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Updated: Jul 3, 2026

Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay
Published on: September 9, 2012
Protamine inhibits formation of the covalent factor IXa-anti-thrombin complex
Arthur S Brecher1, Amie R Roland
1Department of Chemistry, Bowling Green State University, Bowling Green, OH 43403, USA. artbrec@bgsu.edu
Abstract:
The effect of protamine sulfate upon the covalent interaction of human factor IXa with anti-thrombin has been investigated by a quantitative SDS-polyacrylamide gel electrophoresis procedure. Protamine sulfate inhibits the interaction of factor IXa with anti-thrombin upon preincubation with factor IXa, anti-thrombin, or a mixture of the two components. As a consequence of the presence of protamine sulfate, autolysis of factor IXa is promoted to generate a degradation product of a molecular weight (apparent) of 35 kDa. Concomitantly, free factor IXa and free anti-thrombin levels are increased in the mixture. In contrast to the effect of protamine sulfate, heparin promotes a massive increase in the factor IXa-anti-thrombin complex appearing as a doublet. Associated with the increase in the complex is a precipitous drop in free factor IXa and anti-thrombin, as well as the steep decline in the 35 kDa factor IX fragment, the presence of which reflects the degradation of factor IXa. Accordingly, protamine sulfate appears to exert a procoagulant effect upon addition to an intrinsic coagulation system enzyme, in contrast to its modest anticoagulant effect on the overall coagulation process.
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