IMMUNOAFFINITY PURIFICATION OF BOVINE FACTOR-VII

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  • 1 January 1984
    • journal article
    • research article
    • Vol. 63  (2) , 393-398
Abstract
Factor VII was purified to homogeneity from bovine plasma by a procedure that includes affinity purification on an immunoadsorbent column. Recovery was determined by both coagulant assay and liquid scintillation counting, using 3H-factor VII as an internal standard. The purification factor calculated by both methods was .apprx. 120,000-fold, with a final yield of .apprx. 18%. Homogeneity was assessed by sodium dodecyl sulfate (SDS) polyacrylamide gel electrophoresis. The material migrated as a single polypeptide chain of 53,000 daltons, and following activation by factor Xa, the 1-chain zymogen was quantitatively converted to 2-chain factor VIIa. Conversion of affinity-purified factor VII to factor VIIa resulted in up to a 119-fold activation of the coagulant activity, which is 2.7-4 times greater than the activatability reported for factor VII prepared by other methods. Pure factor VII, uncontaminated by traces of factor VIIa, apparently would be activated 123-fold upon conversion to factor VIIa. The close agreement between observed activatability of affinity-purified factor VII and the theoretical prediction suggests that factor VII was isolated essentially free of factor VIIa. The purification data from 3 lots of bovine plasma yield an estimate for the plasma concentration of factor VII from 10.1 nM-18.5 nM.