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The data reported herein were obtained in a search for mechanisms triggering intrinsic clotting which bypass the contact activation factors. An assay system was designed which permitted us to examine the effect of thromboplastin and Factor VII (F.V11) upon F.IX under conditions which produced minimal direct effect of thromboplastin and F.VII on F.X. Purified F.IX was incubated with purified F.VII, brain thromboplastin and Ca ions.Then, a mixture of purified bovine F.VIII (activated with thrombin), phospholipid, Ca ions and F.X was added. The reaction was stopped at 3 min with EDTA, and a subsample assayed for F. Xa with the chromogenic substrate S-2222. When 10 μl of F.IX (15.4 U/ml) was incubated with 10 μl of F.VII (0.2 U/ml), 5 jul brain thromboplastin, and 3 Jul 50 mM CaCl2 for 10 min at 370, F.IXa was formed as shown by generation of 0.830 U/ml of F.Xa. When 10 μl of F.XIa (0.24 U/ml) was substituted for the F.VII and thromboplastin, 1.040 U/ml of F.Xa was generated. Both the thromboplastin-F.VII generation of F.IXa and the F.XIa generation of F.IXa was time-dependent. Only minimal F.Xa was generated in this assay system when the first incubation mixture contained thromboplastin and F.VII without F.IX (0.088 U/ml), thromboplastin and F.IX without F.VII (0.002 U/ml), or F.VII and F.IX without thromboplastin (0.000 U/ml). Only minimal F.Xa was generated when F.VIII was left out of the second incubation mixture. These data establish that thromboplastin and F.VII can activate F.IX in a reaction bypassing F.XIa.

A Bypass Mechanism for Activating Human Factor IX with Human Factor VII and Thromboplastin