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Millipore/17-462 | Phospho-EGFR (Tyr1068) STAR Assay ELISA Kit/17-462/96 assays
  • Millipore/17-462 | Phospho-EGFR (Tyr1068) STAR Assay ELISA Kit/17-462/96 assays

Millipore/17-462 | Phospho-EGFR (Tyr1068) STAR Assay ELISA Kit/17-462/96 assays

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货号: 17-462
品牌: Millipore
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    • Description
      CatalogueNumber17-462
      BrandFamilyUpstate
      TradeName
      • STAR
      • Upstate
      DescriptionPhospho-EGFR(Tyr1068)STARAssayELISAKit
      AlternateNames
      • Epidermalgrowthfactorreceptor
      • Proto-oncogenec-ErbB-1
      • Receptortyrosine-proteinkinaseerbB-1
      • EGFReceptor
      • EGFR
      • ERBB
      • ERBB2
      BackgroundInformationI.TESTPRINCIPLE
      TheUPSTATE®colorimetricSTAR(SignalTransductionAssayReaction)ELISAkitisasolidphasesandwichenzymelinkedimmunosorbentassaythatprovidesafast,sensitivemethodtodetectspecificlevelsofsignalingtargetsinwholecellextracts.TheEGFRplateiscoatedwithaspecificmousemonoclonalEGFRcaptureantibody.SamplelysateorthestandardincludedinthekitareincubatedinthemicrowellsallowingEGFRantigentobecaptured.Theplateisthenwashedtoremoveanyunboundnon-specificmaterial.Aspecificrabbitanti-phospho-EGFRantibodyisaddedtodetectthecapturedphosphorylaterEGFR(Tyr1068)ontheplatewell.TheunbounddetectionantibodyiswashedawayfollowedbyincubationwithanHRP-conjugatedanti-rabbitantibody.AftertheadditionofTMBsubstrateandstopsolutiontheabsorbanceismeasuredat450nmusingaplatereader.Thisallowsforasensitiveenzymaticdetectionofthesample.
      Theentireassaytakeslessthan5hourstocompletewithminimalhands-ontime.Manyofthereagentsaresuppliedinready-touseformulationsforeaseofuse.ThekitalsoincludesastandardthatisrunasbothapositivecontrolandtogenerateastandardcurveforphosphorylatedEGFR(Tyr1068)measurement.

      II.EGFRBACKGROUND

      Epidermalgrowthfactorreceptor(EGFR)isareceptortyrosinekinasethatplaysakeyroleintheregulationofessentialnormalcellularprocessesandinthepathophysiologyofhyperproliferativediseasessuchascancer.EGFRisareceptortyrosinekinaseknowntobeessentialformediationofbothproliferativeandsurvivalsignalstocells.ActivationoftheEGFRsignalingpathwayhasbeenlinkedwithincreasedcellproliferation,angiogenesis,metastasis,anddecreasedapoptosis.EGFRisfoundinmostsolidtumorsanditsautophosphorylationactivatestheintrinsickinaseactivitytowardheterologoussubstrates,aswellascreatingdockingsitesforadapterproteinsformultipleadapterproteinstobindandnucleatesignalingcomplexesthatactivatetheRas,PI3Kinase,andPLCpathways.ManyoftheEGFRmutationsaredirectlyrelatedtovariousdiseasesandarethetargetofnumerousdrugs.Forinstance,theaminoacidsubstitutionL858RisoneofseveralheterozygousmutationsthathavebeenidentifiedinNon-Small-CellLungCancer(NSCLC)patientswhohaveclinicalresponsestotheEGFRinhibitorIressa®.ThereissomeevidencethatthesemutationsresultinelevatedactivityandenhancedsensitivitytoIressa®.InpatientswithtumorsbearingIressa®-sensitivemutations,resistantsubclonescontaininganadditionalEGFRmutation,T790M,emergeinthepresenceofthedrug.IthasbeenshownexperimentallythattheT790Mmutationleadstohigh-levelfunctionalresistancetoIressa®.InpatientswithtumorsbearingIressa®-sensitivemutations(eg.L858R,L861Q),resistantsubclonescontainingtheT790Mmutationemergeinthepresenceofthedrug.
      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
      • CapturePlatepre-coatedwithanti-humanEGFRantibody:(PartNo.17-462A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch.
      • Anti-EGFR(Tyr1068)detectionantibody:(PartNo.17-462B)Onebottle(11mL)ofanti-EGFR(Tyr1068)detectionantibodycontainingsodiumazide,readytouse.
      • ELISADiluent:(PartNo.17-462C)Onebottle(25mL)ofELISADiluentcontainingsodiumazide,readytouse.
      • 25XELISAWashBuffer:(PartNo.17-462D)Onebottle(50mL)of25XELISAWashBuffer.
      • Anti-RabbitIgGHRPconjugate:(PartNo.17-462E)Onevial(125®L)of100Xanti-rabbitHRPconjugate.
      • HRPDiluent:(PartNo.17-462F)Onebottle(25mL)ofHRPDiluent.
      • TMBSolution:(PartNo.17-462G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse.
      • StopSolution:(PartNo.17-462H)Onebottle(25mL)ofstopsolution,readytouse.
      • PhosphorylatedEGFR(Tyr1068)Standard:(PartNo.17-462I)TwovialsofphosphorylatedEGFR(Tyr1068)standard,lyophilized.
      • PlateCovers:Twoplatecovers.
      DetectionmethodChromogenic
      StorageandShippingInformation
      StorageConditionsMaintaintheunopenedkitat2-8°Cuntilexpirationdate.
      Applications
      ApplicationThisPhospho-EGFR(Tyr1068)STARAssayELISAKitisusedtomeasure&quantifyEGFRlevels.
      KeyApplications
      • ELISA
      BIOLOGicalInformation
      SpeciesReactivity
      • Human
      AnalytesAvailable
      • EGFR
      EntrezGeneNumber
      GeneSymbol
      • EGFR
      • ERBB1
      • ERBB
      • mENA
      Modifications
      • Phosphorylation
      UniProtNumber
      UniProtSummaryFUNCTION:SwissProt:P00533#Isoform2/truncatedisoformmayactasanantagoNIST.
      SIZE:1210aminoacids;134277Da
      SUBUNIT:BindsRIPK1.CBLinteractswiththeautophosphorylatedC-terminaltailoftheEGFreceptor.PartofacomplexwithERBB2andeitherPIK3C2AorPIK3C2B.TheautophosphorylatedforminteractswithPIK3C2B,maybeindirectly.InteractswithPELP1.
      SUBCELLULARLOCATION:Cellmembrane;Single-passtypeImembraneprotein.&Isoform2:Secreted.
      TISSUESPECIFICITY:Expressedinplacenta.Isoform2isalsoexpressedinovariancancers.
      PTM:PhosphorylationofSer-695ispartialandoccursonlyifThr-693isphosphorylated.&MonoubiquitinatedandpolyubiquitinateduponEGFstimulation;whichdoesnotaffecttyrosinekinaseactivityorsignalingcapacitybutmayplayaroleinlysosomaltargeting.Polyubiquitinlinkageismainlythrough"Lys-63",butlinkagethrough"Lys-48","Lys-11"and"Lys-29"alsooccur.DISEASE:SwissProt:P00533#DefectsinEGFRareassociatedwithlungcancer[MIM:211980].
      SIMILARITY:SwissProt:P00533##Belongstotheproteinkinasesuperfamily.Tyrproteinkinasefamily.EGFreceptorsubfamily.&Contains1proteinkinasedomain.
      MISCELLANEOUS:BindingofEGFtothereceptorleadstodimerization,internalizationoftheEGF-receptorcomplex,inductionofthetyrosinekinaseactivity,stimulationofcellDNAsynthesis,andcellproliferation.
      PhysicochemicalInformation
      Sensitivity
      • Sensitivity:1unit/mL
        RangeofDetection:1.6to100units/mL
      Dimensions
      MaterialsInformation
      MaterialsInformation
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