NucView™488Caspase-3substrateisanovelcellmembrane-permeablefluorogeniccaspasesubstratedesignedfordetectingcaspase-3activitywithinlivecellsinrealtime.VisitourProductVideopage toseeavideoofNucView™488inaction.
Therateofapoptosistypicallyvariesfromcelltocellevenwithinthesamepopulation.Asaresult,variousapoptoticeventsorMarkersaccompanyingtheapoptoticprocessalsooccurdifferentlyamongcells.Thus,itisimportanttobeabletodetecttheseapoptoticeventsonanindividualcellbasis.TrADItionally,caspaseactivityhasbeendetectedeitherusingamembrane-impermeablefluorogenicenzymesubstratesuchasDEVD-R110,orafluorescently-labeledinhibitorsuchasaFLICAreagent.Intheformercase,celllysisisrequired,thusprecludingthedetectionofcaspaseactivityinlivecells.Inaddition,suchcaspaseassaysmeasureonlytheaveragecaspaseactivityofahighlyheterogeneouscellpopulationatagiventime.Inthelattercase,althoughaFLICAreagentcanenterlivecellstodetectcaspaseactivity,onlytheinitialfluorescentsignalfollowingtheapplicationofthereagentcantrulyreflecttheenzymeactivityorthestateoftheapoptoticcellsbecauseanydetectedsignalaftertheinitial“snapshot”willneedtoconsiderthepotentialinterferenceoftheinhibitortotheenzymeandtheapoptoticcellitself.
Unlikeconventionalcaspaseassays,NucView™488Caspase-3substratedetectscaspase-3activitywithinindividualintactcellswithoutinhibitingcaspase-3activity.ThesubstrateconsistsofafluorogenicDNAdyeandaDEVDsubstratemoietyspecificforcaspase-3.Thesubstrate,whichisbothnon-fluorescentandnonfunctionalasaDNAdye,rapidlycrossescellmembranestoenterthecytoplasm,whereitiscleavedbycaspase-3toformahigh-affinityDNAdyethatstainsthenucleusbrightgreen.Thus,theNucView™488caspase-3substrateisbi-functional,allowingdetectionofintracellularcaspase-3activityandvisualizationofchangesinnuclearmorphologyduringapoptosis.Thefluorescentstainingproducedinresponsetocaspase-3activityiscompatIBLewithsubsequentfixationandpermeABIlizationforimmunostaining.
ThekitcontainsNucView™488Caspase-3substrateandacaspase-3inhibitorAc-DEVD-CHOsufficientfor100flowcytometryormicroplateassays(200uLassayvolume).Thetrialsizekitcontainssufficientreagentsfor25assays(200uLassayvolume).Thenumberoffluorescencemicroscopyassaysthatcanbeperformedwiththekitmayvarybasedonthesizeofculturevesselandstainingvolumeused.Thekitprovidesaconvenienttoolforprofilingapoptoticcellpopulationbasedoncaspase-3activityusingeitherfluorescencemicroscopyorflowcytometry.
KitComponents:
- NucView™488Caspase-3substrate,0.2mMinDMSO
- Caspase-3inhibitorAc-DEVD-CHO,2mMinDMSO
Features:
- Realtimecaspase-3detectioninintactcells:Detectcaspase-3activitywithinindividualcellsinapopulation.
- Bi-functional:Allowsvisualizationofcaspase-3activityandapoptoticnuclearmorphologyatthesametime.
- Simple&Fast:15-minuteassaywithnowashrequired.
- Versatile:Fordetectionbyflowcytometry,fluorescencemicroscopy,orfluorescencemicroplatereaderusingfluoresceindetectionsettings.
- Fixable:Compatiblewithstandardfixationandpermeabilizationmethodsforsubsequentimmunofluorescencestaining.
SeeourfullselectionofNucView™substratesandkits.
NucView™enzymesubstratetechnologyiscoveredbyU.S.PatentNos.8,092,784and8,586,325.
References
Cen,etal.DEVD-NucView488:anovelclassofenzymesubstratesforreal-timedetectionofcaspase-3activityinlivecells.TheFASEBJournalpublishedonlineFebruary8,2008.
Downloadafulllistofreferences:ValidatedcelllinesforNucView488withreferences
