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AAT Bioquest/Cell Meter™ Fluorimetric Cell Cytotoxicity Assay Kit/22781/1000 Tests
Overview | PrinterFriendlyVersion |
Ex/Em(nm) | 571/585 |
MW | N/A |
CAS# | N/A |
Solvent | N/A |
Storage | F/D/L |
Category | CellAnalysis CellCytotoxicity |
Related | ApoptosisandCytotoxicity CellApoptosis BiochemicalAssays |
Spectrum | AdvancedSpectrumViewer |
1.Preparecellsandtestcompounds:
1.1 Plate100to10,000cellsperwellinatissueculturemicroplatewithblackwallandclearbottom.Addtestcompoundsintothecells,andincubateforadesiredperiodoftime(suchas24,48or96hours)ina37°C,5%CO2incubator.Forblankwells(mediumwithoutthecells),addthesameamountofcompoundbuffer.Thesuggestedtotalvolumeis100µLfora96-wellplate,and50µLfora384-wellplate.
1.2 Setupthefollowingcontrolsatthesametime.
•Positivecontrolcontainscellsandknownproliferationorcytotoxicityinducer.
•Negativecontrolcontainscellsbutnotestcompounds.
•Vehiclecontrolcontainscellsandthevehicleusedtodelivertestcompounds.
•Non-cellcontrolcontainsgrowthmediumwithoutcells.
Note:LDHinserumwillcontributetobackgroundfluorescence.
•Testcompoundcontrolcontainsthevehicleusedtodelivertestcompounds[Hank’sbalancesaltsolution(HBSS)orphosphate-bufferedsaline(PBS)]andtestcompound.Sometestcompoundshavestrongautofluorescenceandmaygivefalsepositiveresults.
Note:Matchthetotalvolumeofallthecontrolsto100µLfora96-wellplateor50µLfora384-wellplatewithgrowthmedium.
2.Assayprocedures:
2.1 ThawandwarmuptheAssaySolution(ComponentA)to37°C,andmixitthoroughlybeforestartingtheexperiments.
2.2 Add20µL/well(96-wellplate)or10µL/well(384-wellplate)ofAssaySolution(ComponentA).Mixthereagentsbyshakingtheplategentlyfor30seconds.
2.3 Incubatethecellsina37oC,5%CO2incubatorfor1-24hours,protectedfromlight.
Note1:Theappropriateincubationtimedependsonthemetabolismrateoftheindividualcelltypeandcellconcentrationused.Optimizetheincubationtimeforeachexperiment.
Note2:Extremelyprolongedincubationtimeisnotrecommendedsinceresazurincouldbeconvertedtocolorlessdihydroresorufin.
2.4 Monitorthefluorescenceintensity(bottomread)atEx/Em=540/590nm.Alternatively,readtheO.D.at570nm(thereferencewavelengthshouldbe600nm)todeterminethecellviabilityineachwell.
3.Performdataanalysis:
3.1 ThebackgroundfluorescencereADIngfromthenon-cellcontrolwellissubtractedfromthevaluesforthosewellscontainingthecells.
Note:Thebackgroundfluorescenceoftheblankwellsmayvarydependingonthesourcesofthegrowthmediaorthemicrotiterplates.
3.2 Thefluorescencereadingineachwellindicatesthecellnumberinthatwell.
3.3 Calculatethepercentageofcellviabilityforsamplesandcontrolsbasedonthefollowingformula:
%Cellviability=100×(Fsample-Fo)/(Fctrl-Fo)
Fsampleisthefluorescencereadinginthepresenceofthetestcompound.
Fctrlisthefluorescencereadingintheabsenceofthetestcompound(vehiclecontrol).
Foistheaveragedbackground(non-cellcontrol)fluorescenceintensity.
References&Citations | PrinterFriendlyVersion |
1. McNichollBP,McGrathJW,QuinnJP.(2006)Developmentandapplicationofaresazurin-basedbiomassactivitytestforactivatedsludgeplantmanagement.WaterRes.
2. OriowoMO.(2006)Afluorometricstudyofrelativeocularlenscytosensitivitytomultipurposecontactlenssolutionsusingtheresazurinassaymethod.ToxicolInVitro,20,1548.
3. UmubyeyiAN,MartinA,ZissisG,StruelensM,KaritaE,PortaelsF.(2006)EvaluationoftheresazurinmicrotiterassayforrapiddetectionofofloxacinresistanceinM.tuberculosis.IntJTubercLungDis,10,808.
4. NatecheF,MartinA,BarakaS,PalominoJC,KhaledS,PortaelsF.(2006)ApplicationoftheresazurinmicrotitreassayfordetectionofmultidrugresistanceinMycobacteriumtuberculosisinAlgiers.JMedMicrobiol,55,857.
5. CobanAY,CekicCihanC,BilginK,UzunM,AkgunesA,CetinkayaE,DurupinarB.(2006)RapidsusceptibilitytestforMycobacteriumtuberculosistoisoniazidandrifampinwithresazurinmethodinscrew-captubes.JChemother,18,140.
6. ZrimsekP,KosecM,KuncJ,MrkunJ.(2006)Determinationofthediagnosticvalueoftheresazurinreductionassayforevaluatingboarsemenbyreceiveroperatingcharacteristicanalysis.AsianJAndrol,8,343.
7. RolonM,VegaC,EscarioJA,Gomez-BarrioA.(2006)DevelopmentofresazurinmicrotiterassayfordrugsensibilitytestingofTrypanosomacruziepimastigotes.ParasitolRes,99,103.
8. TizzardAC,BergsmaJH,Lloyd-JonesG.(2006)Aresazurin-basedbiosensorfororganicpollutants.BiosensBioelectron,22,759.
9. Abu-AmeroKK,BosleyTM.(2005)Detectionofmitochondrialrespiratorydysfunctionincirculatinglymphocytesusingresazurin.ArchPatholLabMed,129,1295.
10. Anoopkumar-DukieS,CareyJB,ConereT,O"SullivanE,vanPeltFN,AllshireA.(2005)Resazurinassayofradiationresponseinculturedcells.BrJRadiol,78,945.