| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 646/659 |
| MW | N/A |
| CAS# | N/A |
| Solvent | N/A |
| Storage | F/D/L |
| Category | CellAnalysis CellCytotoxicity |
| Related | ApoptosisandCytotoxicity CellApoptosis |
| Spectrum | AdvancedSpectrumViewer |
1.Foreachsample,preparecellsin0.5mLwarmmediumorbufferofyourchoiceatadensityof5×105to1×106cells/mL.
Note:Eachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensityforapoptosisinduction.
2.Treatcellswithtestcompoundsforadesiredperiodoftimetoinduceapoptosis,andcreatepositiveandnegativecontrols.
3.Add1μLof500XTF5-VAD-FMK(ComponentA)intothetreatedcells(fromStep2),andincubatethecellsina37 °C,5%CO2incubatorfor1-4hours.
Note1:Foradherentcells,gentlyliftthecellswith0.5mMEDTAtokeepthecellsintact,andwashthecellsoncewithserum-containingmediapriortoincubationwithTF5-VAD-FMK.
Note2:Theappropriateincubationtimedependsontheindividualcelltypeandcellconcentrationused.Optimizetheincubationtimeforeachexperiment.
4.Washandspinthecellstwice.ResUSPendthecellsin0.5mLofAssayBuffer(ComponentB)orgrowthmedium.
Note:TF5-VAD-FMKisfluorescent;thereforeitisimportanttowashoutanyunboundreagenttoremovethebackground.
5.Ifdesired,labelthecellswithaDNAstain[suchasGreen-DCSfordeadcells,whichcanbedetectedintheFL1channel(Ex/Em=490/525nm).
6.Ifdesired,fixcells.
7.MonitorthefluorescenceintensitywithaflowcytometerusingtheFL4channel(Ex/Em=650/670nm).Gateonthecellsofinterest,excludingdebris.
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