ProductHighlight- Configuration-specificmonoclonalantibodythatrecognizesGαi-GTP,butnotGαi-GDP
- TheonlyavailableGαi activationassaykitdirectlymeasuresthelevelofGαi-GTP
- ProvidingreliableresultswithconsistentreproducibilitythanotherGαi activationassaykitsbasedoncertaindownstreamsignalingpathways
- Providingshorterexperimentaltime
- CouldbeusedtomonitortheactivationofGαi incellsandintissuesbyimmunohistochemistry
ProductDescriptionAstructurallydiverserepertoireofligands,fromphotonstolargepeptides,activatesGprotein-coupledreceptors(GPCRs)toelicittheirphysiologicalfunctions.Ligand-boundGPCRs,inturn,functionasguaninenucleotideexchangefactorscatalyzingtheexchangeofGDPboundontheGαsubunitwithGTPinthepresenceofGβγ,causingthedissociationoftheGαsubunitfromtheGβγdimertoformtwofunctionalunits(GαandGβγ).BothGαandGβγsubunitssignaltovariouscellularsignalingpathways.Basedonthesequenceandfunctionalhomologies,Gproteinsaregroupedintofourfamilies:Gs,Gi,Gq,andG12. Gαi familyisthelargestfamilyofGproteins.TheyrelaysignalsfrommanyGPCRstoregualtevariousBIOLOGicalfunctions.TherewerenodirectmethodstomeasuretheactivationofGαi proteinsbyreceptors(untilthisassaykit).Mostreportsusedoneofthedownstreampathway,i.e.theinhibitionofadenylylcyclases,asareadout.Alternatively,sensitivitytopertussistoxin(PTX)wasusedasanindicatorofpossIBLeGαi proteinsinvovledinasignalingpathway. NewEastBiosciencesGαi ActivationAssayKitprovidesadirectmeasurementoftheactivationofGαi proteins.ThisisasimpleandfasttooltomonitortheactivationofGαi.Eachkitprovidessufficientquantitiestoperform20assays. NewEastBiosciencesGαi ActivationAssayKitisbasedonthemonoclonalantibodyspecificallyrecognizingtheactiveGTP-boundGαi proteins.ThismonoclonalantibodyhasmuchloweraffinitytowardstheinactiveGαi proteins.Therefore,afteractivationbyreceptorsignals,activeGTP-boundGαi proteinscouldbeimmunoprecipitatedbythismonoclonalantibodyandfurtherquantifiedbywesternblotwithanotheranti-Gαi antibody. AssayPrincipleNewEastBiosciencesGαi ActivationAssayKitisanimmunoprecipitation/westernblotassaytomeasurethelevelsofactiveGTP-boundGαi proteins,eitherfromcellextractsorfrominvitroGTPγSloadedGαiproteins.Briefly,theanti-activeGαi monoclonalantibodywillspecificallybindtoactiveGαi protein.Thisantibody/Gαi complexwillthenbepulleddownbyproteinA/Gagarose.TheprecipitatedactiveGαi proteinswillbedetectedbyimmunoblotswithanotheranti-Gαi antibody. |