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Millipore/17-461 | Phospho-EGFR (Tyr1173) STAR ELISA Kit/17-461/96 assays
  • Millipore/17-461 | Phospho-EGFR (Tyr1173) STAR ELISA Kit/17-461/96 assays

Millipore/17-461 | Phospho-EGFR (Tyr1173) STAR ELISA Kit/17-461/96 assays

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貨號: 17-461
品牌: Millipore
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    • Description
      CatalogueNumber17-461
      BrandFamilyUpstate
      TradeName
      • STAR
      • Upstate
      DescriptionPhospho-EGFR(Tyr1173)STARELISAKit
      AlternateNames
      • Epidermalgrowthfactorreceptor
      • Proto-oncogenec-ErbB-1
      • Receptortyrosine-proteinkinaseerbB-1
      • EGFReceptor
      • EGFR
      • ERBB
      • ERBB1
      BackgroundInformationI.TESTPRINCIPLE
      TheUPSTATEcolorimetricSTAR(SignalTransductionAssayReaction)ELISAkitisasolidphasesandwichenzymelinkedimmunosorbentassaythatprovidesafast,sensitivemethodtodetectspecificlevelsofsignalingtargetsinwholecellextracts.TheEGFRplateiscoatedwithaspecificmousemonoclonalanti-EGFRcaptureantibodyonthemicrowellsofthe96-wellclearplate.SamplelysateorthestandardincludedinthekitareincubatedinthemicrowellsallowingEGFRantigentobecapturedintheplatewells.Theplateisthenwashedtoremoveanynon-boundunspecificmaterial.Thewellsarethenincubatedwithaspecificrabbitanti-EGFRantibodytodetectthecapturedphospho-EGFR(Tyr1173)ontheplatewell.TheunbounddetectionantibodyiswashedawayfollowedbyincubationwithanHRP-conjugatedanti-rabbitantibody.Thisallowsforasensitiveenzymaticdetectionofthesample.AftertheadditionofTMBsubstrateandstopsolutiontheabsorbanceismeasuredat450nmusingaplatereader.Theentireassaytakeslessthan5hourstocompletewithminimalhands-ontime.Manyofthereagentsaresuppliedinready-touseformulationsforeaseofuse.Thekitalsoincludesastandardthatisrunasbothapositivecontrolandtodevelopastandardcurve.

      II.EGFRBACKGROUND

      Epidermalgrowthfactorreceptor(EGFR)isareceptortyrosinekinasethatplaysakeyroleintheregulationofessentialnormalcellularprocessesandinthepathophysiologyofhyperproliferativediseasessuchascancer.Epidermalgrowthfactorreceptor(EGFR)isareceptortyrosinekinaseknowntobeessentialformediationofbothproliferativeandsurvivalsignalstocells.ActivationoftheEGFRsignalingpathwayhasbeenlinkedwithincreasedcellproliferation,angiogenesis,metastasisanddecreasedapoptosis.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.Pipettors&tipscapableofaccuratelymeasuring1-1000%micro;L
      4.GraduatedSEROlogicalpipettes
      5.96-wellmicrotiterPlateReaderwith450nmfilter
      6.Graphingsoftwareforplottingdataorgraphpaperformanualplottingofdata
      7.Microfugetubesforstandardandsampledilutions
      8.Mechanicalvortex
      9.1litercontainer
      10.Distilledordeionizedwater
      ProductInformation
      Components
      • 1.CapturePlatepre-coatedwithanti-humanEGFRantibody:(PartNo.17-461A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch.
      • 2.Anti-EGFR(Tyr1173)detectionantibody:(PartNo.17-461B)Onebottle(11mL)ofanti-EGFR(Tyr1173)detectionantibodycontainingsodiumazide,readytouse.
      • 3.ELISADiluent:(PartNo.17-461C)Onebottle(25mL)ofELISADiluentcontainingsodiumazide,readytouse.
      • 4.25XELISAWashBuffer:(PartNo.17-461D)Onebottle(50mL)of25XELISAWashBuffer.
      • 5.Anti-RabbitIgGHRPconjugate:(PartNo.17-461E)Onevial(125μL)of100Xanti-rabbitHRPconjugatecontainingthimerosol.
      • 6.HRPDiluent:(PartNo.17-461F)Onebottle(25mL)ofHRPDiluentcontainingthimerosol.
      • 7.TMBSolution:(PartNo.17-461G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse.
      • 8.StopSolution:(PartNo.17-461H)Onebottle(25mL)ofstopsolution,readytouse.
      • 9.EGFR(Tyr1173)Standard:(PartNo.17-461I)TwovialsofEGFR(Tyr1173)standard,lyophilized.
      • 10.PlateCovers:Twoplatecovers.
      DetectionmethodChromogenic
      StorageandShippingInformation
      StorageConditions1yearat4°C
      Applications
      KeyApplications
      • ELISA
      BIOLOGicalInformation
      SpeciesReactivity
      • Human
      • Mouse
      • Rat
      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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