Description:
Platelet-derivedgrowthfactor(PDGF)isagrowthfactorthatisimportanttoembryonicdevelopment,cellproliferation,andcellmigration,particularly,inbloodvesselformation(angiogenesis).LikeVEGF,PDGFisabletoindependentlyinitiateangiogenesisandmediatebloodvesselgrowthandbehavior.TheproteinisadimericglycoproteincomposedoftwoA(-AA)ortwoB(-BB)chainsoracombinationofthetwo(-AB).IthasbeenshownthatthecisoncogeneisderivedfromthePDGFB-chaingene.ItbindstoPDGFreceptors,whichactivatesignaltransductionpathwayssuchasthePI3Kpathway,andsubsequentlyregulatedownstreamgeneexpressionandthecellcycle.PDGFhasalsobeenlinkedtoseveraldiseasessuchasatherosclerosis,fibrosisandmalignantdiseases.Principle:
TheELISAisbasedontheprincipleofasolidphaseenzyme-linkedimmunosorbentassay.Theassayutilizesamouseanti-humancytokineantibodyforimmobilizationonthemicrotiterwellsandgoatanti-humancytokineantibodiesalongwithstreptavidinconjugatedtohorserADIshperoxidase(HRP)fordetection.Thetestsampleisallowedtoreactsimultaneouslywiththetwoantibodies,resultinginthemoleculesofthecytokinebeingsandwichedbetweenthesolidphaseandenzyme-linkedantibodies.Afterincubation,thewellsarewashedtoremoveunbound-labeledantibodies.AHRPsubstrate,TMB,isaddedtoresultinthedevelopmentofabluecolor.ThecolordevelopmentisthenstoppedwiththeadditionofStopSolutionchangingthecolortoyellow.Theconcentrationofthecytokineisdirectlyproportionaltothecolorintensityofthetestsample.Absorbanceismeasuredspectrophotometricallyat450nm. |
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