Formulation25mMNaCitrate,100mMNaCl,100mMGlycine,pH6.8Storage-80°CPurity>95%bySDS-PAGEActivityDeterminationN/AShelfLife(properlystored)12monthsMultimericStructureOfVonWillebrandFactorTheformationofa1.8MDavonWillebrandfactor(vWF)multimerwithboundfactorVIIIisillustrated.EachvWFmonomer(Mr=260,000)containsafactorVIIIbindingsiteneartheNH2-terminalendofthemolecule.Themonomersarejoinedend-to-end(NH2toNH2andCOOHtoCOOH)bydisulfidebondstoformlargemultimers.ThematuremultimerscanbindonefactorVIIImoleculepermonomericsubunit.SampleGelInformation:GelNovex4-12%Bis-TrisLoadHumanvonWillebrandFactor,1µgperlaneBufferMOPSStandardSeeBluePlus2;Myosin(191kDa),PhosphorylaseB(97kDa),BSA(64kDa),GlutamicDehydrogenase(51kDa),AlcoholDehydrogenase(39kDa),CarbonicAnhydrase(28kDa),MyoglobinRed(19kDa),Lysozyme(14kDa)Overview:VonWillebrandfactor(vWF)isamultimericplasmaglycoproteinthatisrequiredfornormalhemostaticplateletplugformation(1-8).Thematureplasmaproteiniscomposedofapparentlyidenticalsubunits(Mr=260,000)whichareheldtogetherbydisulfidebonds.ThecirculatingvWFmoleculerangesinsizefromdimers(Mr=520,000)toextremelylargemultimers(Mr>10,000,000).Duringnormalhemostasis,thelargermultimersofvWFareresponsIBLeforfacilitatingplateletplugformationbyformingabridgebetweenplateletglycoproteinIBandexposedcollageninthesubendothelium(9-14).EitheralackofvWFproteinorthepresenceofabnormalitieswhichresultindecreasedpolymerizationmaycausealossofBIOLOGicalactivitywhichischaracteristicofvonWillebrand"sdisease.Inadditiontoitsroleinplateletplugformation,vWFisalsoresponsibleforthebindingandtransportoffactorVIII(antihemophilicfactor)inplasma(15).ItappearsthatthislattereventisresponsibleforboththestABIlityandeffectivedeliveryoffunctionalfactorVIII.StudiesindicatethatfactorVIIIbindstotheNH2-terminalportionofthematurevWFsubunitwithastoichiometryofonefactorVIIImoleculepervWFmonomer(16,17).ThesinglechainvWFmonomercontainsalargenumberofcysteineresiduesatboththeNH2-terminalandCOOH-terminalends,whichareinvolvedinthemultimerformation.Carbohydrateanalysesindicatethatnearly15%ofthemassofvWFiscontributedbycarbohydrate(18).ItappearsthatthecarbohydrateservestoprotectvWFfromproteolysis,butisnotnecessaryforfunctionalactivityormultimerformation.vWFispreparedfromcitratedhumanplasmausingacombinationoftheproceduresdescribedbyThorell(19),andLollar(20).AfactorVIII"free"vWFpreparation,furtherpurifiedtoensureremovaloffactorVIIIprocoagulantactivity(antigenstillpresent),isalsoavailable.Thepreparationsare>95%pureasjudgedbySDS-PAGEunderreducingconditions,andconsistoflargemultimersasdeterminedbyelectrophoresisinSDS/agarosegels.Theproteinisshippedfrozenin0.025Msodiumcitrate,0.1Mglycine,0.1MNaCl,pH6.8,forstorageat-70°C.Properties:Localization/th>PlasmaandsubendotheliumModeofactionfacilitatesplateletplugformationbyformingabridgebetweenplateletglycoproteinIBandexposedcollageninthesubendothelium;alsobindsandtransportsfactorVIIIMolecularweight260,000to>10,000,000(1-8)Isoelectricpoint5.7-5.9(21)ExtinctioncoefficientNotApplicable;concentrationdeterminedbytotalproteinassay.Concentrationinplasma10micrograms/mL(21)Structuremultimericproteincomposedofidentical260,000molecularweightsubunitsPercentcarbohydrateapproximately15%(18)