y., Breidt F., Enumeration of Listeria monocytogenes by Real-Time PCR with Propidium Monoazide and Ethidium Monoazide in the Presence of Dead Cells, Appl. Environ. Microbiol. doi:10.1128/AEM.01198-07 (2007).
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 31 32 Viability & Cytotoxicity Assays » » » Features : adding theCalcein-AMassaysolution. and » » Applications : » » » 000 proportional fluorometric The ■ Calcein Live cell staining Description Annexin V-FluoProbes 488,FlowCytometry Grade(495/519mm) Related products: Calcein Calcein Calcein Cell CountingKit,calcein-AMbased Propidium iodide,1mg/ml λ Calcein ex. by It integrity andpermeability, angiography, liposomes… reproductible screeningassays. reduces The endocytosis, cells, other and neurons in changes volume cell including made it a popular and versatile dye for various applications, emission membranes. exhibits M ester AM should thusbemonitored. physiological respectively. of Calcein Technical tip /λ Non-radioactive cells Ideal forbothsuspensionandadherent cells Suitable forproliferatingandnon-proliferating immunocytochemistry,Microplate assays, flowcytometry, andinvivocelltracing Cell adhesion,chemotaxis,multidrug fluorescein) tootherfluorescentcompounds (i.e. Better retentionandbrightnesscompared Ideal forhigh-throughputassays stepasinan Rapid (nosolubilization em. phenol cells is Calcein-AM pH several (cleaved) :494/517nm;MW994.9 worthy DMSO AM, 5mMinanhydrousDMSO,PureGrade AM, 4mg/mlinanhydrousDMSO AM, 1mg/mlinanhydrousDMSO AM between per dye fluorescence wavelengths solubilization red is determination to ions, well. It to Intracellular membrane-permeant solution is pH gap the is in notice a AM CellCounting&Viability Kit well the (not including 6.5 It number polyanionic junctional microplate can provides retained culture
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.com.com The method of votre source d’innovations 5 96-well a µl fluorescent Assay Kit FP-895515 FP-JQ8140 FP-FI9820 FP-855422 P/N : FP-BH9390 876982 876981 FP-895514 FP-36774A (instead measurement, to microplate measure of dye Fluorescence of calcein atpH9.0 of calcein Fluorescence 10 read cell µl) assay replacing 20 x50µg 200 µl 100 µl 1 ml Qty 100 tests 2 x500tests 500 tests 1 mg 10 ml viability assay at 512 has solution a nm, the and/or detection calcein, cell to cytotoxicity. culture 50 range hydrolyzed µl PBS medium of The less solution by with kit than esterases utilizes PBS per 50 well. cells is calcein necessary in Since to cells, more AM esterases is than directly prior for the 25 to Live cell staining
■ CFDA, SE for microbial and cell enumeration
5-(and 6-)-carboxyfluorescein diacetate, succinimidyl ester (CFDA, SE) efficiently stains gram-negative and gram-positive bacterial genera without causing undesirable effects on cell adhesion or viability. The high throughput method using microplate spectrofluorometry has a detection limit of mid-105 CFDA-stained cells/ml. CFDA, SE tracking technique has applications in bacterial transport, public health microbiology, allowing the movement of pathogen to be monitored in terrestrial, aquatic, and even food-processing environments. The technique may also be useful for studying infection and colonization by pathogens in vivo using animal models.
Reference : Mark F. et al. - Development of a Vital Fluorescent Staining Method for Monitoring Bacterial Transport in Subsurface Environments, Applied and Environmental Microbiology, October 2000, p. 4486-4496, Vol. 66, No. 10 Viability & Cytotoxicity Assays Description P/N : Qty CFDA-SE (CFSE, Green Cell Tracking reagent) FP-52493A 25 mg
λex./em. (cleaved) : 495/519 nm ; MW : 557 ■ UptiBlue Cell Viability Assay Kit
Substrate : Resazurin
λex./em. : 540 / 590 nm Sensitive : 100 cells Principle : the UptiBlue dye enters readily into cells, where it elicits a wavelength shift of absorbance and a strong fluorescence related to redox potential in cell, informing on cell energetic state.
UptiBlue shows excellent correlation to formazan and tritiated thymidine techniques, while being much easier and safer to use. It especially replaces advantagely MTT/XTT in many applications, from cell counting to proliferation assay and cytotoxicity testing. Furthermore it allows longer studies.
Applications
Cell proliferation assay Kinetic / long term assays Cytotoxicity assay
Detection of cell Growth of 4 Cell Lines using Kinetic reduction curve with UptiBlue with plating Determination of Doxorubicin LD50 using UptiBlue UptiBlue density from 500 to 10000 cells A549/ml. and XTT
Description P/N : Qty UptiBlue Cell Viability Assay Kit UP669412 25 ml UP669413 100 ml
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 33 34 Viability & Cytotoxicity Assays Reduced toxicityofassaysolution: » » » » » » ■ WST-8 CellProliferationandCytotoxicity Live cell staining HeLa cellsandHL60cells.Furthermore,thecellproliferationassaydatausingCCK-8correlateswiththat sensitivity the WST-1 asstandaloneproduct WST-1 CellProliferation XTT XTT MTT MTT Also available: CKK-8 CellCountingKit Description CCK-8 is measured More sensitivethanMTT, XTT, MTSandWST-1 steprequired orsolubilization Easy andfast:noharvesting,washing No toxicitytocells required ororganicsolvent Safe :noradioisotope Ready-to-use one-bottlesolution assay Colorimetric microplate yellow UltraPure CellProliferation UltraPure CellProliferation consists using colored in 96 CCK-8 of or formazan WST-8 Assay Kit Assay Kit 384-well Assay Kit is higher and dye plate. 1-methoxy than generated The that wavelength using PMS by dehydrogenases MTT as an
or range electron the F98883 FP-409036A FX873A FP-65939A 45547A 899654 899651 899650 KS0790 P/N : other .com.com for votre source d’innovations the mediator. in tetrazolium cells measurement Cell proliferationassay: is Assay Kit After directly salts the of proportional that plate 100 mg 1 g 1000 tests 1 g 1000 tests 10000 tests 3000 tests 1000 tests 96 tests Qty the produce absorbance is incubated to water-soluble the is number for between WST-8 related reagent. cell n xrcin tp n lmtd sensitivity. limited and Interchim step extraction an including MTT Formazan basedCellViability Technical tip 1-4 viability
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■ aCella™ - TOX Bioluminescence Cytotoxicity Assay (GAPDH) Measurement of Cell-Mediated (T Cells, ADCC, NK) or Complement-Mediated Cytolysis
» Versatile : Assay can be run in serum supplemented media. » Homogenous - One-step, no wash assay. Assay can be run in same plate as samples. » FAST - Results in 3-5 minutes. » Highly Sensitive - Can detect fewer than 500 cells/well. » Works with PRIMARY CELLS for determining cell Cytotoxicity. » Non-destructive assay allows monitoring of additional parameters.
aCella-TOX provides a new and highly sensitive assay using a patented coupled luminescent technology for the detection of cytotoxicity(1). This assay quantitatively measures the Viability & Cytotoxicity Assays release of Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) from Primary Cells, mammalian cell lines, bacterial cells(1,2,3). aCella-TOX can work in different media formulations and allows overnight assays while retaining its sensitivity. The sensitivity of aCella-TOX is also greatly enhanced by the coupled luminescent signal-amplification system (3-Phosphoglyceric Phosphokinase/ATP/Luciferase), which yields a strong luminescent signal from even small amounts of released enzyme.
In the aCella-TOX reaction scheme the release of GAPDH Jurkat cells were plated at various cell concentrations per well. NP-40 cytotoxic agent was is coupled to the activity of the enzyme 3-Phosphoglyceric added to each well. The aCella-TOX kit was used to detected G3PDH enzyme release. Data points show average RLU in triplicate. Phosphokinase (PGK) to produce ATP. ATP is detected via the luciferase, luciferin Bioluminescence methodology. Further, aCella-TOX is a homogeneous cytotoxicity assay ; alternatively in dual mode, aCella-TOX can measure cytotoxicity and cell viability in the same plate. Culture supernatants can also be removed from the original plate and assayed in a different plate, allowing kinetics runs to be set up. The assay is non-destructive, allowing the monitoring of additional parameters such as gene expression. The method is highly general, since all known cells express copious amounts of GAPDH, and, unlike other enzymes, GAPDH is very readily released from the cytoplasm upon cell lysis. Using specially adapted formulations, the sensitivity of the method can be driven below 1 eukaryotic cell (2).
Applications : The aCella-TOX method has been tested with many modes of cytolysis, including : » Cellular cytotoxicity (T cells) » Complement(2,3), pore-forming agents » Antibiotic-mediated lysis of bacteria » Detergent mediated and mechanical lysis
References : 1. Methods and compositions for coupled luminescent assays. United States Patent 6,811,990 Corey and Kinders, issued November 2, 2004. 2. Corey, M. J. and Kinders, R. J. (2005), Drug Discovery Handbook, Ed. Shayne Cox Gad, pp. 689-731 3. Corey, M.J., et al Journal of Immunological Methods 207:43-51, 1997. 4. Corey, M. J., et al., Journal of Biological Chemistry 275: 12917-12925, 2000. 5. Ogbomo H., et al. - Biochemical and Biophysical Research Comunications 339 (2006) pp375-379. 6. Corey, J. and Kinders, J. (2005), Drug Discovery Handbook, Ed. Shayne Cox Gad, pp. 689-731
Description P/N : Qty aCella-Tox bioluminescent Cytotoxicity Assay CA4670 500 tests Kit Content : 4x Enzyme Assay Reagent, 1x Enzyme Assay Diluent, Glyeraldehyde 3-Phosphate (G3P), 50x Detection Reagent, 5.5x Detection Assay Diluent, Lytic Agent
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 35 36 Viability & Cytotoxicity Assays high to fourhours,useourSteadyGlow ARL-67156 Ecto-ATPase inhibitor Reaction Buffer asstandaloneproduct Reaction Buffer asstandaloneproduct ATP Related products: Component D:ReactionBuffer (readytousebuffer) Component C:DithiothreitolDTT Component B:D-Luciferin(todissolveinreactionbuffer) Component Each kitcontains : ATP Description incubation signal concentrations The or Pyrophosphate Assays ATP, ADP, Phosphate & ATP Description Catalysed byfireflyluciferasethesubstrateD-luciferinisoxidizedinan To detect ■ The Sensitivity : shows 20min,1,2,3,4,and5hrsignal). λ Substrate :luciferinwithstabilizer Linear λ Substrate :luciferinwithstabilizer ■ cellular Adenosine Sensitivity : performed The long as4hours.Ithasstableluminescence withnomixingorseparationsrequired,andformulatedtohave minimal hands-ontime. foods. of detection the permits assay this of sensitivity high The format. and proliferation cell of determination the for assay luminescence em. em. luciferin + ATP ATP :560nm :560nm ATP ATP disodiumsalt
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■ ATP Assay Kit, Bright Glow
Substrate : luciferin with stabilizer
λem. : 560 nm Sensitivity : 10 cells/well - 3 pmol ATP 2 h incubation time
Adenosine triphosphate (ATP) plays a fundamental role in cellular energenics, metabolic regulation and cellular signaling. The ATP Assay Kit provides a fast, simple and homogeneous luminescence assay for the determination of cell proliferation and cytotoxicity in mammalian cells. The high sensitivity of this assay permits the detection of ATP in many biological systems, environmental samples and Viability & Cytotoxicity Assays foods. This ATP Assay Kit can detect as low as 10 cells/well. It has stable luminescence with no mixing or separations required, and formulated to have minimal hands-on time.
ATP dose response on 96-well white plate using 2 h incubation time (Z’ factor = 0.6, Blue 30 min, red 1 h, and green 2 h). The integration time was 1 sec. The half life is more than 1.5 h.
Description P/N : Qty ATP Assay Kit, Bright Glow FN0640 96 assays
■ Universal Fluorimetric Kinase Assay Kit, Red Fluorescence
Most of commercial protein kinase assay kits are either based on monitoring of phosphopeptide formation or ATP deletion. For the kinase assay kits that are based on detection of phosphopeptides one has to spend time and efforts to identify an optimized peptide substrate while the ATP depletion method suffers various interferences due to the use of luciferase that are inhibited or activated by various biological compounds. The Universal Kinase Assay Kit is based on the monitoring of ADP formation, which is directly proportional to enzyme phoshphotransferase activity and is measured fluorimetricaly. This kit provides a fast, simple, and homogeneous assay for measure kinases activities. The characteristics of its high sensitivity (<0.2 uM ADP), broad ATP tolerance (1-300 uM), non-antibody based, non-radioactive and no-wash method to detect the amount of ADP produced as a result of enzyme activity make it an ideal kit for determining kinase Michaelis-Menten kinetics and for screening and identifying kinase inhibitors.
» Universal : Can be used for any kinases that used ATP as phosphate donor. » Continuous : Easily adapted to automation with no mixing or separation protocols. » Use of Native substrates : Substrates can be proteins, peptides or sugars. » Non-Antibody-Based : No antibody is used in the kit.
Description P/N : Qty Universal Fluorimetric Kinase Assay Kit (540/590 nm) CL9170 250 assays Contains : ADP sensor buffer, ADP Sensor, ADP standard, ADP Assay Buffer votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 37 38 Viability & Cytotoxicity Assays for assay homogeneous and simple fast, a provides as λ Substrate :redfluorescentsubstrate ■ Most metabolic assay rprinl o nye phosphotransferase enzyme activity to proportional for ADP Large Rangeof Sensitivity : either other broad their to due discovery drug in involved researchers hands-on automation. by is assay formeasurekinasesactivities. Pyrophosphate Assays ATP, ADP, Phosphate & Description ADP ex./em. directly protein ADP assaying monitoring measuring
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■ Phosphate Assay Kit, Blue Fluorescence
Substrate : proprietary substrate
λex./em : 370 / 420 nm
Phosphate is involved in many biological reactions. For example, phosphatases, ATPases and several other enzymes catalyze reactions in which inorganic phosphate (Pi) is released from a substrate. This Phosphate Assay Kit has been developed for measuring the activity of any Pi-generating enzyme. The kit is formulated to give the simplest detection of Pi, neither coupling enzymes nor hazardous radioactive methods are involved. The measurement of Pi is based on our proprietary fluorescent sensor that has its fluorescence intensity proportionally dependent on phosphate concentration. Unlike other phosphate assays, this kit is easy to use. It is a mix and read format, and compatible with all the biological buffers.
Description P/N : Qty
Phosphate Assay Kit, Blue Fluorescence JQ8120 1 kit Viability & Cytotoxicity Assays
■ Phosphate Assay Kit, Red Fluorescence
Substrate : proprietary substrate
λex./em. : 540 / 590 nm Sensitivity : 0.1 μM phosphate
Cells utilize a wide variety of phosphate (Pi) and polyphosphate esters as enzyme substrates, second messengers, membrane structural components and vital energy reservoirs. Phosphate is involved in many biological processes. For example, phosphatases, ATPases and several other enzymes catalyze biochemical reactions in which inorganic phosphate is released from a phosphoester substrate. Detection of many phosphoester– metabolizing enzymes is difficult because suitable substrates are not available. It usually has been necessary to determine inorganic phosphate release using tedious colorimetric assays or radioisotope-based methods. This Fluorimetric Phosphate Assay Kit has been developed for measuring the activity of any Pi- generating enzyme using our red fluorescent Phosphate dose response on 96-well black plate with 1 hr incubation time phosphate sensor. The measurement of Pi is based on the change in the absorbance and fluorescence of our new phosphate sensor. Our kit provides all the essential reagents including phosphate sensor, phosphate standards and assay buffer. It can be used to measure the kinetics of phosphate release from phosphatases (such as GTPases and ATPases) by coupling the two enzymatic reactions. The assay can be performed in a convenient 96-well or 384-well microtiter-plate format and easily adapted to automation with no separation steps required.
Description P/N : Qty Phosphate Assay Kit, Red Fluorescence CI4161 100 assays
Also available : colorimetric phosphate assays* Phosphate Assay, MG method IS2790 1kit (600 assays) Original molybdate and malachyte green dye method. 600-660 nm reading. *The kit can also be used to estimate the phosphate content of proteins (phosphoserine or Phosphate Assay, MG Plus method CI4211 1kit (1000 assays) phosphothreonine post-translational modifications, Improved end-point stable signal (not prone to precipitation) after alkaline hydrolysis.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 39 40 Viability & Cytotoxicity Assays Description Pyrophosphate Pyrophosphate Assays ATP, ADP, Phosphate & assaying pyrophosphate. nye fr hi prpopae detections. pyrophosphate The their for enzymes pyrophosphate and uses method sensor Kit DNA of of concentration the upon dependent proportionally λ Substrate :proprietarysubstrate ■ Pyrophosphate Pyrophosphate Sensitivity : coenzyme the enzymatic activation of fatty acids to form their formation ex./em. biochemical pyrophosphate. provides
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hydrolysis, a least intensity This number easier Assay AMP and two for kit
P/N : .com.com JQ8080 votre source d’innovations Qty 200 assays NAD/NADH, NADP/NADPH Assay Kits
Nicotinamide adenine dinucleotide (NAD+) and nicotinamide adenine dinucleotide phosphate (NADP+) are two important cofactors found in cells. NADH is the reduced form of NAD+, and NAD+ is the oxidized form of NADH. It forms NADP with the addition of a phosphate group to the 2' position of the adenyl nucleotide through an ester linkage. NADP is used in anabolic biological reactions, such as fatty acid and nucleic acid synthesis, which require NADPH as a reducing agent. In chloroplasts, NADP is an oxidizing agent important in the preliminary reactions of photosynthesis. The NADPH produced by photosynthesis is then used as reducing power for the biosynthetic reactions in the Calvin cycle of photosynthesis. The traditional NAD/NADH and NADP/NADPH assays are done by monitoring of NADH or NADPH absorption at 340 nm. This method suffers low sensitivity and high interference since the assay is done in the UV range that requires expensive quartz microplate. Theses NAD/NADH & NADP/NADPH Assay Kits provide a convenient method for sensitive detection. In addition, this assay has very low background since it is run in the red visible range that significantly reduces the interference from biological samples. The assay has demonstrated high sensitivity and low interference with 570 nm excitation 590 nm emission. ■ NADH Assay Kit, Red Fluorescence Viability & Cytotoxicity Assays Sensitivity : 10 nanomoles of NADH in solution
The enzymes in the system specifically recognize NADH in an enzyme cycling reaction which significantly increases detection sensitivity.
NADPH dose response on 96-well black plate was measured with 1 hour incubation time (n=3) while there is no response from NADP (the insert shows the lower detection limit).
Description P/N : Qty Fluorimetric NADPH Assay Kit JQ7320 400 assays
■ NAD/NADH Assay Kit, Red fluorescence
Sensitivity : 100 nM (10 pmol/well) of NADH in solution
λexc./em.: 570/590 nm
The enzymes in the system specifically recognize NAD/NADH in an enzyme cycling reaction. There is no need to purify NAD/NADH from sample mix. The enzyme cycling reaction significantly increases detection sensitivity.
NADH dose response on 96-well black plate was measured with 1 hour incubation time (n=3) while there is no response from NADPH.
Description P/N : Qty NAD/NADH Assay Kit, Red fluorescence JQ7280 400 assays
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 41 42 Viability & Cytotoxicity Assays NADPH The NADPH insolution Sensitivity : ■ NADPH measured recognize ecin Te nye yln reaction cycling significantly increasesdetectionsensitivity. enzyme The reaction. there the lowerdetectionlimit). NAD/NADH, NADP/NADPHAssay Kits NADP/NADPH Description NADPH in ape i. h ezm ccig reaction significantly increasesdetectionsensitivity. cycling enzyme The mix. sample Fluorimetric NADPH incubation measured NADPH is NADP, of detection sensitive for method convenient This in solution Sensitivity : ■ NADP/NADPH Description NADH. the no is enzymes NADP/NADPH no dose system need NADPH dose in with with time response NADPH an response 30 nM(0,3nmol/well)of 10 nM(1pmol/well)ofNADPH Assay Kit,Redfluorescence response 1 NADP/NADPH (n=3) to enzyme hour specifically in and Assay Kit,RedFluorescence purify from Assay Kit,Redfluorescence while the incubation in on their on Assay NADP cycling an 96-well system there NADP/NADPH 96-well ratio. Assay enzyme
recognize Assay Kit,Redfluorescence (the is time black Kit reaction. no black The specifically insert Kit (n=3) response provides plate with cycling enzymes plate shows while NADP/
was 30 There from from was min a P/N : JQ7300 JQ7330 .com.com P/N : votre source d’innovations 400 assays Qty Qty 400 assays Caspases study
■ Caspases Fluorometric HTS Assay Kits
» HTS-compatible : Single-step homogenous assay specifically designed for HTS-based detection. » Fast : Fast enzyme kinetics. » Sensitive : The enzymatic reaction forms intensely green fluorescent rhodamine 110 (R110) product. The long wavelengths of R110 excitation
and emission minimize cellular autofluorescence (λex\λem= 496/520 nm).
Caspases play important roles in apoptosis and cell signaling. Caspases Fluorometric HTS Assay Kits are specifically designed for HTS-based assays. The kits provide a homogenous assay system for fast and highly sensitive detection of specific caspase activity by fluorescence in enzymatic reaction or mammalian cells.
The assay kits include a caspase inhibitor and can be used as a negative control. Also, R110 dye is provided in the kit for generating a standard curve, which can be used for quantifying caspase activity.
The fluorogenic substrate R110-labeled contains two specific tetrapeptides and is completely hydrolyzed by the enzyme in two successive steps 3, 4. Cleavage of the first peptide results in the monopeptide intermediate, which has absorption and emission wavelengths similar to those of R110, but has only about 10% of the fluorescence of the latter. Hydrolysis of the second peptide releases the dye R110, leading to a substantial fluorescence increase. Apoptosis Assays
The fluorogenic substrate (Ac-IETD)2-R110 contains two IETD tetrapeptides and is completely hydrolyzed by the enzyme in two successive steps.
The fluorogenic substrate (Ac-LEHD)2-R110 contains two LEHD tetrapeptides and is completely hydrolyzed by the enzyme in two successive steps.
Reference : 1. Cell Death Diff. 6, 99(1999); 2) J. Biol. Chem.. 274, 11549(1999); 3) J. Biol. Chem. 275, 288(2000); 4) Biochemistry, 38, 13906(1999)
Description P/N : Qty Caspase-3 Fluorometric HTS Assay Kit, DEVD-R110 FP-BR4930 1 ml FP-BR4931 10 ml FP-BR4932 100 ml
Kit Components : Cell lysis/assay buffer , Enzyme substrate (Ac-DEVD)2-R110 , Enzyme inhibitor Ac-DEVD-CHO, R110 Caspase-8 Fluorometric HTS Assay Kit, IETD-R110 FP-BX1510 1 ml FP-BX1511 10 ml FP-BX1512 100 ml
Kit Components : Cell lysis/assay buffer, Enzyme substrate (Ac-IETD)2-R110, Enzyme inhibitor Ac-IETD-CHO, R110 Caspase-9 Fluorometric HTS Assay Kit, LEHD-R110 FP-BX1530 1 ml FP-BX1531 10 ml FP-BX1532 100 ml
Kit Components : Cell lysis/assay buffer , Enzyme substrate (Ac-LEHD)2-R110 , Enzyme inhibitor Ac-LEHD-CHO, R110 Related products : Annexin V-FluoProbes 488, FCM grade (495/519 mm) FP-BH9390 100 tests Staurosporine (apoptosis inducer) 74146D 100 µg 74146E 1 mg See BioScience Innovations catalog, Membrane apoptosis events.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 43 44 Apoptosis Assays Caspases study which canbeusedforquantifyingcaspaseactivity. See BioScienceInnovationscatalog,Membraneapoptosisevents Annexin V-FluoProbes 488 Staurosporine, proteinkinaseinhibitor Related products: Kit Components:Celllysisbuffer, Caspase-9 FluorometricandColorimetric Kit Components:Celllysisbuffer, Caspase-8 FluorometricandColorimetric Kit Components:Celllysisbuffer, Caspase-3 FluorometricandColorimetric Description The based substratessignificantlymoresensitivethanpNA-basedsubstrates,evenbycolorimetricdetection. the Although fluorometricdetectionoftheendproductsispreferredbecause The principleisthesameasforCaspasesFluorometricHTS » » » Continuous measurementofthecaspaseactivity ■ CaspasesFluorimetricandColorimetric Versatile andcolorimetricdetectionsystems. :Compatiblewithbothfluorometric Sensitive :Rhodamine110 (496/520nm)minimizescellularautofluorescence Fast enzymekinetics extinction assay kit coefficient includes a of caspase R110 Assay buffer, Enzymesubstrate(Ac-LEHD) Assay buffer, Enzymesubstrate(Ac-IETD) Assay buffer, Enzymesubstrate(Ac-DEVD) is inhibitor 10 Assay Kit,LEHD-R110 Assay Kit,IETD-R110 Assay Kit,z-DEVD-R110 times and higher can than be used that .com.com as of votre source d’innovations p-nitroaniline a 2 -R110, Enzymeinhibitor 2 Assay Kits. 2 -R110, Enzyme negative -R110, Enzyme FP-BH4140 74146D FP-BX1521 FP-BX1520 FP-BR4941 FP-BR4940 FP-85785B FP-85785C P/N : Assay Kits control. (pNA), inhibitor inhibitor superior sensitivity, detection by absorbanceisalsopossible.Infact, Also, a Ac-IETD-CHO, R110 dye Ac-LEHD-CHO, R110 Ac-DEVD-CHO, R110 R110 commonly is 500 µl 100 µg 100 tests 25 tests 100 tests 25 tests 100 tests 25 tests Qty provided used in in the chromogenic kit for generating substrates, a standard making curve, R110- Caspases study
■ Caspase-6 and GranzymeB based apoptosis/toxicity Assays
Cytotoxicity is measured as functions of fundamental biochemical pathways leading to cell death : -in the CyToxiLux® kit, cleavage of a cell permeable fluorogenic very specific substrate of caspase-6, an established initial activation step in apotosis. Literature : Nature Med. 8:185-189 (2002)
-in the GranToxiLux™ kit, cleavage of a cell permeable substrate for Granzyme B, involve in a early event of cell-mediated apotosis. Granzyme B that is inactive in lysosomal granules, is activated in the presence of Target cells during (degranulation), extremely early (before Perforin). Literature : Nature Med. 8:185-189 (2002); Methods Mol. Biol. 263:125-140 (2004): J. Immunol. 171:27-31 (2003)
These assays provide an extremely early quantitative assessment of caspase-6 and cell-mediated cellular cytotoxicity. Especially Grantoxilux replaces advantageously the classic 51Cr release assay is an end stage of cell mediated cytotoxicity, i.e. after cell lysis. Furthermore that can be used by FCM and microscopy yielding single cell measurements in complex populations.
Benefits : » More versatile in applications : suits CTL/NK and other factor mediated cytotoxicity, cytotoxicity induced by intracellular agents or xenobioteics, physiology and fate of effector cells » More rapid : co-incubation of 0.3-2 H ( vs. 4 H for 51Cr release assay) » More sensitive than the 51Cr method : can detect relatively weak CTL responses against subdominant epitopes whereas the latter cannot. Apoptosis Assays » Large study period : hour to days allow long term studies, that is useful for non- or slow proliferating cells » compatible with multiparametric FCM & Microscopy analysis at the cell level, even in mixed populations » No seric interferences : avoid this limitation of LDH and Formazan methods » No pre-labeling of cells : avoid this limitation of 51Cr method
Cytoxilux 51 Cr Assay
Description P/N : Qty Grantoxilux cytotoxicity assay (Fluo.) BP8891 50 tests Measure the GranzymeB (path of cell-mediated apoptosis) Cytoxilux cytotoxicity assay (Fluo.) BP8881 50 tests Measure the caspase-6 (classic path of apoptosis)
Each kit contains sufficient reagents for 50 assays i n FCM. It may be applied also for microscopy with some modifications (Caspase-6 or GranzymeB Substrate solution, Target cell marker for use with single laser instruments (Ar ion(488 nm), Target cell marker for use with dual laser instruments (Ar ion(488 nm) and Red (633 nm)), Resuspension medium , Wash Buffer bottle, Assay/Culture medium. votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 45 46 Apoptosis Assays DNA Damage&Condensation study and ischemia, neuronal and myocardial of models animal in damage have This and Recent » Applications : extracts. ribose) measures cancer by » » » FITC-NAD Related product: Universal ChemiluminescentPARP Universal ChemiluminescentPARP Description ribose) translational » » » ■ UniversalChemiluminescentPARP important catalyzes damage. Poly » depleting Identify inhibitorsandactivatorsofPARP Quantitate levels Measure caspaseinactivationofPARP Measure activity Sensitivity downto0.0025unitsofPARP Higher throughput96testsize format Chemiluminescent, non-radioactive histone reagent Available plateor eitherwithahistonecoated PARP-1 for assay ADP-ribosylation shown development. measuring onto to data role diabetes, the the adjacent Poly is the in histone event in implicate ideal NAD-dependent incorporation that minimizing DNA cellular (ADP-ribose) the
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votre source d’innovations Assay Kit FX8241 HP9130 HP9090 P/N : 250 µl 96 tests 96 tests Qty DNA Damage & Condensation study
■ HT Chemiluminescent PARP/Apoptosis Assay ELISA assay kit for monitoring PARP activity before and during apoptosis
Sensitivity down to 0.1 mUnits of PARP - less than 500 cells/well
During apoptosis, PARP-1 which catalyzes the NAD dependent addition of poly (ADP-ribose) (PAR) onto various cytoplasmic and nuclear proteins, is cleaved from about 116 kDa to 85 kDa. HT PARP/Apoptosis Assay is ideal for measuring the activity of PARP in cell extracts before and during apoptosis. The HT PARP/Apoptosis Assay is an ELISA which semi-quantitatively detects PAR deposited onto immobilized histone proteins in a 96-well format. An anti-PAR monoclonal antibody, goat anti- mouse IgG-HRP conjugate, and HRP substrate are used to generate a chemiluminescent signal. Thus, absorbance correlates with PARP activity. Etoposide is a topoisomerase II inhibitor that stabilizes this enzyme after it cleaves DNA. It is included as a control apoptosis inducer.
Graphical representation of an example Chemiluminescent Apoptosis Assays readout of a PARP standard curve.
Description P/N : Qty HT Chemiluminescent PARP/Apoptosis Assay CP1990 96 tests Kit content : PARP Buffer, activated DNA, PeroxyGlow™ A & B, Histone coated strip wells, PARP HSA, NAD, Antibody diluent, anti-PAR monoclonal antibody, HRP conjugate diluent, Etoposide.
■ HT Chemiluminescent PARG Assay Kit
» Chemiluminescent format » Higher throughput 96 test size » Sensitivity down to 50 pg of PARG per well
Poly(ADP-ribose) glycohydrolase (PARG) degrades poly(ADP-ribose) (PAR) polymers synthesized by poly(ADP-ribose) polymerase (PARP1). When activated by DNA strand breaks, PARP1 uses NAD as a substrate to form ADP-ribose polymers on itself and on specific acceptor proteins such as histones, DNA polymerases, DNA ligases, p53, and Fos. These polymers are in turn rapidly degraded by PARG, a ubiquitously expressed exo- and endoglycohydrolase. Excessive activation of PARP1 leads to NAD depletion and cell death during ischemia and other conditions that generate extensive DNA damage. PARG may maintain the active state of PARP1 by continuously removing inhibitory ADP-ribose residues from PARP1. The regulation of PARG activity may therefore, influence PARP-mediated cell death. First, PARG inhibition could slow the turnover of PAR and thereby limit NAD depletion. Second, PARG inhibition could prevent the removal of PAR from PARP1. Because PARP1 is inhibited by extensive poly-(ADP-ribosyl)ation, PARG inhibitors could thereby indirectly inhibit PARP1 activity. Prior work has shown that the PARG inhibitor gallotannin can markedly reduce death of astrocytes after oxidative stress. HT Chemiluminescent PARG Assay Kit measures the loss of biotinylated PAR from histones attached to strip wells in a 96 well format and is ideal for the screening of PARG inhibitors and for measuring the activity of PARG in cell extracts.
Applications : » Identify inhibitors and activators of PARG activity » Measure activity of PARG in cell and tissue extracts
Description P/N : Qty HT Chemiluminescent PARG Assay Kit JZ4060 96 tests votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 47 48 Apoptosis Assays Hoechst 33342,10mg/mlinwater Description DNA Damage&Condensation study This biohazardous agents. may to Debbasch Reference : Cell-permeant bis-benzimidethatbindstoDNA ■ Hoechst33342 Propidium iodide/RNasesolution,Nucleotidemix,Fluorescentlabel,Labelingenzyme,Counterstain Kit content:Permeabilizationreagents,Optimizedcation, FlowTACS™ Description DNA Identify andquantitateapoptoticcellsinculture ■ FlowTACS™ Vis Sci.2001Oct;42(11):2525-33. » Features : detection reagents,and » » » » » generate Fast. Requireslessthan Includes Cytonin™permeabilizationreagent. Includes exclusive Works onfixedcells. Unique buffer consistentlabeling. systemproduces more Exclusive, non-toxic
also complete fragmentation C, be et Apoptosis DetectionKits TACS-Nuclease preparingsample-dependentpositive solutionfor positive analyzed al., kit Mitochondrial provides is controls Also, samplesmaybestoredconvenientlyduringtime-courseexperiments. for a TACS-Nuclease forgeneratingpositivecontrolswithyourownsamples. committed TACS Safe Apoptosis DetectionKits DNA all activity and 3 hourstocomplete. for
the content calibration. reagents step TdT™ buffer free. -sodiumcacodylate glutathione using in apoptosis, required the The withfluorescenceenhancement(λ injury TACS-Nuclease, Labelingbuffer, Detectionreagents, included FlowTACS™ in for and apoptosis labeling propidium FP-59046A P/N : 512510 P/N :
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■ JC-10 Mitochondria Membrane Potential Assay Kit
» Increased Signal Intensity : Larger assay window » Increased solubility : Much better water solubility than JC-1 » Convenient and Robust : Formulated to have minimal hands-on time » Versatile applications : Compatible with many cell lines and targets Although JC-1 is widely used in many labs, its poor water solubility makes it hard to use for some applications. Even at 1 µM concentration, JC-1 tends to precipitate in aqueous buffer. JC-10 has been developed to be a superior alternative to JC-1 where high dye concentration is desired. Compared to JC-1, our JC-10 has much better water solubility. JC-10 is capable of entering selectively into mitochondria, and changes reversibly its color from green to orange as membrane potentials increase. This property is due to the reversible formation of JC-10 aggregates upon membrane polarization that causes shifts in emitted light from 520 nm
(i.e., emission of JC-10 monomeric form) to 570 nm (i.e., Apoptosis Assays emission of J-aggregate). When excited at 490 nm, the color of JC-10 changes reversibly from green to greenish orange as the mitochondrial membrane becomes more polarized. This JC-10 Mitochondrial Membrane Potential Assay Kit enable you to monitor mitochondrial membrane potential changes using a simple microplate reader while all JC-10 and JC-1 Comparison on Effect of Campotothecin induced mitochondria the other commercial JC-1 assay kits require the use of a membrane potential change in JurKat cells. Jurkat cells were treated with 10 µM flow cytometer. Our kit provides the most robust method to camptothecin for 4 hours. JC-1 and JC-10 dye loading solution was then added to the monitor mitochondrial membrane potential changes, and can wells for 30 minutes. The fluorescent intensity for both J-aggregates and monomeric be readily used for screening a large compound library. forms of JC-1 and JC-10 was measured at Ex 485 nm/Em 520 and 595 nm.
Description P/N : Qty JC-10 Mitochondria Membrane Potential Assay Kit DT2420 5 plates (96- or 384-well) JC-10 CL0440 5 mg Related products : CCCP 091640 500 mg JC-1, as stand alone product FP-52314A 5 mg
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 49 50 Apoptosis Assays Mitochondrial eventsMitochondrial (MCB), further » » membrane potential(DeltaPsi(m)) lysate withMCB,theintensityoffluorescentsignalgeneratedfromassayreflectsamountGSHpresentincells. to highΔΨm(Feldkamp,2004). procedure Kit contains:Monochlorobimane, CellLysis Buffer, L-Glutathione,GlutathioneS-transferase Live CellGlutathione Description λ ■ LiveCellGlutathioneTransferase Monochlorobimane, asstandaloneproduct Related product: Kit content:Celllysisbuffer, Monochlorobimane(MCB),GST Glutathione DetectionKit,Monochlorobimane Description 1) FASEB J.12(6),479(1998);2)Biochem. Soc. Reference : Diminished » » GSH depletionfollowupduringearlystageofmitochondrion-associatedapoptosis ■ GlutathioneDetectionKit,Monochlorobimane Ouabain Related products: hypoxia/reoxygenation, Reference :FeldkampT. etal.– Safranine O Description Unlike Fluorescence isfollowedat485nmexcitationand586emission. Dynamic ■ SafranineO uptake membranepotentialdecreases » this GSH-mCB It This Sample size:200cells fluorescence released cells S-transferase. abs allows /λ usrt- ΔΨmand transport-mediated dependent, electron Allows directassessmentofbothsubstrate- Suitable fordynamicstudiesofenergization Fast : Simple :Compatiblewithastandardorfluorescenceplatereader. Avoid limitationofJC-1uptakewhentheplasma ΔΨm. hydrolysis-supported approach new em to by the :380/461nm leads form which when energized intracellular kit and Around 1hourassaytime. behavior has complexes. cellular of provides a mitochondrial to to is safranine quantitative fluorescent been cytochrome essentially measure mitochondria Transferase glutathione of AM. J.Physiol.RenalPhysiol.,288:F1092-F1102 (2005) used glutathione reagents JC-1, Unlike O resulting the were to GSH approach monfluorescent Assessment ofmitochondrialmembranepotentialinproximaltubulesafter safranine c measure GSH means other Activity Kit release de-energized and (GSH) mono-chlorobimane S-transferase from ΔΨm : content methods bimanes that of O level and GSH quenching low was changes Trans. 28,56(2000). with occurs of caspase fluorescence until content to rapidly such the FCCP. simply detection it of cytosolic at of reacts as safranine of 3 and the mitochondrial adduct The monobromobimane, and ATP induction positivecontrol cultured corresponds early completely decreased with by quickly
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Technical tip Oxidative metabolism study (ROS, NO)
The production of free radicals primarily results from O2 catched by cells and reduced in mitochondria. 98% is fully utilized by cytochrome c oxidase to form water, but this enzyme can release partly reduced species. Other respiratory chain enzymes, and in particular complexes I and III, also produce partly reduced oxygen species including superoxide. These reactive oxygen species can react with nitric oxide to produce reactive nitrogen species including peroxynitrite. A significant proportion of the reactive oxygen and nitrogen species diffuse with controlled rate into the cytosol, where they react with various molecules, lipids, proteins, sugars and nucleotides. But a major portion remains in the mitochondrion where they causes oxidative damage. When the electron transfer efficiency decreases, more radicals are produced, and so more cytosolic proteins are damaged. Moreover, oxidative and nitrative damage of mitochondrial proteins adds to OXPHOS dysfunction further exacerbating free radical production. A protective mechanism against ROS is SOD metabolism.
Enhanced oxidative stress occurs in number degenerative diseases. In human, ROS are considered to be one of the main causes of aging-related diseases, Parkinsons disease, Alzheimers and other vascular-damage-related brain diseases, Cancer, Artherioschlerosis and diabetes.
In plants, the SOD activity is increased by the use of herbicides such as paraquat, by the SO2 Apoptosis Assays concentration in the atmosphere, by drought, or by exposure to high concentration of zinc and magnesium. ROS probes have high selectivity and sensitivity in enzymatic oxidation reactions, favorising their use for diagnostic analysis. Also, peroxidase is a common enzyme for signal amplification in immunoassays (EIA).
Interchim provides several ROS probes for fluorimetry and colorimetry, chromogenic, fluorogenic or luminogenic.
Selection guide
Fluorescent ROS probes
Reactive Oxygen Species (ROS) P/N : Probes Hydrogen Peroxide Hydroxy radical Hypochlorous acid Peroxyl radical Peroxynitrite anion Superoxide anion H202 H0- HOCl COO- ONOO- O2- FP-83775 Dihydrorhodamine 123 + + + FP-46731 H2DCFDA + + + + FP-46915 Lucigenin* + FP-97233 Coelenterazine + + FP-38544 MCLA* + + 1 Dihydrocalcein AM + ( O2) + + Dihydroethidium - + 24200A tMPV CA7170 HPF + + CA7270 APF - + + - + - U3238A ADHP +
*ROS-generating Enzyme Detection kits MyeloPeroxidase detection kit Hemoprotein of PMNs cells / Cl- oxidation to HOCl
Catalase detection kit antioxidant enzyme / decomposes H2O2 Cis-Parinaric acid fatty acid to monitor lipid peroxidation Malachite Green IT produce a Hydroxyl radical burst upon irradiation -● SuperOxideDismutase (SOD) converts O2 into H2O2 and O2
*Other oxidative species : aldehydes SSAO detection kit deamination / formaldehyde, methylglyoxal MonoAmineOxidase (MAO A&B) oxidate a variety of neurotransmitters / aldehydes
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 51 52 Apoptosis Assays Reactive OxygenSpecies(ROS) Reactive oxidase–dependent the chemiluminescence (Em=466nm)onoxidationbysuperoxide. leading (λ 2',7'-Dichlorodihydrofluorescein diacetate . KondoM.etal., of cyclomaltononaose(delta-cyclodextrin)." burst inphagocyticcells. λ 2',7'-Dichlorodihydrofluorescein diacetate,succinimidylester Carboxy-2',7'-Dichlorodihydrofluorescein diacetate an 2-Methyl-6-(4-methoxyphenyl)-3,7-dihydroimidazo[1,2-a]pyrazin-3-one, hydrochloride MCLA Description . Reference : MCLA Superoxide orsingletoxygenchemiluminescentprobe ■ MCLA Coelenterazine (native) Description Margaret Reference : Coelenterazine ■ Coelenterazine(native) λ Description Jiyoung Reference : H ■ H neutral rangethanarethepHoptimaofluminolandlucigenin.MCLA . Sakurai Sciences Carboxy-H viability andcytotoxicityassays,apoptosis.ItcanbeusedwithPropidiumiodidetofollowoxidantproductionnuclearinjury. H of 485nmandemissionat535nm. 84:1203-1211 (1999) Teranishi exc. exc. 2 2 exc. DCFDA, DCFDA /λ /λ enhanced cell, /λ em. em. em. 2
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highly is ; oxygen amine sensitive EC retention S., used an results. chemiluminescence Calcium-Mediated : indicator 11 green is reactive to consumption, a 000) chemiluminescent superior of An detect Detection fluorescent the additional and for form Carbohydr Res338,987-93(2003) (H probe. The ROS. reactive nonfluorescent 2 2 alternative DCFDA) DCFDA, H Activation of of of endothelial 6-(4-methoxyphenyl)imidazo[1,2-a]pyrazin-3(7H)-one the Oxidants On advantage 2',7'-dichlorofluorescein (CH H oxygen penetration 2 fluorescence 2 DCFD) of marker DCFDA to -1 c-Jun cm in -1 cm lucigenin (H until cell Vascular -1 of 2 -1 DCFDA-SE) species NH 2 FP-38544A P/N : for
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Reactive Oxygen Species (ROS)
■ Dihydroethidium (hydroethidine) – Measurement of O2
The superoxide oxidizes dihydroethidium to a specific fluorescent product (oxyethidium) that differs from ethidium by the presence of an additional oxygen atom in its molecular structure. Exposure of dihydroethidium to hydrogen peroxide or peroxynitrite caused no formation of oxyethidium from dihydroethidium. Fluorescence detection at 590 nm (emission) and 530 nm (excitation) is used to monitor oxyethidium production.
Reference : Debbasch C, et al., Mitochondrial activity and glutathione injury in apoptosis induced by unpreserved and preserved beta-blockers on Chang conjunctival cells. Invest Ophthalmol Vis Sci. 2001 Oct;42(11):2525-33.
Description P/N : Qty Dihydroethidium, special air-free packaging FP-52492A 25 mg FP-52492B 10 x 1 mg Dihydroethidium, 5 mM in DMSO FP-R5919A 1 ml Dihydroethidium FP-524929 20 x 50 µg
■ Dihydrorhodamine-1,2,3 (DHR 123) Measurement of peroxonitrite (ONOO–) Apoptosis Assays
The level of ONOO– can be measured by monitoring the oxidation of dihydrorhodamine-1,2,3 (DHR 123), using microplate reader at excitation and emission wavelengths of 485 nm and 530 nm, respectively. It is nonfluorescent until oxidized to the mitochondrial probe rhodamine 123.
Reference : Ji Young Lee, et al., Induction of endothelial apoptosis by 4-hydroxyhexenal, Eur. J. Biochem. 271, 1339-1347 (2004) Description P/N : Qty DiHydroRhodamine 123, 5 mM stabilized solution in DMSO FP-R6805A 1 ml Dihydrorhodamine 123 FP-83775A 10 mg Dihydrorhodamine 123, air-free packaging FP-IT397A 10 x 1 mg Dihydrorhodamine 123, 5 mM in DMSO FP-R6805A 1 ml
■ Dihydrocalcein A
This cell permeant probe is oxidized to the calcein with better cell retention than H2DCFDA.
Reference : Keller A, et al. Analysis of dichlorodihydrofluorescein and dihydrocalcein as probes for the detection of intracellular reactive oxygen species. Free Radic Res. 38: 1257–1267 (2004)
Description P/N : Qty Dihydrocalcein AM FP-BA1970 1 mg FP-T7996A 20 x 50 µg
■ trans-1-(2'-Methoxyvinyl)pyrene (tMPV) Sensitive singlet oxygen chemiluminescence probe
trans-1-(2'-Methoxyvinyl)pyrene is the most sensitive singlet oxygen probe that could be used to detect picomole quantities of singlet oxygen in chemical and biological systems. Furthermore, this highly selective probe does not react with other activated oxygen species such as hydroxyl radical, 1 superoxide or hydrogen peroxide. It generates chemiluminescence (Em = 465 nm in 0.1 M SDS) upon reaction with O2.
Reference : Nat Biotechnol 24, 95-9 (2006). Plant and Cell Physiology 46(6):947-954 (2005) Methods Enzymol 133, 569-584 (1986)
Description P/N : Qty trans-1-(2'-Methoxyvinyl)pyrene (353/401 nm) FP-24200A 1 mg votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 53 54 Apoptosis Assays Chemistry Vol. 278,No.5,IssueofJanuary31,pp.3170–3175,2003 ( Oxygen Radical:1O reactive Peroxynitrite :ONOO (H Reactive OxygenSpecies(ROS) Reactive Aminophenyl Fluorescein (APF) Description 1- Reference : ROS (RFU) » » » » » » Selective IndicatorforHighlyReactiveOxygenSpecies ■ Hydroxyl Radical: Autoxidation Alkylperoxyl Radical:ROO. Nitric Oxide:NO Hydrogen Peroxide:H Superoxide :O Hypochlorite : A oxide (NO•),andalkylperoxide(RO2 ueoie (0 superoxide detection. uh s hpclrt ( hypochlorite : as such FeTPPS, specificperoxynitritescavenger Related product: Hydroxyl /PeroxynitriteDetectionKit(HPF) Description Chemistry, Vol. 278,No.5,IssueofJanuary31,pp.3170–3175,2003 Ken-ichi Reference : Fluorescent Hydroxyl( ■ Hydroxyphenylfluorescein(HPF) » » » » » A hydroxyl is permeable highly - OH), Peroxynitrite:(ONOO
new Setsukinai 2 novel O a APF (forhROSdetection) Microscope/Flow Reader/Fluorescent Applications -FluorescencePlate parameters monitoringofadditional Non-destructive cellbasedassayallows Adaptable forHighThroughputformat One-step, nowashassay Quenched cellpermeabledye realtimekinetics Can monitormultipletimepointstofollow Non-destructive realtimekinetics Can monitormultipletimepointstofollow One step,nowash,homogenousassay Can beusedwithcelllysates,tissue Quenched Cellpermeabledye 2 ), highly Reactive novel nitric Setsukinai, oxygen radical probe, It K., highly - has probe, oxide OCl 2 selective - et al., • 2 - , yrgn eoie (H peroxide hydrogen •), species Oxygen • little OH et al.: et (OH•), 2 Hydroxyphenyl (NO•), - Development Aminophenyl sensitive monitoringofadditionalparameters cell basedassayallows 2 0 2 reactivity •OH) /Peroxynitrite(ONOO J. of Bilogical of J. Species. Specific Distinguish and Species Oxygen Reactive Detect Reliably Can That Probes Fluorescence Novel of Development probe (hROS). Species - and OCl), and - ) andhypochlorite( alkyl fluorescent peroxynitrite of for towards singlet Novel The fluorescein (hROS). fluorescein peroxide the •). 3600 1200 APF (RFU) Ex :499Em515 <1 6 9 560 <1 2 <1 probe Fluorescence homogenates oxygen detection other 2 It probe 0 has (ONOO (RO 2 (APF) is , nitric ), hROS (HPF) a - little OCl) productionincells. (0 2 •) Probes cell - 2 for ) assay 1 of (see developed ), - ) reactivity That table 888810 CA7170 P/N : .com CA7270 P/N :
.com Assay Kit Can towards below) votre source d’innovations by Reliably Tetsuo 86 7400 DCFH-DA Ex :500Em520 190 67 26 6600 2000 710 150 1 . other APF Detect Nagano is ROS (RFU) Reactive a Cytometry cell such et. permeable al. Oxygen as: 250 mg 150 tests Qty 150 tests Qty (1) , is a general selective indicator for the detection of highly of detection the for indicator selective general a is , singlet Species indicator oxygen and Distinguish that (O 2 1 can ), superoxide be Specific used Species. to (O detect 2 - •), hydrogen peroxide The Hydroxyl journal of Biological Radical Reactive Oxygen Species (ROS)
■ Hypochlorite Detection Kit Fluorescent OCl (Hypochlorite) assay
» Quenched Cell permeable dye » Can be used with Cell Lysates, Tissue Homogenates » One Step, No wash Homogenous assay » Can monitor multiple time points to follow real time kinetics » Non-destructive cell based assay allows monitoring of additional parameters
The two novel probes, Aminophenyl fluorescein (APF) and Hydroxyphenyl fluorescein (HPF) are selective for the detection of highly reactive oxygen
species (hROS). They offer greater selectivity and stability than does H2DCFDA. Both probes have little reactivity towards other ROS such as: singlet 1 - oxygen (02 ), superoxide (O2 •), hydrogen peroxide (H202), nitric oxide (NO•), and alkyl peroxide (RO2•). HPF/APF are cell permeable and can be used in combination to detect hypochlorite (-OCl) production in cells (see fig 1). Hypochlorite can be detected by loading two samples, one with APF and the other with HPF. Hypochlorite production is visualized by increase in fluorescence of APF loaded cells and no increase in fluorescence in HPF loaded cells.
Reactivity Profile of APF/HPF: ROS (RFU) HPF (RFU) APF (RFU) DCFH-DA (RFU) Apoptosis Assays λex./em.: 499 / 515 nm λex./em.: 499 / 515 nm λex./em.: 500 / 520 nm Hydroxyl Radical: •OH 730 1200 7400 Peroxynitrite: ONOO- 120 560 6600 Hypochlorite: -Ocl 6 3600 86 1 Oxygen Radical: O2 5 9 26 - Superoxide: O2 • 8 6 67
Hydrogen Peroxide : H202 2 1 190 Nitric Oxide: NO 6 1 150 Alkylperoxyl Radical: ROO 17 2 710 Autoxidation 1 1 2000
Description P/N : Qty Hypochlorite Detection Kit (HPF, APF) CA7250 150 tests
■ ADHP Hydrogen peroxide Assay Kit
"Read and mix" sensitive H202 Detection Kit
Sensitive : 10 picomoles of H2O2 in solution.
Hydrogen peroxide (H2O2) is a reactive oxygen metabolic by-product that serves as a key regulator for a number of oxidative stress- related states. It is involved in a number of biological events that have been linked to asthma, atherosclerosis, diabetic vasculopathy, osteoporosis, a number of neurodegenerative diseases and Down's
syndrome. Perhaps the most intriguing aspect of H202 biology is the recent report that antibodies have the capacity to convert molecular oxygen into hydrogen peroxide to contribute to the normal recognition and destruction processes of the immune system. Measurement of this reactive species will help to determine how oxidative stress modulates varied intracellular pathways. Hydrogen Peroxide Assay Kit uses our Red peroxidase substrate to quantify hydrogen peroxide in solutions, in cell extracts and in live cells. It can also be used to detect a variety of oxidase activities through H2O2 dose response on 384-well black plate with 30 minutes incubation time (n=3). The enzyme-coupled reactions. The kit is an optimized "mix and read" insert shows the low levels of H2O2 detection. assay that is compatible with HTS liquid handling instruments. Description P/N : Qty ADHP Hydrogen peroxide Assay Kit CL2580 500 assays Also available :
ADHP Hydrogen peroxide/Peroxidase Assay Kit dual mode, can detect H2O2 or peroxidase activity. U3238A 500 tests
10-acetyl-3,7-dihydroxyphenoxazine (ADPH) λabs/λem. : 563/587 nm FP-39423B 25 mg votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 55 56 Apoptosis Assays Reactive OxygenSpecies(ROS) Reactive including cytochrome molecular expressed detect H substrate, tissue result organelles calledperoxisomes neutrophils 12.CancerRes.,61, 2766-2733(2001). 11. FEBSLett.,473,177-182(2000). 10.FEBSLett.,414, 552-556(1997). 9.FEBSLett.,442, 65-69 (1999). 8.J.Biol.Chem.,Sep1999;274:26217- 26224 7. 6.J.ImmunolMethods202,133(1997). 5. 4.ProgressinBiophys.Mol.Biol.,72,19-66 (1999). 3.Physiol.Rev., 50,319-375(1970). 2. 1.Physiol.Rev., 50,319-375(1970). References : Catalase ■ CatalaseDetectionKit Myeloperoxidase DetectionKit Description a containing group protoporphyrin acting independently an contains chain heavy Each disulfide Myeloperoxidase » » » » » ■ MyeloperoxidaseDetectionKit The lipids thus resulting in cell death and mutagenisis and death cell in resulting thus lipids stress relateddiseaseshavebeenwidelystudied H of The bactericidal yrgn eoie (H peroxide hydrogen contain leukocytes to is seen in connective tissue stress leadingtooxidativetissueinjury. produced H central produce 2 Sensitive Adaptable forHigh One-step, nowashassay kinetics Can monitormultipletimepointstofollow Readout :Fluorescenceorabsorbance Archives ofBiochemistryandBiophysics,431:138-144 (2004). Anal Biochem253,162(1997). Analytical BiochemistryVol. 245,Issue1,1February1997,Pages55-60. O 2 Catalase production 0 2 of 2 to can + Detectionreagent(non-fluorescent)MPOfluorescentanalog. only various 2 tissue. bridge. 0 bacteria by is iron. 2 weight substrateleftoverfromthecatalasereaction 10-Acetyl-3, in (PMNs) a result to activity. system. one an neutrophils. strong mammalian detection oxidize Highest MPO oxidases of Each antioxidant third in cellular (MPO) of and hydrogen and This nonradical The 144kD. dimmer of is a 2 Throughput format plant 0 activity is kit 7-dihydroxyphenoxazine the varity 2 enzyme present ) and Excessive enzyme is unique and is to (3) damage MPO a enzyme sensitive peroxide cells The . superoxide is water of In is non-mammalian highly (4) oxidant,HOCl. composed . seen is to eukarotic aromatic in found hemoprotein has (1-3) unique neutrophils production the through and . cationic that in assay in Catalase been in liver azurophilic eukaryotic dismutase oxygen. PMN’s. however of catalyses compounds cells, and isolated that oxidation a HOCl glycosolated heavy and of aerobic consists activity catalase kidney, utilizes (8,12) (8-11) (ADHP, these MPO Catalase DetectionKit Description Catalase monocytes. in reactions. is cells granules . (53kD) . H . from the that the of to cells utilizes radicals while 2 varies a (5-6) 0 of proteins, is in decomposition most CF2980 P/N : give 2 it 530/590 non various .com oxidative in role .com . an two concentrated hemoprotein can and is containing lowest Accumulation votre source d’innovations of - substrate greatly end powerfull H ubiquitously However, oxidize fluorescent dimers can light 2 polymorphonuclear 0 DNA sources, 2
nm), product produced activity cause (15kD) from and chloride the linked bactericidal radicals to monocytes of that in oxidative subunit.
by has via ions the for a a The Catalase activitydetectedusingthekit. of at well temperature. catalase room 500 tests Qty reaction and temperature. the in added incubated inhibitor). Ex. :530nmEm.590nm. Reaction MPO contained reaction 1X reaction standard to Biochem. Pharmaco., J. Biochem., Biochem. Pharmaco., Proc. Nat. J Neural References : cocktail individual incubated Next Next at 20 room Buffer. µM curve 100 50 Transm Suppl23:55–72.(1987) CA7280 P/N : (RC) H Acad. Sci.U.S.A.,85,4934–4938(1988) μL and well 2 µL for 0 The 79, 1297–1299(1976)
was 2 Ex. /Em. :530-571590-600nm of
was (final) of temperature another of MPO reaction Reaction serially a prepared 96 per 27, 1995–2000(1978) 17, 1285–1297.l(1968) standard well 10 well diluted was minutes cocktail black in as and incubated the described and in the plates. dark. 1X in was indicated 50 the Reaction added Data μL for The (without dark 500 tests Qty
of 30 plate collected amounts RC minutes at to Buffer. room each EPO was was Reactive Oxygen Species (ROS)
■ Cis-Parinaric Acid (CPA) Measurement of lipid peroxidation
Cis-parinaric acid (CPA) is a fluorescent poly-unsaturated fatty acid used as a probe to directly monitor lipid peroxidation. Fluorescence measurement is using 318 nm excitation and 420 nm emission filters.
Description P/N : Qty Cis-Parinaric Acid FP-46900A 10 mg
■ SOD Assay Kit with colorimetric substrate WST-1
» Measures 100% inhibition by SOD » pH independent IC50 determination » Convenient 96-well microplate colorimetric assay » Low-background noise measurement
Superoxide dismutase (SOD), which catalyzes the Apoptosis Assays - dismutation of the superoxide anion (O2 •) into hydrogen peroxide and molecular oxygen, is one of the most important antioxidative enzymes. In order to determine the SOD activity, several direct and indirect methods have been developed. Among these methods, an indirect method using nitroblue tetrazolium (NBT) is commonly used due to its convenience and ease of use. However, there are several disadvantages to the NBT method, such as poor water solubility of the formazan dye and the interaction with the reduced form of xanthine oxidase.
SOD Assay Kit-WST allows very convenient SOD assaying by utilizing highly water-soluble tetrazolium salt, WST-1 that produces a water-soluble - formazan dye upon reduction with a superoxide anion. The rate of the reduction with O2• are linearly related to the xanthine oxidase (XO) activity, and
is inhibited by SOD, as shown in the Figure above. Therefore, the IC50 (50% inhibition activity of SOD or SOD-like materials) can be determined by measuring the decrease in the color development at 440 nm.
Description P/N : Qty SOD Assay Kit S07411 100 tests S07410 500 tests Related products : WST-1 as stand alone product F98883 100 mg NADPH Q91330 40-wells
■ Malachite green isothiocyanate Localized production of hydroxyl radicals by amine-reactive probe
This non-fluorescent photosensitizer probe (λexc./em. 628 nm/none) can be conjugated to specific antibodies. Enzymes and other proteins within ~10 Å of the binding site of the malachite green–labeled antibody can then be selectively destroyed by production of hydroxyl radicals upon irradiation with long-wavelength light.
References : Stresser DM. et al., Drug Metab Dispos 30, 845-52 (2002) Tolosa L. et al., Lifetime-based sensing of glucose using energy transfer with a long lifetime donor.". Anal Biochem 250, 102-108 (1997) Beermann A. et al., "Chromophore-assisted laser inactivation of cellular proteins." Methods Cell Biol 44, 715-732 (1994)
Description P/N : Qty Malachite green isothiocyanate FP-98782A 10 mg
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 57 58 Apoptosis Assays Aldehyde oxidation study biogenic aminesincludingtyramine. Furthermore disease andvariousotherinflammatorydiseases. cancer, agingandatherosclerosis. production SSAO’s vascular systemandcirculatesinplasma. Semicarbazide-sensitive Fluorescent Semicarbazide-Sensitive ■ SSAODetectionKit Monoamine Oxidase Description specific serotonin, Monoamine ■ MonoamineOxidase Localized and benzylamineisinhibitedbydeprenylpargyline. properties. MAO-A which aretheproductsoftwodistinctgenes. SSAO DetectionKit Description H Benzylamine +O
Substrate +O2H2OMAO-A/B H 2 2 O O 2 2 + Detectionreagent(non-fluorescent)+Peroxidase Prxds + Detectionreagent(non-fluorescent)+Peroxidase
organ and functional in norepinephrine, MAO-A of oxidase the B by-products hydrogen and/or exhibit outer
role A&B DetectionKit acts 2 +H (MAO) within mitochondrial has different preferentially peroxide. 2 amine O +SSAO of epinephrine been a SSAO is specific a specificities oxidase suggested flavin-containing Reports deamination, membrane, on cell Amine OxidaseDetectionKit A&B DetectionKit and
serotonin (SSAO) type. of to dopamine to elevated be In substrates involved these is such addition and enzyme a common to norepinephrine, levels as enzymes yield in: to formaldehyde P/N : CA7290 and CA7310 P/N :
.com.com that apoptosis, their of the votre source d’innovations name inhibitor SSAO catalyses are corresponding role for found in and have atherogenesis, regulating selectivities. a and the widely is Benzaldehyde +NH
aldehyde +NH throughout been inhibited methylglyoxal, oxidation
aldehydes. distributed reported neurotransmitters, Extensive by cell 500 tests Qty Qty 500 tests the of clorgyline. adhesion, in a body. 3 MAO +N have enzyme variety congestive 2 studies 3 O +N Often exists been Resorufin (fluorescent) 2 Resorufin (fluorescent) Ex. /Em. :530-570590-600nm MAO-B leucocyte of in 2 these O amine-containing only nature. 2 in proposed have heart two acts enzymes one isoforms, been trafficking, failure, In preferentially form man to preformed be are diabetes of this namely involved the also glucose neurotransmitters enzyme enzyme on involved to mellitus, MAO-A 2-phenylethylamine in transport characterize is pathogenesis is present in
and present alzheimer’s processing and MAO-B, such in local their in the as of a Nitric Oxide (NO) assay
■ DAF-2 diacetate
» Cell permeable » No wash homogenous assay » Real time detection of NOS activity » – NO and NO2 detection limit : ~5 nM
NO scavenging can be measured by monitoring 4,5-diaminofluorescein (DAF-2). DAF-2, as a specific NO indicator, selectively traps NO between two amino groups in its molecule, and yields triazolofluorescein, which emits green fluorescence when excited at 490–515 nm. The diacetate form is cell-permeable derivative of DAF-2. DAF-2 diacetate can be used to detect NOS activity in cell culture or tissue sections. This reagent is not species specific and can also be used to detect NOS activity in plant cells (but not in barley aleurone cells1). References : 1) Journal of Experimental Botany 2006 57(3):463-470
Description P/N : Qty DAF-2 diacetate S03720 100 µg NOS Detection kit CA7150 125 µg (2.22 mg/ml) CA7151 250 µg Apoptosis Assays Related products : SNAP, photoactivable NO donor FP-71646A 25 mg Spermine NONOate, pH controlled NO donor FP-M16259 10 mg Carboxy-PTIO potassium salt, specific NO scavenger 199500 5 mg L-NMMA, NO synthase inhibitor FP-85524A 50 mg
■ DAF-FM
DAF-FM is important reagent for quantitating low concentrations of nitric oxide in solution. This compound is essentially nonfluorescent until it reacts with NO to form a fluorescent benzotrizole (495/515 nm). The diacetate form is membrane permeant and is deacetylated by intracellular esterases to DAF-FM.
» Fluorescence independent of pH above pH 5.5 » Significantly more photostable than that of DAF-2 » – NO and NO2 detection limit : ~3 nM versus Description P/N : Qty DAF-FM FP-R1227A 1 mg DAF-FM diacetate FP-R1228A 1 mg
■ 2,3-Diaminonaphthalene 2,3-Diaminonaphthalene reacts with nitrosonium, which is formed from NO, to form the fluorescent dye 1 H-naphthotriazole (365/415 nm).1,2 Using this method, 10 nM to 10 µM of nitrite (NO2-) can be detected and the detection is compatible with 96-well format.3 References : Luminescence 14, 283 (1999) Methods Enzymol. 268, 105(1996) Anal. Biochem.214, 11(1993) Description P/N : Qty 2,3-Diaminonaphthalene FP-04832F 100 mg
■ NBD Methylhydrazine for nitrite assay
– NBD methylhydrazine (N-methyl-4-hydrazino-7-nitrobenzofurazan) reacts with NO2 in the presence of mineral acids leads to formation of fluorescent products (468/537 nm). NBD methylhydrazine has been used to quantitate nitrite in waters.
Reference : Anal Chem 71, 3003-3007 (1999) Description P/N : Qty NBD Methylhydrazine FP-R1315A 50 mg votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 59 60 Biochemistry Assays Fluorescent DNAAssay DNA Hoechst 33258,forhighconcentrationDNA DNA Related products: EvaGreen 20000XinDMSO EvaGreen 20XinPBS Description to PCRtubescontaining1.25µlofEvaGreen(20Xconcentrate),andwaterwasaddedmakeafinalvolume25µl. Relationships betweenfluorescenceintensityandtheamountofDNA Wang » » » ■ EvaGreen-DNA » the tube. Other applicationoftheEvaGreeninBioScienceInnovationcatalog. The 303-305 (2006)):Figure Standard filterset:λ Sample size:50µl Linear responseontherangeof1–100ng Requires smallamountsofDNA dosagebyUVwithonly5µlsample,usingIMAplates fromlambdaphage,asstandardforDNA intensity W. et al. has of fluorescence (relativeunits) used A the andB. exc. EvaGreen /λ A stronglinearrelationshipisobservedwhentheamountofDNA em. quantificationinsolution : 490/520nm for DNA anddye quantification
quantification quantification on 25 is FP-61248A UP947860 CA6770 BI1790 P/N : see descriptionpage72 .com µl
.com proportional to sample votre source d’innovations pertube:Triplicate samplesof_DNA with linear the response total islessthan100ng(Fig. amount from 100 mg 100 µg 1 ml(200000assays) 5 x1ml(1000assays) Qty 1 ng of to DNA intherangeof1–1000ng(A)or0–100ng,(B)wereadded 100
per ng (Analytical A), andthisisveryreproducible(Fig.B). tube rather Biochemistry, than the Volume concentration of 356, Issue 2, DNA Pages
in Protein assays
Interchim proposes 3 methods to assay proteins in microplates :
Colorimetric/BCA method : Colorimetric/Bradford method : Fluorescent/Epicocconone : BC Assay & MicroBC Assay CooAssays () LavaPep method
0.5 µg/ml (MicroBC Assay) 1-25 µg/ml (max sens.protocol) Sensitivity : 50-160 pg/ml peptide, proteins 20 µg/ml (BCA assay) 50-1500 µg/ml (Broad range protocol) 1-250 µg/ml (MicroBC Assay) 1-25 µg/ml (max sens.protocol) Linear range : 100 ng/ml-160 µg/ml 20-2000 µg/ml (BCA assay) 50-1500 µg/ml (Broad range protocol) Reading 562 nm 570-610 nm 405-500/560-610 nm The most reliable method Ultimate sensitivity and linearity Biochemistry Assays Compatible with most detergents, bases, Peptide dosage Quick (1-10 min) Comments : advantages DNA, lipids Biodegradable Compatible with reducers Broader linearity Robust, amenable to N-term Lowest variations between proteins sequencing, MS and functionnal assays Limited compatibility Comments : limitations Need 37°C incubation or longer time Limited compatibility (reducers, acids, some detergents)
Description P/N : Qty BC Assay protein determination kit UP4080A 1 kit-1 L Bicinchoninic acid based method-590nm. UP4080B 1 kit-250 ml Contains : 1L reagent sufficient for 500/5000 tests (tube/µplate), and 10 x 1 ml BSA standard 2 mg/ml MicroBC Assay protein determination kit UP75860A 1 kit-1 L Version of BC Assay with sensitivity 0.5 µg/ml. UP75860B 1 kit-50 ml Contains : 1L reagent sufficient for 500/3400 tests, and 10 x 1 ml BSA standard 2 mg/ml Coo Assay protein determination kit UPF86400 1 kit-1 L Order together the following reagent to Modified Bradford (Coomassie based method). UPF86401 1 kit-250 ml render these protein assays compatible Contains : 1L reagent sufficient for 500/4000 tests (tube/µplate), and 10 x 1 ml BSA standard 2 mg/ml with any interfering substance !
LavaPep peptide & proteins assay CH4191 1 Kit Protein Preparation kit Epicocconone based method. (up to 2000 tests) Réf. : R5594A 500 ml Contains : Dye concentrate 10X, Buffer concentrate 10X Biochemistry tests for biologicals samples Interchim provides a whole range of biochemistry assays for clinical chemistry (Blood, Urine, CerobroSpinal fluids,...) as well as for other samples (food, soil, water...) analysed in agro-alimentary industry or environement study. Here is a short selection of assay kits.
Description P/N : Qty Glucose assay BD1850 1 Kit (120 ml) Hexokinase/G6PDH based method. Reading at 500 nm. Linear to 400 mg/dL. For serum, plasma or urine. 5 min procedure. Creatinine assay BP9991 1 Kit (30 ml) Enzymatic method. Reading at 546 nm. Linear to 30 mg/dL. For serum, plasma and urine samples. β-Hydroxybutyrate assay AL2230 1 Kit (60 ml) Enzymtic method. Reading at 505 nm. Linear to 4.5 nmol/L. For serum or plasma. Glucose assay U67120 1 Kit (40 tests) Hexokinase/G6PDH based method. Reading at 340 nm. Linear 1-80µg/ml. For food stuff and bewerage. Glycerol assay R51065A 1 Kit (4 x 10 tests) Enzymatic method. Reading at 330/334/365 nm. One component. For food stuff and bewerage. Cholesterol assay R5756A 1 Kit (4 x 10 tests) Enzymatic CHOD/PAP method. Reading at 546 nm. For food stuff and bewerage. Contact us for other analytes in biological samples (Calcium, Bilirubin, HDL, Urea, ... / Maltose, Fructose, Starch, Malate, Lactate,...). votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 61 62 Immunological Assays UptiLight ELISA Also available: UptiLight ELISA ■ UptiLightELISA ELISA -Luminescent (ECL) detection AssaysImmuno A Description » » » » » » immunoassays. Increase 10timesthenetsignalofyourusualluminolreagent,forultra-sensitiveHRP (S/N) ratios. background Interchim alkaline ImmunoAssays in significantly more flexibleandquickertocalibrateusingfewlabeledstreptavidinsformanydifferent assays luminescent We range, sensitivityandnotablyallowformultiplexedanalysis. Signal Sensitivity comparisonwithUptiLightandcompetitors respective integration PEF, IgG(H+L) found withUptiLightreagent. (S/N) costeffective reagentforroutineanalysis,when onlypicomoledetectionisrequired. screening Easy touse:mix,incubate5min,read Reagent stable>18monthsat4°C Great forHTSapplications Signal stableupto1hour Sensitivity inthepicoandfemtorange High signalwithminimalbackground finally BEF, for to each noise - phosphatase AEA, supplier. time,
HRP provides provide the is tested experiments. in substrates,
HRP HRP and after (#UP446330), lower. sensitivity ELISA have Luminescence PBP. HighSensitivityChemiluminescent Substrate. UltraSensitivityChemiluminescentSubstrate. biotin/streptavidin a HRP a 5 As
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offering your by techniques, sensitivity, Noise with competitors anti to usual 0,1 Mouse ratios Linked get their -some was include sec an reagent, and Signal that amplification crucial Immunosorbent based not fluorescent 200 ml 300 ml 120 ml 60 ml Qty Photostability 30 minpreincubationperiod. was improves to but only points Noise recorded the highly of that can increase the dynamic the increase can that immunoreagents the Assay), with of page 18. See alsochemiluminescent kitsfor sensitive Technical tip signal. achievable detectionlimit(highestsensitivity). or detection of range dynamic extended requiring assays quantitative for recommended most is microplates). 425 of peroxidase substrates the principle sensitivity) overcoming and The light 0,1 glow by-products qualitative luminol nm especially emission effective sec Additionnally, is colorimetric consists (in integration of then upon (incl. the The was classical tubes, with chemiluminescent of assays performances recorded use the using of introduced OPD, the the of UptiLight substrate. or time biotin chemical luminescent insoluble substrates, generation peroxidase requiring in TMB, by after as wells Assay #JQ6760 label luminometer (and USLuminescence The reaction a chromogenic a DAB). convenient substrates of is 1, of substrate, the emission first, (HRP) generaly light but 5, ELISA from best The 10 the also by at or
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ELISA - Luminescent detection (ECL)
■ Luminometric Alkaline Phosphatase Assay Kit
Alkaline Phosphatase Assay Kit uses a proprietary luminogenic phosphatase substrate, to quantify alkaline phosphatase activity immobilized on surfaces, i.e. ELISA, as well as in solutions (i.e. cell extracts, live cells; see page 17). This proprietary phosphatase substrate generates a luminescent product (Em (nm) 560 nm) that produces strong luminescence upon interaction with phosphatase. The kit provides all the essential components with our optimized ‘mix and read’ assay protocol that is compatible with HTS liquid handling instruments. It has extremely high sensitivity, and can be used for the assays that require demanding sensitivity.
Description P/N : Qty Chemiluminescent AP assay kit JQ6760 100 tests See description in enzyme detection/Alkaline Phosphatase section, page 18.
Related products : Selected HRP labeled secondary antibodies (anti Igs) Host a Specificity Biotin labeled Qty HRP labeled Qty *Anti Human antibodies
Gt IgG Anti-Human IgG , min X Bov, Hrs, Ms sr prot. UP892650 0.5 mg UP783493 1 mg Immunological Assays Gt IgG Anti-Human IgG, Fc fragment specific, min X Bov, Hrs, Ms sr prot. 722640 1 mg 802020 1 ml Ms Mab Anti-Human Kappa Light Chain UPB91830 0.5 mg UPB91850 0.5 mg Rt Mab Anti-Human Lambda Light Chain UPB91870 0.5 mg UPB91890 0.5 mg Rt Mab Anti-Human IgG Gamma Chain UPB92190 0.5 mg UPB92210 0.5 mg Rt Mab Anti-Human IgM Mu Chain UPB91910 0.5 mg UPB91930 0.5 mg Rt Mab Anti-Human IgD Delta Chain UPB91990 0.5 mg UPB92010 0.5 mg Rt Mab Anti-Human IgE Epsilon Chain UPB92030 0.5 mg UPB92050 0.5 mg Rt Mab Anti-Human IgA Alpha1&2 Chain UPB92150 0.5 mg UPB92170 0.5 mg *Anti Mouse antibodies Dk IgG Anti-Mouse IgG(H+L), min X Bov, Ck, Gt, GP, Sy Hms, Hrs, Hu & Rb, Shp sr prot. 946220 0.5 ml UP973171 0.5 mg Gt IgG Anti-Mouse IgG(H+L), min X Hu, Bov, Hrs, Rb, Sw sr prot. 668610 1.5 ml UP215731 1 mg Rb IgG Anti-Mouse IgG(H+L), min X Hu sr prot. 794090 1 ml 794560 1 ml Anti Rabbit antibodies Dk IgG Anti-Rabbit IgG(H+L), minX Bov, Ck, Gt, GP, Sy Hms, Hrs, Hu & Ms, Rt, Shp sr prot. UP944471 0.5 mg 338370 0.5 ml Gt IgG Anti-Rabbit IgG(H+L), min X Hu sr prot. 812230 1.5 ml UP687714 1 mg Gt IgG Anti-Rabbit IgG(H+L), min X Ms, Rt sr prot. UP668621 1 mg UP669631 1 mg *Anti Rat antibodies Gt IgG Anti-Rat IgG(H+L) 740470 2 ml UP399892 1 mg Secondary antibodies are available with different preadsorbtions, formats (Fab'2), raised in 6 hosts, and against 22 target Ig species. Please inquire. HRP (strep)avidin reagents Description P/N : Qty HRP labeled Streptavidin UP395888 1 mg HRP labeled Avidin 35445A 2 mg HRP labeled Neutralized Avidin UP36570A 1 mg
Other Colorimetric & Fluorimetric Enzy- matic Substrates for Microplates assays
Peroxidase (HRP) and Alkaline phosphatase (AP) enzymatique labels are widely used in various biological assays including ELISAs, as well as in immunohistochemical techniques and Western blot analyses. Following are effective ready to use substrates for colorimetric and fluorometric detections in EIA.
Following is a selection of conventional and unique formulations of standard enzymatic substrates to fulfill the requirements of your microplates assays. Below are research catalog quantities, please as for HTS and bulk needs. votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 63 64 Immunological Assays Contains ADHP See assaykits#JQ6730(Blue-MUP),#JQ6740Green-Red)insection "Apoptosis" page17. ■ Fluorimetric Description ADHP ■ molar assays ina384-wellformat. use Substrates for Microplates assays Colorimetric &Fluorimetric Enzymatic Other Recommended inparticularfordiagnostickits TMB, "Standard"solution » » » Highest sensitivity;Stability>1yearat+4°C. 100% waterbasedformulationtomaximizethesafety(noregulationconcerns). (does notcontainDMF, DMSO) Non-hazardous, non-volatile,non-organic,non-toxic TMB, "Aqueous"solution Includes areddyethatdonotinterferewithreactionnorfinalreading.Highsensitivity;Stability>1yearat+4°C reagent distribution. Recommended forroutineassays,tobettercontrolfailurein TMB, "Check"solution -Stability>24monthsat+4°C -Highestsensitivity The originalformulation,idealforclassicapplications Description 3,3’,5,5’-tetramethylbenzidine (TMB),hydrogeneperoxide(H Solutions ■ UptimaTMBchromogenicsubstrateforHRP ADHP Ready-to-use Reproducible lots Highest sensitivity convenient HRP
detection provides ADHP ELISA are HRP substrateandPeroxide optimized format higher sensitivity. assaykit– ELISA sensitivity for AP chromogenic HTS ADHP It provides ELISA applications. based
in Assay Kit Assay buffer (50ml) ELISA the substrates
Assay kits
that reagents The chromogenic fluorescent for to peroxidase, perform HS6241 P/N : UPS08181 UP664780 UPS08180 UPS08170 P/N : .com.com UPS08183 UPS08182 UPS08171 UP664782 UP664781 substrates. 2 signal O votre source d’innovations 2 500 ), andproprietarycatalyzingstabilizingagents. designed is ELISA read The
assays at kit for 590 ELISA provides ELISA nm in a (λ 500 tests Qty 200 ml 200 ml 100 ml 100 ml Qty 1 L 500 ml 500 ml 1 L 500 ml 96-well
exc the techniques, \λ em substrate
= format. 530-571 manual and The / the 590-600 protocol or stabilized automatic can nm) Peroxide readily and systems. achieves be buffer modified They down in ready-to contain to femto run Other Colorimetric & Fluorimetric Enzyma- tic Substrates for Microplates assays
■ Substrates for horseradish peroxidase HRP substrates P/N : (a) Qty Format (c) Type (b) Comment Each tab contains 15 mg of OPD for quick and easy OPD tablets of 15 mg 270861 50 tabs Tabs Chromo. preparation of substrate solution OPD, Ultrapure powder 02673F 25 g Pow. Chromo. (o-Phenylene DiAmine) CAS: [95-54-5] ; MW : 108.1 02673G 50 g ABTS, Liquid substrate UP732550 100 ml Soln Chromo. Ready-to-use solution Each tab contains 10 mg of ABTS for quick and easy ABTS tablets 42387C 50 tablets Tabs Chromo. preparation of substrate solution. ABTS, Ultra pure grade UP423876 1 g Pow. Chromo. CAS : [30931-67-0] ; MW : 548.7 UP423877 10 g TMB solution for ELISA UP66478 Soln Chromo. Highest sensitivity. See description page 64. TMB, Ultrapure powder UP15426D 1 g Pow. Chromo. (tetramethylBenzidine) CAS : [64285-73-0] ; MW : 331.3 UP15426E 5 g Immunological Assays Ready-to-use solution less toxic for manipulation DAB solution (50X) UP732320 500 ml Soln Chromo. Supplied as a 50x concentrate with a 10x dilution buffer. Each tab contains 5 mg of DAB for quick and easy preparation DAB tablets of 5 mg UP732310 50 tabs Tabs Chromo. of substrate solution DAB, Ultrapure powder UP01012G 5 g Pow. Chromo. (3.3'-DiAminoBenzidine) CAS : [868272-85-9] ; MW : 360.1 UP01012H 10 g ADHP HRP Assay Kit. HS6241 500 tests Kit Fluo. Page 64. ADHP, Pure Grade powder FP-39423A 5 mg Pow. Fluo. Highly and stable fluorigenic substrate
CAS : [119171-73-2] ; MW : 257.2 FP-39423B 25 mg λabs/λem. : 563/587 nm (10-Acetyl-3,7-Dihydroxyphenoxazine) Resorufin, Pure Grade FP-95432B 100 mg Pow. Fluo CAS:[635-78-9]; MW : 213.04
Luminol, Pure Grade powder FP-57578A Pow. Lum. λabs/λem. : 355/413 nm ; EC : 7650 l/mol x cm CAS : [521-31-3] ; MW : 177.2 Luminol, Na salt FP-CA9611 2.5 g Pow. Lum. See also UptiLight page 62. CAS : [20666-12-0] ; MW : 199.1 (a) all products are available as bulk formats. Please inquire. (b)Chromo. : Chromogenic ; Fluo. : Fluorigenic ; Lum. : Lumigenic (c)Pow; : powder ; Soln : Solution mono-component or bi-component (2 Soln) See also substrates for reporter assays (B-galactosidase,...) page 16. ■ Substrates for Alkaline Phosphate HRP substrates P/N : (a) Qty Format (c) Type (b) Comment pNPP ELISA solution BP7080 500 tests Kit Contains Assay buffer, wash buffer, stop solution and AP II Ab. pNPP tabs of 30 mg UP732500 100 tabs Tabs Chromo Each tablet contains 30 mg of pNPP for quick and easy UP732501 1000 tabs preparation of substrate solution. pNPP tabs of 5 mg UP89562G 100 tabs Tabs Chromo Each tablet contains 5 mg of pNPP for quick and easy UP89562F 1000 tabs preparation of substrate solution. pNPP, Ultrapure powder UP89562C 25 g Pow. Chromo. (p-NitroPhenylPhosphate). CAS : [4264-83-9] ; MW : 301.3 UP89562D 100 g MUP Na salt, Ultra Pure grade FP-30045A 100 mg Pow. Fluo. (MethylUmbelliferyl Phosphate). CAS : [22919-26-2] ; MW : 277.1 FP-30045B 500 mg Used for detecting phosphatases in solution. FP-30045C 5 g Maximum fluorescence at pH value of >10. FP-30045D 10 g Maximum fluorescence above pH 7.0, for continuous assays. MUP Plus, Na salt FP-JQ6710 25 mg Pow. Fluo. Also used for the assays that require acidic pH such as acid phosphatases. See description page 18. MUP Free acid FP-24119A 100 mg Pow Fluo See also MUP based AP assay kit #JQ6730 page 17. CAS : [3368-04-5] ; MW : 256.1 FDP FP-72573A 5 mg Pow. Fluo. (Fluorescein DiPhosphate). CAS : [217305-49-2] ; MW : 560.4 See also FDP based AP assay #JQ6740 page 17. FDP ELISA kit HT0790 1000 tests Kit Fluo. Contains Assay buffer, wash buffer, stop solution and AP II Ab. votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 65 66 Immunological Assays selection ofgreatFluoProbesdyes,the782,682,647HandR-PEbeingmostusefulinmicroplatesassays: Streptavidin ■ Streptavidinconjugates Description ELISA (IIAbs, Avidin) Fluorescent reagents for secondary FluoProbes FluoProbes Other labeled(strept)avidinconjugatesareavailablefromInterchim Related products FluoProbes FluoProbes R-Phycoerythrin FluoProbes FluoProbes FluoProbes 2005, 174:3359-3368. Rowell References : For otherwavelengths,pleasecontact usorseeinInterchimBioScienceInnovationscatalog. E. et ® ® ® ® ® ® ® 488 547H 565A 594A 647H 682 782 al., conjugates Opposing λ λ abs. em. 200 200 Roles are λ 493 557 565 563 601 653 690 783 exc. second for 305 (nm) 295 the Cyclin-Dependent step 355 345 reagents 405 395 800 518 572 576 592 627 675 709 λ em. Kinase (nm) for 445 455 staining Inhibitor .com.com 480 471 with votre source d’innovations p27 P/N : FP-IS1810 FP-BA2221 FP-CA5570 FP-77776A FP-CA5610 FP-CA5620 FP-CA5640 FP-BE8050 biotinylated kip1 505 495
in Wavelength (nm) the Control 530 520 antibodies. of CD4 555 545 Qty 1 mg 1 mg 1 mg 1 mg 1 mg 1 mg 1 mg 1 mg +
T
Cell Interchim 580 571 Proliferation 605 595 covers See Fluoprobesdyesdescriptionspage 67. and a 645 655 Effector wide 695 705 range Function, of 755 745 The wavelengths.
Journal 805 800 of Immunology, Here is a Fluorescent secondary reagents for ELISA (II Abs, Avidin) ■ Secondary Antibodies conjugates
We are offering a range of high quality secondary antibodies conjugated with our fluorophores called FluoProbes. Showing superior fluorescence properties, these fluorophores are an excellent alternative to the conventional dyes such Cyanines and other dyes. Following are selected fluorophores compatible using common exitation sources and standard filters set such as FITC/Cy2, TRITC/Cy3, TR/Cy5, and InfraRed, .
Benefits of FluoProbes-antibody conjugates
» High brightness : FluoProbes® dyes show an enhanced fluorescence compared to other similar conjugates (very high Signal/Noise ratio) » High photostability : Our FluoProbes dies are more photostable than most other conjugates allowing longer reading/scaning. » Color choice : FluoProbes® dyes range in color from green to infra-red. » Ready to use solution : no dissolution step and risk of contamination. » High species-specificity : The secondary antibodies were selected to have highest Immunological Assays specificities. It is then possible to have very species-specific secondary antibodies conjugated with the best fluorochromes.
Description λexc. (nm) λem. (nm) P/N : Qty Goat anti-Mouse IgG (H+L) - FluoProbes® 782 783 800 FP-BW7970 1 mg - FluoProbes® 682 690 709 FP-BE7250 1 mg - FluoProbes® 647H Min x Rat, Hu, Bov, Hrs, Rb sr Prot 653 675 FP-SC4000 1 mg - FluoProbes® 547H Min x Rat, Hu, Bov, Hrs, Rb sr Prot 557 572 FP-SB4000 1 mg - FluoProbes® 488 Min x Rat, Hu, Bov, Hrs, Rb sr Prot 493 518 FP-SA4000 1 mg Goat anti-Rabbit IgG (H+L) - FluoProbes® 782 783 800 FP-BW7980 1 mg - FluoProbes® 682 690 709 FP-BF1690 1 mg - FluoProbes® 647H Min X Bov,Ck,Gt,GP,Hms,Hrs,Hu,Ms,Rat,Shp Sr Prot 653 675 FP-SC5000 1 mg - FluoProbes® 547H Min X Bov,Ck,Gt,GP,Hms,Hrs,Hu,Ms,Rat,Shp Sr Prot 557 572 FP-SB5000 1 mg - FluoProbes® 488 Min X Bov,Ck,Gt,GP,Hms,Hrs,Hu,Ms,Rat,Shp Sr Prot 493 518 FP-SA5000 1 mg See all our secondary antibodies items in our catalog Biosciences Innovations.
Related products : PrimAb™ : a large collection of primary antibodies. Search our web PrimAbs engine tool at http://www.interchim.com/interchim/PrimAb/search.cfm. Research areas include : CellSignaling (ionic, cytokines, hormones), Apotosis/cell cycle, DNA replications/transcription/Repair, post-translational modifications, Cell adhesion, Enzymes, Membrane study, Lipids, Metabolism, Angio/Histogenesis, Infectious agents, biomarkers for Cancer/Hypoxia, Cardiology, Neurosciences, Drug and resistance, Allergens.
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 67 68 Immunological Assays Agents ImmunoAssay -BuffersSaturating & SeaBlock, serumfreein SeaBlock, serumfreeinPBS SeaBlock (standard,excelsasablockerinELISA) TBS PBS tablets BSA TBS tablets ■ ReadytouseBuffersforImmunoassays PBS with PBS TBS powderpack PBS powderpack Description ■ StandardBuffersforImmunoassays Note :afullrangeofother gradesofBSA Highly pureandpackaged insealedampulsunderargontoincrease theaccuracyofimmunoassays. Tween BSA Prionex Water (KF) :<12%;Viscosity :35-45mpa CAS [9000-70-8];Bloomnumber:240-270 ;pH(28°C):4.5-5.5 Gelatin A Non-fat Milk positively chargednylonorPVDFmembranesinnucleicacidproteinblottingapplications. An economicstandardblockerbasedoncasein,optimizedfor BioBlock membraneblockingagent(inPBS) Polymerised BSA Polymerised BSA,30%solution BSA Using thissolution,forgetthehassleofweightinganddissolving biotechnologies, includingimmuno-saturations. Our standardgradeandeconomicBSA,ubiquitousformost Description ■ OthersaturatingagentsandBufferscomponents Description (whenBSA,caseinandotheragents » » » ■ SeaBlocksaturatingagent(nonmammalianserum) Ab dilutedinthissolutioncanbestoredforupto18monthsat+4°C. TBS with TBS withBSA TBS withNon-FatPowderedMilk3% Description These poweredblendssaveyourtimeandarehighproteomicsgradeforoptimalresultsinimmunodetections. BSA Tween Antibody Diluent(ReadytoUse) PBS with PBS withBSA PBS withNon-FatPowderedMilk3% popularbloquer andGlutaraldehydeactivated tosaturatehighbindingsurfaces, Excellent Lower background antibodies) withimmunoreagents(i.e. mammalian Non-mammalian naturepreventsinteractions alternative,stronglyreducesunspecificbinding ofproteintoplasticmicroplatewalls powder(noagregates!).Savetimeandmoney! 30%solution powder (Tris Buffered Saline) (Tris Buffered Saline) ® ® 20,20%solution,oxidant free 20,pure ® , 10%sterilesolution Tween Tween Tween (1 (1 1% 1% Tab.=100 mlof1Xsolution) Tab.makes 100mlof1Xsolution) ® improveseveraldetectionsystems ® ® 0.05% 0.05% , pH7.5 (makes 20L) (makes 10L) TBS , 20Xsolution , 20Xsolution andotheralbuminsare alsoavailable. blockers). aregoodbutnotexcellentorevenpoor GS4170 GS4160 P/N : HH6690 GS4250 GS4190 GS4180 GS4200 N13810 N13761 N14580 P/N : GS3660 307150 UP74004A UP68723A UPAP1380 UPAP1370 UP40301A P/N : 901770 N13361 N13650 UP900101 UP900100 UPQ84170 UPQ84171 N13360 768701 N13660 BJ1440 P/N : UPQ84170 .com.com UP158740 15874A votre source d’innovations Amine polystyrene 500 ml 1 L 4 L Qty 100 100 1 pack 1 pack 5 pk(22g/1L) 5 pk(42g/1L) Qty 125 ml 5 pk(10.4g/1L) 5 pk(19.8g/1L) 5 pk(39.8g/1L) 5 pk(12.5g/1L) 500 ml 500 ml 500 ml Qty 100 ml 500 g 1 L 500 ml 50 ml 1 kg 500 g 100 g 500 g 1 L 50 ml Qty 100 g 5 x10ml 1 L (in
Tabs Tabs TBS ) Antibody Labeling Kits
For direct conjugation of primary antibodies or your biomolecules of interest, Interchim provides a large range of biotinylation and fluorescent agents as well complete kits with desalting tools.
For general use, we recommend to use NHS-, Maleimide, or Hydrazide-activated reagents.
Also, complete biotinylation kits are proposed for non experienced investigators, as well as for convenient labeling (spin format) while avoiding to buy separate reagents.
■ Microspin biotinylation labeling kits-NH2 and -SH Efficient and convenient biotinylation reagent
» Quick : only 1 hour (/NH2) or 3 hours (/SH) to get conjugates » Easy : all processes in a single filtration tube » Reliable : high recovery of conjugates, even for 500 µg of IgG !
Biotin Labeling Kits are primarily used for the preparation of biotin-labeled IgG for Immunological Assays immunoassays. We offer kits with a very convenient format : spin filters where reaction and washes take place, that are available with 2 coupling strategies, and for 2 sample sizes (50-200 μg IgG, or ca 1 mg).
The kit BG767 biotinylates on amines, the most standard strategy. It uses a succinimidyl ester activated biotin, and contains all necessary reagents for labeling 3 samples of IgG antibody (10 μg to 200 μg). It can also be used to biotinylate any protein greater than MW 50 000 Da. The labeling process is simple.
Just add the NH2-reactive biotin to IgG solution on a filter membrane, and incubate at 37 ºC for 10 min. On the average, 5 to 8 biotin molecules conjugate to each IgG molecule. Exceeding biotin molecules can be removed using a Filtration tubes included in this kit.
The kit BT3591 biotinylates on sulfhydryls to obtain oriented and defined biotinylation. It uses a maleimide- activated biotin. Features are similar to kit BG7670, except 1/there is an additional step to create a free sulhydryl in those protein (IgG) that do not have one, without loss of affinity ; 2/maleimide incubation occurs at 37 ºC for 30 min.
Description P/N : Qty
Biotinylation kit-NH2 reactive BG7671 3 lab. of 50-250 mg BG7671 1 lab. of 10 mg Biotinylation kit-SH reactive BT3591 3 lab. of 50-250 mg Also available : biotinylation agents as stand alone sulfoNHS-lc-Biotin UP54398A 100 mg
NHS-PEO4-Biotin UPR20279A 50 mg PEO spacer confers hydrophilicity allowing to reach higher couplig ratio and to yield more bioactive and stable conjugates. It is also available with extended spacer length (PEO12 ). NHS-SS-Biotin UPS073A 100 mg cleavable spacer
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 69 70 Immunological Assays Antibody LabelingKits See Fluoprobesdyesdescriptionspage67. can especially Description FluoProbes Easy antibodydirectconjugationwiththebrilliantandphotostableFluoProbesdyes! ■ FluoProbes FluoProbes For otherwavelengths,pleasecontactusorseeinInterchimBioScienceInnovations catalog. FluoProbes FluoProbes FluoProbes FluoProbes on inquire). be labeled ® ® ® ® ® antibodies. 488 547H 647H 682 782 ®
labeling in a 1h30 kits They ® ProteinLabelingKits procedure. are use designed a succinimidyl They
for are the available ester easy-to-use of with fluorescent .com.com many and votre source d’innovations efficient of labels our λ 493 557 653 690 783 exc. FluoProbes (nm) labeling that form of a ®
protein labels. covalent 518 572 675 709 800 λ em. Following with (nm) stable molecular linkage. is a list weights P/N : FP-BE3750 FP-BZ9600 FP-BZ9610 FP-BE8280 FP-CA6070 Up of selected 100µg greater to and 1.5 than popular mg Qty 1 kit(5labelings) 1 kit(5labelings) 1 kit(5labelings) 1 kit(5labelings) 1 kit(5labelings) 25 of kD, protein ones including (others (IgG) FPlyte Microplates
Microplate innovation enables leading edge screening assays
» Long wavelength UV microplates » Limited well-to-well light cross talk » Improved cell binding efficiency
The FPlyte microplates are microplates fully compatible with commercially available plate readers, robotic sample processors and automated liquid handling systems. They are available in 2 formats, 96- and 384-wells and 3 colors :
The black plate provides the all-absorbing background needed to minimise background interference for sensitive fluorescence measurements. The opaque white plate maximises reflectivity enabling even weakly emitting luminescence assays to be routinely undertaken. In addition to optimised luminescence and fluorescence measurements the unique design offers improved cell binding efficiency and allows the convenience of direct measurements on bottom reading spectrophotometers and inverted microscopes.
FPlyte microplates are ideal for quantitative assays at excitation wavelengths in the long-wavelength UV area between 325 nm – 425 nm. They offer excellent photometric performance down to 325 nm (80%T at 325 nm, 100%T at 335 nm).
Wavelengths below 350 nm are particularly useful for a variety of fluorescence assays such as HNK-1 (λexc./em.: 325/380 nm), Thiguanine
(λexc./em.: 330/410 nm) using black FPlyte microplates, as well as many absorbance assays including Vitamin A (325 nm), retinol and retinyl acetate (325 nm), caspase (325 nm), acid phosphatase (330 nm) and hydroxyproline (335 nm) using white FPlyte microplates. Accessory tools
Description Colour P/N : Qty 96-well FPlyte Microplate, standard Black FP-BA7991 50 u Black FP-BA7990 100 u White FP-BA7950 100 u 96-well FPlyte black well, clear bottom Black FP-KT225A 50 u Black FP-KT225B 100 u 96-well FPlyte Microplate, Tissue Culture Treated, with lids Black FP-BA8010 100 u White FP-BA7970 100 u Hi Bind, 96-well FPlyte Microplate Black FP-BA8000 100 u White FP-BA7960 100 u Twister™ High Throughput Screening Pack, with lids, 96-well Black FP-BA8020 80 u White FP-BA8030 80 u 384-well FPlyte Microplate, standard Black FP-BA8170 100 u White FP-BA8130 100 u 384-well FPlyte Microplate, Tissue Culture Treated, with lids Black FP-BA8180 100 u White FP-BA8160 100 u Related products : Seal film for fluorescent assays FP-CD5130 25 m x 78 mm (1 roll) FP-CD5110 500 m x 78 mm (1 roll) FP-CD5150 125 mm x 78 mm (100 units)
Please contact us for other wavelengths of fluorescent reference standards.
Technical tip Fluorescein detection limit of an instrument Begin with a hard weigh-out of at least 4-5 mg and solubilize in 100 mM sodium borate (pH9.5). Borate is the NIST buffer used, but it can be replaced by 50 mM phosphate (pH9). The detection limits may vary slightly. To check the absorbance spectrum and back-calculate to confirm the concentration by a known extinction coefficient as calibrated against the NIST standard. Calculate the concentration of fluorescein stock solution by C (M)=(Amax / extinction coefficient) x dilution fold, the light path is 1 cm, A max at 492+-5 nm, extinction coefficient is 78,000 M-1 cm-1. Make a dilution series into the same buffer, starting in the low nM range and dilute down. For most standard curves, triplicate measurements at each concentration are sufficient. But closer to the detection limit, it is recommand to take 8 replicates. This is important for the blank sample, as well. With detection limits using Z-factor analysis, a result > or = -1 is considered to be a detectable signal. The equation for determining Z-factors is 1-((3*Sample+3*Std Dev Blank)/(Ave. read Sample - Ave. read Blank)). Description P/N : Qty New applications are under Fluorescein, standard solution, 100 nM (494/519 nm) FP-DO6630 50 ml development. Contact us for Fluorescein, standard (494/519 nm) FP-19365A 1 g your special needs. votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 71 72 Accessory tools absorbencies IMAplate equal orsuperioronmicrovolumes(5µL)! IMAplate™ (96µcuves)-white IMAplate™ StartKit Description IMAplate™ (96µcuves)-yellow IMAplate™ (96µcuves)-black (Intelligent MultiFunctional Analysis) IMAplate Technology Berthold technologiesGmbh Powered byInterchimincollaborationwith ex. do96reactionschemicalorenzymaticonauniquesupport,miniaturizeELISAswithonly5μlofreagents,highersignalsandsavingtime! » ex. assayDNA/RNA » ex. pipetteandempty96x5µL » ■ § * contains5IMAplates andReaderadapter
andyellowplatesforUV-visWhite platearerecommended forluminescencemeasurments,Black forfluorescencemeasurements, spectrometry andsamplehandling. 100 µL Applications forparallelreactions,immuno-assaysandcellassays 96 microwellplate UV,96 bottom-freemicro-cuvettearrayfor VISorIRspectroscopy: 96-channel pipettingforliquidtransfer: Méthode classique/Microplaquesstandard is L minutes /10mL a in polystyrene UV, andproteinsonjust5µL visible inonly10seconds! plate or § § § InfraRed, of 96 withyourusualmicroplatereader! "microcuves". of fluorescence It Analysis withamicroplatereader or is DR9621 P/N : DT5441 DT5431 DR9611 luminescence, .com.com used votre source d’innovations to simplify this and with : accelerate any 1 Kit* Qty 5 ea 5 ea 5 ea standard 5 µL the L seconds /1mL handling reader » » » » » andcostlyreagents >3-20x/loweredprice » Added valueofIMAplates: Technologie IMAplate of Increase sensitivity (solid-phase assays) Reduce reactionstimes Robust -highlyreproducible High analysisthroughput,evenmanually Transfer integratedtoassays ofliquid per test :savinglimitedsamples Miniaturization of microplate, reagents, while AND yielding to make signals measures at least of IMAPlate Microplates
■ How IMAPlate work?
» Loading, un-loading and washes are simplified, accelerated and reliable : samples and reagents and buffers are loaded simultaneously by capillary force (a)(precise volume), assayed, then drawn away by an absorbent paper (c) or by centrifugation. ex. 1 plate/ samples can be washed in just 10 seconds, without machine ! » Microcuves of 5 µL save up 20 fold (rare) samples and any (costly) detection reagents (ex in ELISA). » Reading (b) : the optical path is perfectly defined, and longer to those of standard microplates ! Hence detection sensitivity is superior. » The microcuves have no bottom ! Thus no parasite optical absorption take place, and you can work in UV, IR..., with superior sensitivity. You even can recover the samples (c). » The microcuves have a geometry more favorable for immunoenzymatic reactions (surface/volume 3.8x superior), compared with wells of standard microplates : hence kinetic is speeded at each step (ex incubations 2 fold shorter in ELISA).
(a) (b) (c) Accessory tools
IMAplate technology combines advantageously notably in ELISA, and for multiplexed analysis.
IMAplate offer a solution at the same time more flexible, quicker and cost-effective, when : » Samples are in limited quantity or precious, » Reagents are costly (case of commercial kits), » Several analysis are performed on each sample (multiplex), » To speed steps and handling with reliability.
Examples of applications particularly appropriate : » Serological analysis of many analytes in small animals serums » Multiplex screening (pharma, cosmeto, vaccines)
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 73 74 Accessory tools on auniquesupport,flexible&efficient,withoutinstrumentinvestment! ■ Integrateyourdifferentmethods/analysis Microplates compound InfraRedanalysis Proteins dosage(UV280nm) DNA/RNA ex : SPECTROMETRY Commercial Kits Reduce by3-20thecostofreagents "home made" Reduce cost/testby2-5fold Homogenous-Phase assays–CELL Cell Biology:MTT, SOD, Biochemistry (clinical,IAA): Proteins Dosages:BC ex : Colorimetric assays- dosage(260nm) Assays ABS Assay, Coo Thromboxanes... Tests Glucose...
Assay Assays Toxicology :LDH,Grantoxilux... Cell viability:UptiBlue... Calcium Flux:Fluo-3,Fluo-8... ofDNA/RNA Assay ofProteins:LavaPep ex : Fluorimetric Cell Reduce timesofincubation,transfer, samples &costlyreagentsby3-20X Increase thesensitivityofdetection Reduce thevolumes&costofrare .com.com votre source d’innovations IMAplate technology and washes,... Assay -FL :Evagreen Other systemsofmuliplexedanalysis Reduce by3-20thecostofbuffers, saturants,antibodies,enz.substrates "home made"ELISA Reduce cost/testby2-5fold ELISA Solid-Phase assays-ELISA CommercialKits : TMB, UptiLightECL... Cell viability: Calcium Flux:Coelenterazine/aequaporin... ELISA ex : (Chemi)Luminescent Reporter assay:Luciferase,Coelenterazine... AND MicroCuvette :5µlvolume (automatable) centrifugation) Easy µcuveun-loading(10sec/absorbant, (10 sec/capillaryforce,automatable) Easy µcuveloading ex : LIQUID HANDLING TIME BY avecHRP 20-50% ATP/lum... (Luminex, SpotELISA,Micro-Arrays) /UptiLightECL Assays -ECL ■ Antifade Kit for Microplate
When exposed to excitation light, fluorescence intensity of dyes decreases due to their photooxidation or other photoreactions. There are very few fluorescent dyes that completely resist photobleaching. Frequently, when a section has been scanned repeatedly under strong excitation light, dyes could lose significant fluorescence signal before visual evaluation or photography can be accomplished. For examples, the photobleaching of fluoresceins (such as FITC-labeled antibodies) has become a major problem in fluorescence microscopy. In severe cases (such as phycoprotein-labeled bioconjugates), a fluorescence image of high resolution can not even be taken due to the extremely high photobleaching rate. The Antifade Kit is to reduce the dye photobleaching rate, giving researchers longer observation time. The kit contains all the essential components that can be readily applied to imaging experiments. They are all premixed and ready- to-use solutions. This kit is designed for microplate format.
Description P/N : Qty Antifade Kit for Microplate FP-CL0530 1 plate
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Acella from CellTechnologies Cytonin, PeroxyGlow, and TACS from Trevigen CyToxiLux and GranToxiLux from OncoImmun DeepBlue C from BioSignal Packard FluoProbes, PrimAb and UptiBlue from Interchim MUP plus, Phospholite, and Rhod-4 from ABD PMA from Biotium Twister from Caliper
votre source d’innovations Tél 33 (0)4 70 03 88 55 - Fax 33 (0)4 70 03 82 60 - www.interchim.com 75