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    當前位置 ： Millipore >>> Millipore/APT409 | CaspaTag Caspase 9 In Situ Assay Kit, Fluorescein/APT409/100 assays
    Millipore/APT409 | CaspaTag Caspase 9 In Situ Assay Kit, Fluorescein/APT409/100 assays
    • Millipore/APT409 | CaspaTag Caspase 9 In Situ Assay Kit, Fluorescein/APT409/100 assays

    Millipore/APT409 | CaspaTag Caspase 9 In Situ Assay Kit, Fluorescein/APT409/100 assays

    價格: ￥6876.00 市場價: 11460.00
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    貨號: APT409
    品牌: Millipore
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      • Description
        CatalogueNumberAPT409
        BrandFamilyChemicon®
        TradeName
        • CaspaTag
        • Chemicon
        DescriptionCaspaTagCaspase9InSituAssayKit,Fluorescein
        BackgroundInformationApoptosisisanevolutionarilyconservedformofcellsuicide,whichfollowsaspecializedcellularprocess.Thecentralcomponentofthisprocessisacascadeofproteolyticenzymescalledcaspases.Theseenzymesparticipateinaseriesofreactionsthataretriggeredinresponsetopro-apoptoticsignalsandresultinthecleavageofproteinsubstrates,causingthedisassemblyofthecell1.

        CaspaseshavebeenidentifiedinorganismsrangingfromC.eleganstohumans.Themammaliancaspasesplaydistinctrolesinapoptosisandinflammation.Inapoptosis,caspasesareresponsIBLeforproteolyticcleavagesthatleadtocelldisassembly(effectorcaspases),andareinvolvedinupstreamregulatoryevents(initiatorcaspases).Anactivecaspaseconsistsoftwolargeandtwosmallsubunitsthatformtwoheterodimers,whichassociateinatetramer2-4.Incommonwithotherproteases,caspasesaresynthesizedasprecursorsthatundergoproteolyticmaturation,eitherautocatalyticallyorinacascadebyenzymeswithsimilarspecificity5.

        Caspaseenzymesspecificallyrecognizea4or5aminoacidsequenceonthetargetsubstrate,whichnecessarilyincludesanasparticacidresidue.Thisresidueisthetargetforcleavage,whichoccursatthecarbonylendoftheasparticacidresidue6.Caspasescanbedetectedviaimmunoprecipitation,immunoblottingtechniquesusingcaspasespecificantibodies,orbyemployingFluorochromesubstrates,whichbecomefluorescentuponcleavagebythecaspase.
        MaterialsRequiredbutNotDelivered·Culturedcellswithmedia

        ·Reagentstoinduceapoptosis

        ·15mLpolystyrenecentrifugetubes

        ·Ambervialsortubesforstorageof150Xconcentrateat-20°C

        ·600mLgraduatedcylinder

        ·Microscopeslides

        ·Hemocytometer

        ·Centrifuge(400xg)

        ·37°Cincubator

        ·Vortexer

        ·Adjustablevolumepipettorwithdisposabletips

        ·Deionizedwater

        ·PBS,pH7.4

        ·DMSO
        ProductInformation
        Components
        • FLICAReagent(FAM-LEHD-FMK):Fourlyophilizedvials
        • 10XWashBuffer:60mL
        • Fixative:6mL
        • PropidiumIodide:1mLat250μg/mL,ready-to-use
        • HoechstStain:1mLat200μg/mL,ready-to-use
        DetectionmethodFluorescent
        StorageandShippingInformation
        StorageConditions·Storeunopenedkitmaterialsat2-8°Cuptotheirexpirationdate.

        ·ReconstitutedFLICAReagent(150X)shouldbefrozenat-20°Cforupto6months,andmaybethawedtwiceduringthistime.Aliquotintoseparateambertubesifdesired.Protectfromlightatalltimes.

        ·Storediluted(1X)washbufferupto-20°Cfor2weeks.
        Applications
        ApplicationTheCaspaTagCaspase-9InSituAssayKit,Fluoresceinforflowcytometryisafluorescent-basedassayfordetectionofactivecaspase-9incellsundergoingapoptosis.
        KeyApplications
        • FlowCytometry
        • ActivityAssay
        ApplicationNotesTheCHEMICON?CaspaTag?Caspase-9InSituAssayKit,Fluoresceinisafluorescent-basedassayfordetectionofactivecaspase-9incellsundergoingapoptosis.Thekitisforresearchuseonly.Notforuseindiagnosticortherapeuticprocedures.

        TestPrinciple

        CHEMICON?"sCaspaTag?Caspase-9InSituAssayKitsuseanovelapproachtodetectactivecaspases.ThemethodologyisbasedonFluorochromeInhibitorsofCaspases(FLICA).Theinhibitorsarecellpermeableandnon-cytotoxic.Onceinsidethecell,theinhibitorbindscovalentlytotheactivecaspase7.Thiskitusesacarboxyfluorescein-labeledfluoromethylketonepeptideinhibitorofcaspase-9(FAM-LEHD-FMK),whichproducesagreenfluorescence.Whenaddedtoapopulationofcells,theFAM-LEHD-FMKprobeenterseachcellandcovalentlybindstoareactivecysteineresiduethatresidesonthelargesubunitoftheactivecaspaseheterodimer,therebyinhibitingfurtherenzymaticactivity.Theboundlabeledreagentisretainedwithinthecell,whileanyunboundreagentwilldiffuseoutofthecellandiswashedaway.Thegreenfluorescentsignalisadirectmeasureoftheamountofactivecaspase-9presentinthecellatthetimethereagentwasadded.Cellsthatcontaintheboundlabeledreagentcanbeanalyzedby96-wellplate-basedfluorometry,fluorescencemicroscopy,orflowcytometry.
        BIOLOGicalInformation
        SpeciesReactivity
        • Mammals
        EntrezGeneNumber
        EntrezGeneSummaryThisgeneencodesamemberofthecysteine-asparticacidprotease(caspase)family.Sequentialactivationofcaspasesplaysacentralroleintheexecution-phaseofcellapoptosis.Caspasesexistasinactiveproenzymescomposedofaprodomain,alargeproteasesubunit,andasmallproteasesubunit.Activationofcaspasesrequiresproteolyticprocessingatconservedinternalasparticresiduestogenerateaheterodimericenzymeconsistingofthelargeandsmallsubunits.ThisproteinisinvolvedintheprogrammedcelldeathinducedbyFasandvariousapoptoticstimuli.TheN-terminalFADD-likedeatheffectordomainofthisproteinsuggeststhatitmayinteractwithFas-interactingproteinFADD.ThisproteinwasdetectedintheinsolublefractionoftheaffectedbrainregionfromHuntingtondiseasepatientsbutnotinthosefromnormalcontrols,whichimplicatedtheroleinneurodegenerativediseases.Manyalternativelysplicedtranscriptvariantsencodingdifferentisoformshavebeendescribed,althoughnotallvariantshavehadtheirfull-lengthsequencesdetermined.
        GeneSymbol
        • CASP8
        • MCH5
        • MGC78473
        • CASP-8
        • MACH
        • ALPS2B
        • procaspase-8
        • FLICE
        • CAP4
        • EC3.4.22.61[Contains:Caspase-8subunitp18
        • Caspase-8subunitp10].
        UniProtNumber
        UniProtSummaryFUNCTION:SwissProt:Q14790#MostupstreamproteaseoftheactivationcascadeofcaspasesresponsiblefortheTNFRSF6/FASmediatedandTNFRSF1Ainducedcelldeath.BindingtotheadaptermoleculeFADDrecruitsittoeitherreceptor.Theresultingaggregatecalleddeath-inducingsignalingcomplex(DISC)performsCASP8proteolyticactivation.TheactivedimericenzymeisthenliberatedfromtheDISCandfreetoactivatedownstreamapoptoticproteases.ProteolyticfragmentsoftheN-terminalpropeptide(termedCAP3,CAP5andCAP6)arelikelyretainedintheDISC.CleavesandactivatesCASP3,CASP4,CASP6,CASP7,CASP9andCASP10.MayparticipateintheGZMBapoptoticpathways.CleavesADPRT.Hydrolyzesthesmall-moleculesubstrate,Ac-Asp-Glu-Val-Asp--AMC.LikelytargetforthecowpoxvirusCRMAdeathinhibitoryprotein.Isoforms5,6,7and8lackthecatalyticsiteandmayinterferewiththepro-apoptoticactivityofthecomplex.
        SIZE:479aminoacids;55391Da
        SUBUNIT:Heterotetramerthatconsistsoftwoanti-parallelarrangedheterodimers,eachoneformedbya18kDa(p18)anda10kDa(p10)subunit.InteractswithFADD,CFLARandPEA15.Isoform9interactsattheendoplasmicreticulumwithacomplexcontainingBCAP31,BAP29,BCL2and/orBCL2L1.
        SUBCELLULARLOCATION:Cytoplasm.
        TISSUESPECIFICITY:Isoforms1,5and7areexpressedinawidevarietyoftissues.Highestexpressioninperipheralbloodleukocytes,spleen,thymus,andliver.Barelydetectableinbrain,testis,andskeletalmuscle.DOMAIN:SwissProt:Q14790Isoform9containsaN-terminalextensionthatisrequiredforinteractionwiththeBCAP31complex.
        PTM:Generationofthesubunitsrequiresassociationwiththedeath-inducingsignalingcomplex(DISC),whereasadditionalprocessingislikelyduetotheautocatalyticactivityoftheactivatedprotease.GZMBandCASP10canbeinvolvedintheseprocessingevents.&PhosphorylateduponDNAdamage,probablybyATMorATR.
        DISEASE:SwissProt:Q14790#DefectsinCASP8arethecauseofcaspase-8deficiency(CASP8D)[MIM:607271].CASP8Disadisorderresemblingautoimmunelymphoproliferativesyndrome(ALPS).Itischaracterizedbylymphadenopathy,splenomegaly,anddefectiveCD95-inducedapoptosisofperipheralbloodlymphocytes(PBLs).ItleadstodefectsinactivationofT-lymphocytes,B-lymphocytes,andnaturalkillercellsleADIngtoimmunodeficiencycharacterizedbyrecurrentsinopulmonaryandherpessimplexvirusinfectionsandpoorresponsestoimmunization.
        SIMILARITY:BelongstothepeptidaseC14family.&Contains2DED(deatheffector)domains.
        PhysicochemicalInformation
        Dimensions
        MaterialsInformation
        MaterialsInformation
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