Cacl2 solution

Coagulation times were determined in human plasma using a STAGO STart 4 coagulation analyzer (Diagnostica). Human single donor plasma was used (Innovative Research). For the extrinsic coagulation, 50 μl of plasma with or without inhibitor was placed in the incubating chamber of the instrument for 2 min at 37°C. After incubation, 100 μl of innovin (recombinant human tissue factor, synthetic phospholipids, and calcium in stabilized Hepes buffer system; Dade Behring/Siemens) was added using the pipet connected to the instrument. Upon addition of this reagent, the electromagnetically induced movement of a steel ball in the plasma was monitored. The time until the ball stops moving was recorded as coagulation time. For the intrinsic coagulation, 100 μl of plasma with or without inhibitor was incubated with 100 μl of Pathromtin SL (silicon dioxide particles and plant phospholipids in Hepes buffer system, Siemens) for 2 min at 37°C. Subsequently, the coagulation was triggered by addition of 100 μl of CaCl₂ solution (25 mM, Siemens).