Dade innovin

The following commercial reagents were used: thrombin (Enzyme Research Laboratories, South Bend, IN, USA), hirudin (Calbiochem, San Diego, CA, USA), corn trypsin inhibitor (CTI) (Enzyme Research Laboratories, South Bend, IN, USA) thrombin calibrator (Synapse BV, Maastricht The Netherlands), Substrate Z-Gly-Gly-Arg-AMC (Bachem, Bubendorf, Switzerland), recombinant human tissue factor (TF), Dade Innovin®, (Dade Behring, Newark, DE, USA), mouse anti-human FVIII (GMA8015) and isotype antibody (GMA650) (Green Mountain Antibodies, Burlington, VT, USA), rhFVIIa, NovoSeven® (Novo Nordisk, Bagsvaerd, Denmark), recombinant tissue plasminogen activator, Activase® (Genentech, San Francisco, CA, USA), FVIII, Kogenate® (Bayer Healthcare, Leverkusen, Germany), and carboxypeptidase inhibitor from potato tuber (CPI) (Sigma-Aldrich, USA). Pooled normal Human Plasma (NHP) and plasma from FVIII-deficient patients (FVIIIdP) with and without inhibitors (titer specified in Bethesda Units (BU)), were purchased from George King Bio-Medical (Overland Park, KS, USA). Phosphatidylcholine (PC), phosphatidylserine (PS) and phosphatidylethanolamine (PE) were purchased from Avanti Lipids (Alabaster, AL,USA) and phospholipid vesicles containing 80% PC and 20% PS or 60% PC, 20% PS, and 20% PE were prepared as described (22 (link)).

Dade^® Innovin^® was purchased from Siemens. All other materials were purchased from Sigma-Aldrich or previously cited sources.^{65 (link)} Hepes-Tyrode buffer (129 mM NaCl, 20 mM HEPES, 12 mM NaHCO₃, 2.9 mM KCl, 1 mM MgCl₂, 0.34 mM Na₂HPO₄·12H₂O; pH7.3) was modified with 0.045 g of glucose per 50 mL of buffer the day of experiment and kept in the 30°C bath until use.