Bovine Platelet Proteins II. Purification of Platelet Fibrinogen

Abstract

Summary1. Platelet fibrinogen has been purified from washed bovine platelets. The procedure was based on the methods for purification of plasma fibrinogen by fractionated precipitations and extractions with ethanol and glycine below 0°, and precipitation of proteins by dimethylformamide at 0°.2. The platelet extract obtained by freezing and thawing of the cells, freed from insoluble material by centrifugation at 23,000 x g for 30 min, contained 0.22 ±0.003mg fibrinogen per 109 platelets. Total protein of this fraction was 0.77 ±0.08 mg per 109 platelets whereas that of the insoluble fraction was 0.79 ±0.09 mg per 109 platelets.3. The most purified platelet fibrinogen fraction contained 91-98% of the protein in a thrombin-clottable state. The yield was approx. 20%. It showed homogeneity in analytical ultracentrifugation, in immunoelectrophoresis using an antiserum produced by immunization of rabbits against platelet extract, and in starch gel electrophoresis using a discontinuous system of Tris HCl and borate buffers offering a high resolution power towards the platelet proteins. Polyacrylamide disc electrophoresis revealed two to three faint lines behind the main fibrinogen line. At least one such line was also observed with purified plasma fibrinogen.

Bovine Platelet Proteins III. Some Properties of Platelet Fibrinogen

“…Plasma fibrinogen is considered to b e synthesized mainly in the liver ( 4 7). The finding of others (26) t hat there was no exchange between the intraplatelet and t h e plasma fibrinogen pools lends no support to t he view t hat the platelet fibrinogen is m erely a " pinocytized" plasm a fibrinogen nor does the high intracellular concentration fit such a hypothesis (11). Human platelet s contain some RNA but have no nucleus and no det ectable DNA (48).…”

“…Electrophoretic mobilities were the same for platelet and plasma fibrinogen in starch gel electrophoresis and polyacrylamide disc electrophoresis under all conditions used (see ref. 11,Fig.4).…”

Adenine Nucleotide Metabolism during the Aggregation of Human Platelets by Bovine Factor VIII

“…Human platelets aggregate when treated with preparations of bovine plasma and platelet fibrinogen [2,7,15,16]. Recent information [4,6] strongly suggests that factor VIII present in the fibrinogen preparations is solely responsible for the aggregation effect.…”