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    當前位置 ： Millipore >>> Millipore/17-484 | phospho-IR (β Subunit) (Tyr 1162/1163) STAR ELISA Assay Kit/17-484/96 assays
    Millipore/17-484 | phospho-IR (β Subunit) (Tyr 1162/1163) STAR ELISA Assay Kit/17-484/96 assays
    • Millipore/17-484 | phospho-IR (β Subunit) (Tyr 1162/1163) STAR ELISA Assay Kit/17-484/96 assays

    Millipore/17-484 | phospho-IR (β Subunit) (Tyr 1162/1163) STAR ELISA Assay Kit/17-484/96 assays

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    貨號: 17-484
    品牌: Millipore
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      • Description
        CatalogueNumber17-484
        BrandFamilyUpstate
        TradeName
        • STAR
        • Upstate
        Descriptionphospho-IR(βSubunit)(Tyr1162/1163)STARELISAAssayKit
        AlternateNames
        • InsulinReceptor
        • IR
        BackgroundInformationI.TESTPRINCIPLE
        TheUPSTATE?colorimetricSTAR(SignalTransductionAssayReaction)ELISAkitisasolidphasesandwichenzymelinkedimmunosorbentassaythatprovidesafast,sensitivemethodtodetectspecificlevelsofsignalingtargetsinwholecellextracts.TheIRplateiscoatedwithaspecificmousemonoclonalIR(βsubunit)captureantibodyonthemicrowellsofthe96-wellclearplate.SamplelysateorthestandardsincludedinthekitareincubatedinthemicrowellsallowingIRtobecapturedintheplatewells.Theplateisthenwashedtoremoveanyunboundnon-specificmaterial.Thewellsarethenincubatedwithaspecificrabbitanti-phospho-IRantibodytodetectthecapturedIRphosphorylatedonTyrosines1162and1163.TheunbounddetectionantibodyiswashedawayfollowedbyincubationwithanHRP-conjugatedanti-rabbitantibody.Thisallowsforasensitiveenzymaticdetectionofthesample.AftertheadditionofTMBsubstrateandstopsolutiontheabsorbanceismeasuredat450nmusingaplatereader.
        Theentireassaytakeslessthan5hourstocompletewithminimalhands-ontime.Manyofthereagentsaresuppliedinready-touseformulationsforeaseofuse.Thekitalsoincludesastandardthatisrunasbothapositivecontrolandtodevelopastandardcurve.

        II.IRBACKGROUND
        TheInsulinReceptor(IR)issynthesizedasasinglepolypeptide,whichissubsequentlycleavedtogenerateanextracellularαchainandatransmembraneandintracellularβchain,tetheredtogetherbydisulfidebonds.Theβchainhasmultipletyrosinephosphorylationsites,includingthreeautophosphorylationsitesatitsactivationloop.Insulinstimulatesthesignalingcascadebybindingtotheinsulinreceptorthroughitsextracellularαsubunits.Thisstimulatesthetyrosinekinaseactivityoftheβsubunitsofthereceptor.ThereceptorthenautophosphorylatesitselfandphosphorylatestheIRS(InsulinReceptorSubstrate)proteinsthatareveryimportantmodulatorsofinsulinsignaling.PhosphorylatedIRS,inturn,interactswithseveraldownstreameffectorsofinsulinsignaling,includingPI3Kinasep85,GRB2,SHP2,Nck,Crk,andFyn.TheoverallstructureoftheIRishighlyhomologoustotheIGF-IReceptor,exceptintheirc-termini,wherethetwoproteinsdivergesomewhat.InsulinsignalingishighlydependentonthePI3KinasepathwayandAkt,whichappeartomediatethefunctionsofinsulin.
        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-IRantibody:(PartNo.17-483A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch.
        • Anti-phospho-IR(Tyr1162/1163)detectionantibody:(PartNo.17-483B)Onebottle(11mL)ofanti-phospho-IR(Tyr1162/1163)detectionantibodycontainingsodiumazide,readytouse.
        • ELISADiluent:(PartNo.17-483C)Onebottle(25mL)ofELISADiluentcontainingsodiumazide,readytouse.
        • 25XELISAWashBuffer:(PartNo.17-483D)Onebottle(50mL)of25XELISAWashBuffer.
        • Anti-RabbitIgGHRPconjugate:(PartNo.17-483E)Onevial(125μL)of100Xanti-rabbitHRPconjugatecontainingthimerosal.
        • HRPDiluent:(PartNo.17-483F)Onebottle(25mL)ofHRPDiluentcontainingthimerosal.
        • TMBSolution:(PartNo.17-483G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse.
        • StopSolution:(PartNo.17-483H)Onebottle(25mL)ofstopsolution,readytouse.
        • phospho-IRStandard:(PartNo.17-483I)TwovialsofIR((Tyr1162/1163)standard,lyophilized.
        • PlateCovers:Twoplatecovers.
        DetectionmethodChromogenic
        StorageandShippingInformation
        StorageConditionsMaintaintheunopenedkitat2-8°Cuntilexpirationdate.
        Applications
        ApplicationThisphospho-IR(βSubunit)(Tyr1162/1163)STARELISAAssayKitisusedtomeasure&quantifyInsulinlevels.
        KeyApplications
        • ELISA
        BIOLOGicalInformation
        SpeciesReactivity
        • Human
        • Mouse
        • Rat
        AnalytesAvailable
        • Insulin
        EntrezGeneNumber
        EntrezGeneSummaryAfterremovaloftheprecursorsignalpeptide,theinsulinreceptorprecursorispost-translationallycleavedintotwochains(alphaandbeta)thatarecovalentlylinked.Bindingofinsulintotheinsulinreceptor(INSR)stimulatesglucoseuptake.Twotranscriptvariantsencodingdifferentisoformshavebeenfoundforthisgene.
        GeneSymbol
        • INSR
        • CD220
        • IR
        • HHF5
        Modifications
        • Phosphorylation
        UniProtNumber
        UniProtSummaryFUNCTION:SwissProt:P06213#Thisreceptorbindsinsulinandhasatyrosine-proteinkinaseactivity.IsoformShorthasahigheraffinityforinsulin.Mediatesthemetabolicfunctionsofinsulin.BindingtoinsulinstimulatesassociationofthereceptorwithdownstreammediatorsincludingIRS1andphosphatidylinositol3"-kinase(PI3K).CanactivatePI3Keitherdirectlybybindingtothep85regulatorysubunit,orindirectlyviaIRS1.
        SIZE:1382aminoacids;156307Da
        SUBUNIT:Tetramerof2alphaand2betachainslinkedbydisulfidebonds.Thealphachainscontributetotheformationoftheligand-bindingdomain,whilethebetachainscarrythekinasedomain.InteractswithSORBS1butdissociatesfromitfollowinginsulinstimulation.BindsSH2B2.InteractswiththePTB/PIDdomainsofIRS1andSHC1invitrowhenautophosphorylatedontyrosineresidues.ThesequencessurroundingthephosphorylatedNPXYmotifcontributedifferentiallytoeitherIRS1orSHC1recognition.InteractswiththeSH2domainsofthe85kDaregulatorysubunitofPI3K(PIK3R1)invitro,whenautophosphorylatedontyrosineresidues.InteractswithSOCS7.
        SUBCELLULARLOCATION:Membrane;Single-passtypeImembraneprotein.
        TISSUESPECIFICITY:IsoformLongandisoformShortareexpressedintheperipheralnerve,kidney,liver,striatedmuscle,fibroblastsandskin.IsoformShortisexpressedalsointhespleenandlymphoblasts.
        PTM:AfterbeingtransportedfromtheendoplasmicreticulumtotheGolgiapparatus,thesingleglycosylatedprecursorisfurtherglycosylatedandthencleaved,followedbyitstransporttotheplasmamembrane.&Autophosphorylatedontyrosineresiduesinresponsetoinsulin.&PhosphorylationofTyr-999isrequiredforIRS1-andSHC1-binding.
        DISEASE:SwissProt:P06213#DefectsinINSRarethecauseofinsulinresistance(Insresistance)[MIM:125853].&DefectsinINSRarethecauseofRabson-Mendenhallsyndrome[MIM:262190];alsoknownasMendenhallsyndrome.Itisasevereinsulinresistancesyndromecharacterizedbyinsulin-resistantdiabetesmellituswithpinealhyperplasiaandsomaticabnormalities.Typicalfeaturesincludecoarse,senile-appearingfacies,dentalandskinabnormalities,aBDominaldistension,andphallicenlargement.Inheritanceisautosomalrecessive.&DefectsinINSRarethecauseofleprechaunism[MIM:246200];alsoknownasDonohuesyndrome.Leprechaunismrepresentsthemostsevereformofinsulinresistancesyndrome,characterizedbyintrauterineandpostnatalgrowthretardationanddeathinearlyinfancy.Inheritanceisautosomalrecessive.&DefectsinINSRmaybeassociatedwithnoninsulin-dependentdiabetesmellitus(NIDDM)[MIM:125853];alsoknownasdiabetesmellitustype2.&DefectsinINSRarethecauseoffamilialhyperinsulinemichypoglycemia5(HHF5)[MIM:609968].Familialhyperinsulinemichypoglycemia[MIM:256450],alsoreferredtoascongenitalhyperinsulinism,nesidioblastosis,orpersistenthyperinsulinemichypoglycemiaofinfancy(PPHI),isthemostcommoncauseofpersistenthypoglycemiaininfancyandisduetodefectivenegativefeedbackregulationofinsulinsecretionbylowglucoselevels.&DefectsinINSRarethecauseofinsulin-resistantdiabetesmellituswithacanthosisnigricanstypeA(IRANtypeA)[MIM:610549].Thissyndromeischaracterizedbytheassociationofsevereinsulinresistance(manifestedbymarkedhyperinsulinemiaandafailuretorespondtoexogenousinsulin)withtheskinlesionacanthosisnigricansandovarianhyperandrogenisminadolescentfemalesubjects.Womenfrequentlypresentwithhirsutism,acne,amenorrheaoroligomenorrhea,andvirilization.ThissyndromeisdifferentfromthetypeBthathasbeendemonstratedtobesecondarytothepresenceofcirculatingautoantibodiesagainsttheinsulinreceptor.
        SIMILARITY:SwissProt:P06213##Belongstotheproteinkinasesuperfamily.Tyrproteinkinasefamily.Insulinreceptorsubfamily.&Contains2fibronectintype-IIIdomains.&Contains1proteinkinasedomain.
        PhysicochemicalInformation
        Sensitivity
        • Sensitivity:1Unit/mL
          RangeofDetection:1.6-100Units/mL
        Dimensions
        MaterialsInformation
        MaterialsInformation
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