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Millipore/17-479 | STAR FAK ELISA Kit/17-479/96 assays

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貨號: 17-479

品牌: Millipore

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Description
CatalogueNumber	17-479
BrandFamily	Upstate
TradeName	STAR Upstate
Description	STARFAKELISAKit
AlternateNames	Focaladhesionkinase1 FADK1 Focaladhesionkinase-relatednonkinase FRNK Proteinphosphatase1regulatorysubunit71 PPP1R71 Protein-tyrosinekinase2 p125FAK pp125FAK
BackgroundInformation	I.TESTPRINCIPLE TheUPSTATE?colorimetricSTAR(SignalTransductionAssayReaction)ELISAkitisasolidphasesandwichenzymelinkedimmunosorbentassaythatprovidesafast,sensitivemethodtodetectspecificlevelsofsignalingtargetsinwholecellextracts.TheFAKplateiscoatedwithaspecificmousemonoclonalFAKcaptureantibody.SamplelysateorthestandardincludedinthekitareincubatedinthemicrowellsallowingFAKantigentobecaptured.Theplateisthenwashedtoremoveanyun-boundnon-specificmaterial.Aspecificrabbitanti-FAKantibodyisaddedtodetectthecapturedFAKontheplate.TheunbounddetectionantibodyiswashedawayfollowedbyincubationwithanHRP-conjugatedanti-rabbitantibody.AftertheadditionofTMBsubstrateandstopsolutiontheabsorbanceismeasuredat450nmusingaplatereader.Thisallowsforasensitiveenzymaticdetectionofthesample. Theentireassaytakeslessthan5hourstocompletewithminimalhands-ontime.Manyofthereagentsaresuppliedinready-touseformulationsforeaseofuse.ThekitalsoincludesastandardthatisrunasbothapositivecontrolandtogenerateastandardcurveforFAKmeasurement. II.FAKBACKGROUND FAK(FocalAdhesionKinase)isanon-receptortyrosinekinasewhichlocalizestofocaladhesionsandplaysakeyroleinintegrin-mediatedsignalingpathwaysregulatingcellmigration.Itautophosphorylatesontyrosine397duringadhesionandspreADIng,andisthoughttobeacriticaltransducerofadhesion-dependentgrowthandsurvival.FAKanditphosphorylationstateshavebeenimplicatedincancermetastasisandtumorcellsurvivalandadhesion-independentgrowth.Additionally,recentevidenceindicatesthatelevationofFAKactivityinhumancarcinomacellsisassociatedwithincreasedinvasivepotential.AcentralroleintumorformationandprogressionsuggestthatFAKisanattractivetargetfortherapeuticintervention. Cellsurfaceintegrinswhenengagedwiththeirligandsleadtotherecruitmentofanumberofintracellularproteinstospecializedsitesofthecytoplasmicfaceintofocaladhesions.Thesefocaladhesionplaquesarecomposedofmanyproteinsthatincludekinases,phosphatases,scaffoldingproteins,andG-proteins.Theactivationofintegrinsleadstothetyrosinephosphorylationofseveralcytoplasmicproteinscriticaltothebiochemicalprocess,includingFAK,Src,paxillin,tensin,andp130Cas.Thisclusteringandactivationofproteinsatthemembraneleadstotheactivationofmanydivergingsignalingpathwaysthatresultincytoskeletalre-organization.ThisstartswitheithertherecruitmentofSrcandFAKtotheplasmamembranefollowingintegrinorPI3Kinaseactivation.Thenon-receptorprotein-tyrosinekinases(PTKs)FAKandSrcarekeymembersofthefocaladhesioncomplex.ThesePTKsco-Localizeandassociatewiththetransmembraneintegrinsandvariousdownstreamtargetsofcertaingrowthfactorreceptors.TheoutcomeofSrcbindingtoFAKisthephosphorylationofseveraltyrosineresiduesinFAKanditsrelatedproteins,includingtheFAKautophosphorylationsiteTyr397.ThisautophosphorylationoccursimmediatelyafterintegrinclusteringandallowsfortheassociationofFAKwithvarioussignalingproteinssuchasPI3kinase,Grb2-Sos,p130Cas,andpaxillin.ThephosphorylationofFAKTyr397iscrucialformanyoftheestablishedBIOLOGicalrolesofFAK,includingcellmigration,cellcycleprogression,andpreventionofanoikis,aformofdetachment-inducedapoptosisandiscorrelatedwiththeinvasivenessoftumorsandisthoughttobeapivotalplayerbymodulatingtheturnoveroffocaladhesionstoregulatecellmigration.Phosphoinositide3-Kinase(PI3kinaseorPI3K)alsoplaysaroleincellmigrationviaitsbindingtotheautophosphorylationsite(Tyr397)ofFAKthroughoneorbothofitsSH2domainsinthep85subunitofPI3K.ThisisthesamesiteinFAKthatisboundbySrc.Intheory,thebindingofPI3kinasetoFAKmaybeinvolvedinactivitieslikecellproliferation,apoptosis,andmigrationthatarerelatedtothephosphorylationofTyr397.ThisissubstantiatedbytheobservationthatAKTisdownstreamofPI3kinaseandisamediatorinpreventingapoptosis.ThereforeitispossIBLethatFAK/PI3kinaseassociationmayhelpregulateapoptosis.
MaterialsRequiredbutNotDelivered	1.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-FAKantibody:(PartNo.17-479A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch. Anti-FAKdetectionantibody:(PartNo.17-479B)Onebottle(11mL)ofanti-FAKdetectionantibodycontainingsodiumazide,readytouse. ELISADiluent:(PartNo.17-479C)Onebottle(25mL)ofELISADiluentcontainingsodiumazide,readytouse. 25XELISAWashBuffer:(PartNo.17-479D)Onebottle(50mL)of25XELISAWashBuffer. Anti-RabbitIgGHRPconjugate:(PartNo.17-479E)Onevial(125μL)of100Xanti-rabbitHRPconjugatecontainingthimerosal. HRPDiluent:(PartNo.17-479F)Onebottle(25mL)ofHRPDiluentcontainingthimerosal. TMBSolution:(PartNo.17-479G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse. StopSolution:(PartNo.17-479H)Onebottle(25mL)ofstopsolution,readytouse. FAKStandard:(PartNo.17-479I)TwovialsofFAKstandard,lyophilized. PlateCovers:Twoplatecovers.
Detectionmethod	Chromogenic

ProductInformation

Components

CapturePlatepre-coatedwithanti-FAKantibody:(PartNo.17-479A)Onepre-coated96-stripwellimmunoplatesealedinafoilpouch.
Anti-FAKdetectionantibody:(PartNo.17-479B)Onebottle(11mL)ofanti-FAKdetectionantibodycontainingsodiumazide,readytouse.
ELISADiluent:(PartNo.17-479C)Onebottle(25mL)ofELISADiluentcontainingsodiumazide,readytouse.
25XELISAWashBuffer:(PartNo.17-479D)Onebottle(50mL)of25XELISAWashBuffer.
Anti-RabbitIgGHRPconjugate:(PartNo.17-479E)Onevial(125μL)of100Xanti-rabbitHRPconjugatecontainingthimerosal.
HRPDiluent:(PartNo.17-479F)Onebottle(25mL)ofHRPDiluentcontainingthimerosal.
TMBSolution:(PartNo.17-479G)Onebottle(25mL)ofstABIlizedtetramethylbenzidine(TMB),readytouse.
StopSolution:(PartNo.17-479H)Onebottle(25mL)ofstopsolution,readytouse.
FAKStandard:(PartNo.17-479I)TwovialsofFAKstandard,lyophilized.
PlateCovers:Twoplatecovers.

Detectionmethod

Chromogenic

StorageandShippingInformation
StorageConditions	Maintaintheunopenedkitat2-8°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.

StorageandShippingInformation

StorageConditions

Maintaintheunopenedkitat2-8°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
Application	ThisSTARFAKELISAKitisusedtomeasure&quantifyFAKlevels.
KeyApplications	ELISA

BiologicalInformation
SpeciesReactivity	Human Mouse Rat
AnalytesAvailable	FAK
EntrezGeneNumber	NM_005607.3 NM_153831.2
EntrezGeneSummary	Thisgeneencodesacytoplasmicproteintyrosinekinasewhichisfoundconcentratedinthefocaladhesionsthatformbetweencellsgrowinginthepresenceofextracellularmatrixconstituents.TheencodedproteinisamemberoftheFAKsubfamilyofproteintyrosinekinasesbutlackssignificantsequencesimilaritytokinasesfromothersubfamilies.Activationofthisgenemaybeanimportantearlystepincellgrowthandintracellularsignaltransductionpathwaystriggeredinresponsetocertainneuralpeptidesortocellinteractionswiththeextracellularmatrix.Atleastfourtranscriptvariantsencodingfourdifferentisoformshavebeenfoundforthisgene,butthefull-lengthnaturesofonlytwoofthemhavebeendetermined.
GeneSymbol	PTK2 pp125FAK FADK FAK1 FAK
UniProtNumber	Q05397
UniProtSummary	FUNCTION:SwissProt:Q05397#Non-receptorprotein-tyrosinekinaseimplicatedinsignalingpathwaysinvolvedincellmotility,proliferationandapoptosis.Activatedbytyrosine-phosphorylationinresponsetoeitherintegrinclusteringinducedbycelladhesionorantibodycross-linking,orviaG-proteincoupledreceptor(GPCR)occupancybyligandssuchasbombesinorlysophosphatidicacid,orviaLDLreceptoroccupancy.Playsapotentialroleinoncogenictransformationsresultinginincreasedkinaseactivity. SIZE:1052aminoacids;119233Da SUBUNIT:InteractswithCASfamilymembersandwithGIT1,SORBS1andBCAR3.InteractswithSHB(Bysimilarity).InteractswithTGFB1I1. SUBCELLULARLOCATION:Celljunction,focaladhesion;Cytoplasmicside.Note=Constituentoffocaladhesions. TISSUESPECIFICITY:Expressedinallorganstested,inlymphoidcelllines,butmostabundantlyinbrain. DOMAIN:SwissProt:Q05397ThefirstPro-richdomaininteractswiththeSH3domainofCRK-associatedsubstrate(BCAR1)andCASL(Bysimilarity).&Thecarboxy-terminalregionisthesiteoffocaladhesiontargeting(FAT)sequencewhichmediatesthelocalizationofFAK1tofocaladhesions. PTM:Phosphorylatedon6tyrosineresiduesuponactivation. SIMILARITY:Belongstotheproteinkinasesuperfamily.Tyrproteinkinasefamily.FAKsubfamily.&Contains1FERMdomain.&Contains1proteinkinasedomain.

PhysicochemicalInformation
Sensitivity	Sensitivity:1.6ng/mL RangeofDetection:1.6to100ng/mL SpeciesReactivity:Human,mouseandrat

Dimensions

MaterialsInformation

MaterialsInformation

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Millipore/17-479 | STAR FAK ELISA Kit/17-479/96 assays

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