EVALUATION OF THE INITIATION PHASE OF BLOOD-COAGULATION USING ULTRASENSITIVE ASSAYS FOR SERINE PROTEASES

The initiation phase of enzyme generation in a reconstituted model of the tissue factor (TF) pathway to thrombin was evaluated, At 1.25 pM a dded TF, no thrombin generation was observed in the absence of factor V, The substitution of factor Va for factor V increased the rate of th rombin generation, Factor X activation during the initiation phase was not influenced by the absence of factor VIII or thrombin, leading to the conclusion that initially factor Xa is generated exclusively by th e factor VIIa-TF complex, When thrombin was eliminated from the system , no contribution of the factor Ma-factor VIIIa complex to factor X ac tivation was observed during the propagation phase, Similarly, factor V activation was also not observed in the absence of thrombin, indicat ing that thrombin is the only enzyme responsible for factor V and fact or VIII activation, Only subnanomolar amounts of factor VII were activ ated when prothrombin activation was almost complete, In the absence o f coagulation inhibitors, factor XI did not influence thrombin generat ion initiated by 1.25 pat factor VIIa-TF complex. The termination of f actor XIa generation by added hirudin in the factor XI experiment indi cates that factor XI activation occurs exclusively by thrombin.