| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 492/516 |
| MW | 1031.07 |
| CAS# | N/A |
| Solvent | DMSO |
| Storage | F/D/L |
| Category | EnzymeDetection PeptidasesandProteases |
| Related | CellApoptosis ApoptosisandCytotoxicity BiochemicalAssays |
| Spectrum | AdvancedSpectrumViewer |
1. GeneralSolutionCaspaseAssaysUsingAMC,AFC,pNA,R110andProRedSubstrates
1.1. Preparea10mMstocksolutioninDMSO.
1.2. Preparea2Xcaspase substrate(50µM)assaysolutionasthefollowing:
50µLsubstratestocksolution
100 µLDTT(1M)
400 µLEDTA(100mM)
10mLTrisBuffer(20mM),pH=7.4
1.3. Mixequalvolumeofthecaspasestandardsorsampleswith2Xcaspasesubstrateassaysolution(fromStep1.2),andincubatethesolutionsatroomtemperatureforatleast1hour.
1.4. Monitorthefluorescenceusingafluorescencemicroplatereader,orabsorbanceusinganabsorbancemicroplatereader.
2. CellCaspaseAssaysUsingCell-PermeableFMKCaspaseProbes
2.1. Preparea2-5mMstocksolutioninDMSO.
2.2. Treatcellsasdesired.
2.3. Preparea2Xpermeablecaspasesubstrate(20µM)assaysolutionbydilutingtheDMSOstocksolution(fromStep2.1)inHankswith20mMHepesbuffer(HHBS).
2.4. Mixequalvolumeofthetreatedcellswith2Xcaspasesubstrateassaysolution(fromStep2.3),andincubatethecellsina37°C,5%CO2incubatorforatleast1hour.
2.5. WashthecellswithHHBSforatleastonce.
2.6. Monitorthefluorescenceintensitybyaflowcytometer,afluorescencemicroscopeorafluorescencemicroplatereader.
3. CellCaspaseAssaysUsingCell-PermeableFMKCaspaseProbes(For#13470-13476only)
3.1. Preparea250Xstocksolutionbyadding50µLDMSOintothevial.
3.2. Treatcellsasdesired.
3.3. Add250XDMSOstocksolution(fromStep3.1)intothecellsolutionata1:250ratio(suchas2uLto500uLcells),andincubatethecellsina37°C,5%CO2incubatorfor1hour.
3.4. WashthecellswithHHBSforatleastonce.
3.5. Monitorthefluorescenceintensitybyflowcytometer,fluorescencemicroscopyorfluorescentmicroplatereader.
| References&Citations |  PrinterFriendlyVersion |
1. BrewsterJL,LinsemanDA,BouchardRJ,LoucksFA,PrechtTA,EschEA,HeidenreichKA.(2006)Endoplasmicreticulumstressandtrophicfactorwithdrawalactivatedistinctsignalingcascadesthatinduceglycogensynthasekinase-3betaandacaspase-9-dependentapoptosisincerebellargranuleneurons.MolCellNeurosci,32,242.
2. SordilloLM,WeaverJA,CaoYZ,CorlC,SylteMJ,MullarkyIK.(2005)Enhanced15-HPETEproductionduringoxidantstressinducesapoptosisofendothelialcells.ProstaglandinsOtherLipidMediat,76,19.
3. WangQ,WangL,FengYH,LiX,ZengR,GorodeskiGI.(2004)P2X7receptor-mediatedapoptosisofhumancervicalepithelialcells.AmJPhysiolCellPhysiol,287,C1349.
4. CheongJW,ChongSY,KimJY,EomJI,JeungHK,MaengHY,LeeST,MinYH.(2003)Inductionofapoptosisbyapicidin,ahistonedeacetylaseinhibitor,viatheactivationofmitochondria-dependentcaspasecascadesinhumanBcr-Abl-positiveleukemiacells.ClinCancerRes,9,5018.
5. SylteMJ,LeiteFP,KuckleburgCJ,InzanaTJ,CzuprynskiCJ.(2003)CaspaseactivationduringHaemophilussomnuslipooligosaccharide-mediatedapoptosisofbovineendothelialcells.MicrobPathog,35,285.
6. SumitomoM,OhbaM,AsakumaJ,AsanoT,KurokiT,HayakawaM.(2002)ProteinkinaseCdeltaamplifiesceramideformationviamitochondrialsignalinginprostatecancercells.JClinInvest,109,827.
7. SchlesingerM,JiangJD,RobozJP,DennerL,LingYH,HollandJF,BekesiJG.(2000)3-m-bromoacetylaminobenzoicacidethylester:anewcancericidalagentthatactivatestheapoptoticpathwaythroughcaspase-9.BiochemPharmacol,60,1693.
8. ZhuH,McElwee-WitmerS,PerroneM,ClarkKL,ZilbersteinA.(2000)Phenylephrineprotectsneonatalratcardiomyocytesfromhypoxiaandserumdeprivation-inducedapoptosis.CellDeathDiffer,7,773.
9. ChatterjeeS,MatsumuraA,SchradermeierJ,GillespieGY.(2000)Humanmalignantgliomatherapyusinganti-alpha(v)beta3integrinagents.JNeurooncol,46,135.
10. FadeelB,HassanZ,Hellstrom-LindbergE,HenterJI,OrreniusS,ZhivotovskyB.(1999)CleavageofBcl-2isanearlyeventinchemotherapy-inducedapoptosisofhumanmyeloidleukemiacells.Leukemia,13,719.
