The effect of varying protein S levels on thrombin generation in plasma in the absence of APC. (A) Normal pooled plasma was incubated with antibodies against protein C and 2.73 μM (▵), 1.37 μM (▿), 0.68 μM (□), 0.34 μM (⋄), 0.17 μM (○), or no (•) antibodies directed against protein S. (A, inset) ETP as function of the concentration antibodies against protein S. (B) Protein S–depleted plasma was mixed with varying amounts of normal pooled plasma resulting in protein S levels of 0% (▵), 40% (▿), 55% (□), 70% (⋄), 85% (○), and 100% (•). (B, inset) ETP as function of the protein S concentration. All plasma mixtures contained antibodies against protein C. Thrombin generation was initiated with 3.5 pM tissue factor, 10 μM phospholipid vesicles (20/60/20 DOPS/DOPC/DOPE), and 16 mM CaCl₂ (final concentrations) and quantified with the fluorogenic substrate I-1140 as described in “Materials and methods.”