Purification of porcine and human ristocetin-Willebrand factor.
A plasmatic component required for the ristocetin-induced aggregation of platelets has been purified from normal human and porcine plasma by gel filtration (4% agarose) and anion-exchange chromatography (DEAE cellulose). No factor VIII coagulant activity was found associated with the purified human or porcine component. Urea sodium dodecylsulfate electrophoretic analysis of the purified component of both species indicated that the apparent molecular weight with intact disulfides is in excess of 500,000; after disulfide reduction with 2-mercaptoethanol, single components with an apparent subunit molecular weight of 230,000 were observed. Purified porcine ristocetin-Willebrand factor (RWF) co-sedimented in sucrose gradients with the factor present in normal plasma. Amino acid analysis of both human and porcine RWF indicated that all normal amino acids are present, whereas amino sugars were undetected. However, lipid analysis indicated 1% to 2% lipids present, including monoglycerides, di- and tri-glycerides, cholesterol, cholesterol esters, some free fatty acids, and a trace of phospholipid. A single line of identity was observed between normal human plasma and purified human RWF when immunodiffusion plates were run with purified rabbit anti-human RWF immunoglobulins. Antisera raised against human and porcine RWF's do not inhibit the factor VIII coagulant activity of the homologous plasma, nor is "spontaneously occurring" human factor VIII inhibitor neutralized by the isolated material of either species.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001779:Blood Coagulation Factors / Q000302:isolation & purification*; D001780:Blood Coagulation Tests; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D005779:Immunodiffusion; D008055:Lipids / Q000032:analysis; D008722:Methods; D010974:Platelet Aggregation / Q000187:drug effects; D011506:Proteins / Q000032:analysis; D012310:Ristocetin / Q000302:isolation & purification* / Q000494:pharmacology; D013552:Swine; D014842:von Willebrand Diseases / Q000097:blood; D014841:von Willebrand Factor / Q000302:isolation & purification*
D000596:Amino Acids; D001779:Blood Coagulation Factors; D008055:Lipids; D011506:Proteins; D014841:von Willebrand Factor; D012310:Ristocetin