The direct binding of human factor VII in plasma to recombinant human tissue factor

The extrinsic pathway of coagulation is initiated when zymogen factor VII binds to its cell surface receptor tissue factor. Recently recombinant human tissue factor has become available and therefore in this study the direct binding of human factor VII in plasma to recombinant tissue factor was explored. Factor VII binding was quantitated by a standard ELISA protocol using monospecific polyclonal rabbit anti-human factor VII as the primary antibody and goat anti-rabbit IgG conjugated to alkaline phosphatase as the second antibody. Both the oxidation state of the recombinant tissue factor and calcium ion concentration were found to be critical for the efficient binding of factor VII. A linear relationship was observed between absorbance and factor VII concentration when normal pooled human plasma was diluted in the range 1:25 to 1:1000 (factor VII concentration 0.5-20 ng/ml). Evidence is provided to show that binding is both specific for human coagulation factor VII and can be utilized to detect factor VII molecular variants with impaired tissue factor binding.