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Millipore/17-488 | phospho-p38α (Thr180/Tyr182) STAR ELISA Kit/17-488/96 assays
  • Millipore/17-488 | phospho-p38α (Thr180/Tyr182) STAR ELISA Kit/17-488/96 assays

Millipore/17-488 | phospho-p38α (Thr180/Tyr182) STAR ELISA Kit/17-488/96 assays

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货号: 17-488
品牌: Millipore
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    • Description
      CatalogueNumber17-488
      BrandFamilyUpstate
      TradeName
      • STAR
      • Upstate
      Descriptionphospho-p38α(Thr180/Tyr182)STARELISAKit
      BackgroundInformationI.TESTPRINCIPLE
      TheUPSTATE®colorimetricSTAR(SignalTransductionAssayReaction)ELISAkitisasolidphasesandwichenzymelinkedimmunosorbentassaythatprovidesafast,sensitivemethodtodetectspecificlevelsofsignalingtargetsinwholecellextracts.Thep38MAPKplateiscoatedwithaspecificmousemonoclonalp38MAPKcaptureantibodyonthemicrowellsofthe96-wellclearplate.Samplelysateorthestandardincludedinthekitareincubatedinthemicrowellsallowingp38MAPKantigentobecapturedintheplatewells.Theplateisthenwashedtoremoveanynon-boundunspecificmaterial.Thewellsarethenincubatedwithaspecificrabbitanti-phospho-p38(Thr180/Tyr182)antibodytodetectthecapturedp38(Thr180/Tyr182)ontheplatewell.TheunbounddetectionantibodyiswashedawayfollowedbyincubationwithanHRP-conjugatedanti-rabbitantibody.Thisallowsforasensitiveenzymaticdetectionofthesample.AftertheadditionofTMBsubstrateandstopsolutiontheabsorbanceismeasuredat450nmusingaplatereader.
      Theentireassaytakeslessthan5hourstocompletewithminimalhands-ontime.Manyofthereagentsaresuppliedinready-touseformulationsforeaseofuse.Thekitalsoincludesastandardthatisrunasbothapositivecontrolandtodevelopastandardlineofdetection.

      II.p38BACKGROUND
      Thep38MAPKfamilyofSer/Thrkinaseproteinsismadeupof4isoforms,p38alpha(MAPK14andSAPK2a),p38beta(MAPK11andSAPK2b),p38delta(MAPK13,SAPK4),andp38gamma(MAPK12,SAPK3).Theotherp38isoformsshare74%,57%,and60%sequenceidentitywiththealphaisoform,respectively.AllfourisoformspossestheTGYmotifandareactivatedbythedualphosphorylationofthethreonineandtyrosineresiduesinthismotif,whichcorrespondtoThr180/Tyr182inhumanp38alpha.p38,especiallythealphaandbetaisoforms,areabletophosphorylateawidearrayoftargetsincludingnumerouskinasesincludingMAPKAP-K2,MAPKAP-K3,PRAK,MSK,andMNKaswellastranscriptionfactorsincludingSTAT1,p53,NFATp,ATF-2,Elk-1(althoughweakly),andMEF2a/c.p38MAPKfamilymembershavetrADItionallybeenassociatedwithstress,immuneresponse,andregulationofcellproliferationandapoptosis.Theyalsohavearoleinregulatingtheproductionofpro-inflammatorycytokinesandhavebeenassociatedwithdiseasessuchasasthmaandseveralautoimmunediseases,includingrheumatoidarthritis,diabetes,andinflammatoryboweldisease.JustasERKisphosphorylatedandactivatedbyMEK,p38alphaMAPKinaseisactivatedpredominantlybyitsupstreamregulatorykinasesMKK6andMKK3aswellasMKK4whichalsoactivatesJNK,butisnotasubstrateforMKK7likeJNKis.
      MaterialsRequiredbutNotDelivered1.Multi-channelorrepeatingPipettes
      2.Plateshaker(optional)
      3.Pipettors&tipscapableofaccuratelymeasuring1-1000µL
      4.GraduatedSEROlogicalpipettes
      5.96-wellmicrotiterPlateReaderwith450nmfilter
      6.Graphingsoftwareforplottingdataorgraphpaperformanualplottingofdata
      7.Microfugetubesforstandardandsampledilutions
      8.Mechanicalvortex
      9.1litercontainer
      10.Distilledordeionizedwater
      ProductInformation
      Components
      • CapturePlatepre-coatedwithanti-p38alphaMAPKantibody:(PartNo.17-487A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch.
      • Anti-phospho-p38alpha(Thr180/Tyr182)detectionantibody:(PartNo.17-487B)Onebottle(11mL)ofanti-phospho-p38alpha(Thr180/Tyr182)detectionantibodycontainingsodiumazide,readytouse.
      • ELISADiluent:(PartNo.17-487C)Onebottle(25mL)ofELISADiluentcontainingsodiumazide,readytouse.
      • 25XELISAWashBuffer:(PartNo.17-487D)Onebottle(50mL)of25XELISAWashBuffer.
      • Anti-RabbitIgGHRPconjugate:(PartNo.17-487E)Onevial(125µL)of100Xanti-rabbitHRPconjugatecontainingthimerosal.
      • HRPDiluent:(PartNo.17-487F)Onebottle(25mL)ofHRPDiluentcontainingthimerosal.
      • TMBSolution:(PartNo.17-487G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse.
      • StopSolution:(PartNo.17-487H)Onebottle(25mL)ofstopsolution,readytouse.
      • phospho-p38alpha(Thr180/Tyr182)Standard:(PartNo.17-487I)Twovialsofphospho-p38alpha(Thr180/Tyr182)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-p38α(Thr180/Tyr182)STARELISAKitisusedtomeasure&quantifyp38/SAPKlevels.
      KeyApplications
      • ELISA
      BIOLOGicalInformation
      SpeciesReactivity
      • Human
      AnalytesAvailable
      • p38/SAPK
      EntrezGeneNumber
      EntrezGeneSummaryTheproteinencodedbythisgeneisamemberoftheMAPkinasefamily.MAPkinasesactasanintegrationpointformultiplebiochemicalsignals,andareinvolvedinawidevarietyofcellularprocessessuchasproliferation,differentiation,transcriptionregulationanddevelopment.Thiskinaseisactivatedbyvariousenvironmentalstressesandproinflammatorycytokines.TheactivationrequiresitsphosphorylationbyMAPkinasekinases(MKKs),oritsautophosphorylationtriggeredbytheinteractionofMAP3K7IP1/TAB1proteinwiththiskinase.ThesubstratesofthiskinaseincludetranscriptionregulatorATF2,MEF2C,andMAX,cellcycleregulatorCDC25B,andtumorsuppressorp53,whichsuggesttherolesofthiskinaseinstressrelatedtranscriptionandcellcycleregulation,aswellasingenotoxicstressresponse.Fouralternativelysplicedtranscriptvariantsofthisgeneencodingdistinctisoformshavebeenreported.
      GeneSymbol
      • MAPK14
      • PRKM14
      • RK
      • SAPK2A
      • CSBP1
      • p38
      • CSPB1
      • CSBP
      • PRKM15
      • Mxi2
      • p38ALPHA
      • MXI2
      • EXIP
      • CSBP2
      Modifications
      • Phosphorylation
      UniProtNumber
      UniProtSummaryFUNCTION:SwissProt:Q16539#Respondstoactivationbyenvironmentalstress,pro-inflammatorycytokinesandlipopolysaccharide(LPS)byphosphorylatinganumberoftranscriptionfactors,suchasELK1andATF2andseveraldownstreamkinases,suchasMAPKAPK2andMAPKAPK5.Playsacriticalroleintheproductionofsomecytokines,forexampleIL-6.MayplayaroleinstabilizationofEPOmRNAduringhypoxicstress.IsoformMxi2activationisstimulatedbymitogensandoxidativestressandonlypoorlyphosphorylatesELK1andATF2.IsoformExipmayplayaroleintheearlyonsetofapoptosis.
      COFACTOR:Magnesium.
      SIZE:360aminoacids;41293Da
      SUBUNIT:Bindstoakinaseinteractionmotifwithintheproteintyrosinephosphatase,PTPRR.ThisinteractionretainsMAPK14inthecytoplasmandpreventsnuclearaccumulation.InteractswithSPAG9(Bysimilarity).InteractswithNP60.
      SUBCELLULARLOCATION:Cytoplasm(Bysimilarity).Nucleus(Bysimilarity).
      TISSUESPECIFICITY:Brain,heart,placenta,pancreasandskeletalmuscle.Expressedtoalesserextentinlung,liverandkidney.DOMAIN:SwissProt:Q16539TheTXYmotifcontainsthethreonineandtyrosineresidueswhosephosphorylationactivatestheMAPkinases.
      PTM:DuallyphosphorylatedonThr-180andTyr-182,whichactivatestheenzyme.&PhosphorylateduponDNAdamage,probablybyATMorATR.
      SIMILARITY:Belongstotheproteinkinasesuperfamily.CMGCSer/Thrproteinkinasefamily.MAPkinasesubfamily.&Contains1proteinkinasedomain.
      PhysicochemicalInformation
      Sensitivity
      • Sensitivity:1Unit/mL.
        RangeofDetection:1.6to100Units/mL
        SpeciesReactivity:Human
      Dimensions
      MaterialsInformation
      MaterialsInformation
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