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Millipore/17-472 | Phospho-GSK-3β (Ser9) STAR ELISA Kit/17-472/96 assays
  • Millipore/17-472 | Phospho-GSK-3β (Ser9) STAR ELISA Kit/17-472/96 assays

Millipore/17-472 | Phospho-GSK-3β (Ser9) STAR ELISA Kit/17-472/96 assays

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貨號: 17-472
品牌: Millipore
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    • Description
      CatalogueNumber17-472
      BrandFamilyUpstate
      TradeName
      • STAR
      • Upstate
      DescriptionPhospho-GSK-3β(Ser9)STARELISAKit
      AlternateNames
      • glycogensynthasekinase3beta
      BackgroundInformationI.TESTPRINCIPLETheUPSTATE?colorimetricSTAR(SignalTransductionAssayReaction)ELISAkitisasolidphasesandwichenzymelinkedimmunosorbentassaythatprovidesafast,sensitivemethodtodetectspecificlevelsofsignalingtargetsinwholecellextracts.TheGSK-3βplateiscoatedwithaspecificmousemonoclonalGSK-3&betacaptureantibodyonthemicrowellsofthe96-wellclearplate.SamplelysateorthestandardincludedinthekitareincubatedinthemicrowellsallowingGSK-3βantigentobecapturedintheplatewells.Theplateisthenwashedtoremoveanynon-boundunspecificmaterial.Thewellsarethenincubatedwithaspecificrabbitanti-GSK-3βantibodytodetectthecapturedphosphor-GSK-3β(Ser9)ontheplatewell.TheunbounddetectionantibodyiswashedawayfollowedbyincubationwithanHRP-conjugatedanti-rabbitantibody.Thisallowsforasensitiveenzymaticdetectionofthesample.AftertheadditionofTMBsubstrateandstopsolutiontheabsorbanceismeasuredat450nmusingaplatereader.Theentireassaytakeslessthan5hourstocompletewithminimalhands-ontime.Manyofthereagentsaresuppliedinready-touseformulationsforeaseofuse.Thekitalsoincludesastandardthatisrunasbothapositivecontrolandtodevelopastandardlineofdetection.

      II.GSK-3βBACKGROUNDGlycogenSynthaseKinase-3differsfrommostserine/threoninekinasesinthatitisactiveintheabsenceoftheactionofsignalingpathways.Whenitbecomesphosphorylated,theproteinbecomesinactive.Therearetwoisoformsthatexist,GSK-3αandGSK-3β.ThefunctionofGSK-3istophosphorylateGlycogenSynthaseandtherebyinactivateit.InsulinactionstimulatesthePI3KinasepathwaythatresultsintheactivationofAkt.AktphosphorylatesGSK-3,therebyinactivatingit.GlycogenSynthase,GSK-3"sdownstreamsubstrate,isthenrapidlydephosphorylatedandactivated.OtherGSK-3substratesincludeJun(oninhibitorysites),andeIF2B.DetectionofGSK-3"sphosphorylationstatusoftheAktsite(Ser21)onGSK-3issuitableforsurrogateassaysoftheactivationstateofthepathway.
      MaterialsRequiredbutNotDelivered1.Multi-channelorrepeatingPipettes
      2.Plateshaker(optional)
      3.Pipettorsandtipscapableofaccuratelymeasuring1-1000μL
      4.GraduatedSEROlogicalpipettes
      5.96-wellmicrotiterPlateReaderwith450nmfilter
      6.Graphingsoftwareforplottingdataorgraphpaperformanualplottingofdata
      7.Microfugetubesforstandardandsampledilutions
      8.Mechanicalvortex
      9.1litercontainer
      10.Distilledordeionizedwater
      ProductInformation
      Components
      • GSK-3βCapturePlate:(PartNo.17-472A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch.
      • Anti-phospho-GSK-3β(Ser9)detectionantibody:(PartNo.17-472B)Onebottle(6mL)ofanti-phospho-GSK-3β(Ser9)detectionantibodycontainingsodiumazide,readytouse.
      • ELISADiluent:(PartNo.17-472C)Onebottle(25mL)ofELISADiluentsodiumazide,readytouse
      • 25XELISAWashBuffer:(PartNo.17-472D)Onebottle(50mL)of25XELISAWashBuffer.
      • Anti-RabbitIgGHRPconjugate:(PartNo.17-472E)Onevial(125μL)of100Xanti-rabbitHRPconjugate.
      • HRPDiluent:(PartNo.17-472F)Onebottle(25mL)ofHRPdiluent.
      • TMBSolution:(PartNo.17-472G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse.
      • StopSolution:(PartNo.17-472H)Onebottle(25mL)ofstopsolution,readytouse.
      • PhosphorylatedGSK-3β(Ser9)Standard:(PartNo.17-472I)TwovialsofphosphorylatedGSK-3β(Ser9)standard,lyophilized.
      • PlateCovers:Twoplatecovers.
      DetectionmethodColorimetric
      StorageandShippingInformation
      StorageConditionsMaintaintheunopenedkitat2-8°Cuntilexpirationdate.
      Precautions
      ?Theinstructionsprovidedhavebeendesignedtooptimizethekit"sperformance.Deviationfromtheinstructionsmayresultinsuboptimalperformanceofthekitandthefailuretoproduceaccuratedata.
      ?CausticMaterial:StopSolution.Caution:Eye,hand,face,andclothingprotectionshouldbewornwhenhandlingthismaterial.
      ?SafetyWarningsandPrecautions:Thiskitisdesignedforresearchuseonlyandnotrecommendedforinternaluseinhumansoranimals.Allchemicalsshouldbeconsideredpotentiallyhazardousandprinciplesofgoodlaboratorypracticeshouldbefollowed.
      ?TheDetectionAntibodyandELISADiluentcontainsodiumazide.Sodiumazidemayreactwithcopperandleadplumbingtoformhighlyexplosivemetalazides.Upondisposal,flushwithlargeamountsofwatertopreventazidebuild-up.Avoidcontactwithskin.
      ?TheAnti-RabbitIgGHRPConjugateandHRPDiluentcontainthimerosal.Thimerosalishighlytoxicbyinhalation,contactwithskinorifswallowed.ThimerosalisapossIBLemutagenandshouldbehandledaccordingly.
      Applications
      ApplicationThisPhospho-GSK-3β(Ser9)STARELISAKitisusedtomeasure&quantifyGSK-3βlevels.
      KeyApplications
      • ELISA
      BIOLOGicalInformation
      SpeciesReactivity
      • Human
      • Mouse
      • Rat
      AnalytesAvailable
      • GSK-3β
      EntrezGeneNumber
      EntrezGeneSummaryGlycogensynthasekinase-3(GSK3)isaproline-directedserine-threoninekinasethatwasinitiallyidentifiedasaphosphorylatingandinactivatingglycogensynthase.Twoisoforms,alpha(GSK3A;MIM606784)andbeta,showahighdegreeofaminoacidhomology(StambolicandWoodgett,1994[PubMed7980435]).GSK3Bisinvolvedinenergymetabolism,neuronalcelldevelopment,andbodypatternformation(Plyteetal.,1992[PubMed1333807]).[suppliedbyOMIM]
      GeneSymbol
      • GSK3B
      Modifications
      • Phosphorylation
      UniProtNumber
      UniProtSummaryFUNCTION:SwissProt:P49841#ParticipatesintheWntsignalingpathway.Implicatedinthehormonalcontrolofseveralregulatoryproteinsincludingglycogensynthase,MYBandthetranscriptionfactorJUN.PhosphorylatesJUNatsitesproximaltoitsDNA-bindingdomain,therebyreducingitsaffinityforDNA.
      SIZE:420aminoacids;46744Da
      SUBUNIT:Monomer(Bysimilarity).InteractswithNINandCABYR.
      TISSUESPECIFICITY:Expressedintestis,thymus,prostateandovaryandweaklyexpressedinlung,brainandkidney.
      PTM:PhosphorylatedbyAKT1andILK1.
      SIMILARITY:SwissProt:P49841##Belongstotheproteinkinasesuperfamily.CMGCSer/Thrproteinkinasefamily.GSK-3subfamily.&Contains1proteinkinasedomain.
      PhysicochemicalInformation
      Sensitivity
      • Sensitivity:1.6Units/mL
        RangeofDetection:1.6to100Units/mL
      Dimensions
      MaterialsInformation
      MaterialsInformation
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