Inhibition of FXa by the TFPI forms in the absence of PS. Reactions were performed in TBS/BSA. Progress curves for the hydrolysis of S-2222 by FXa as measured by ΔA₄₀₅ nm are presented at different concentrations of each TFPI. Reaction mixtures in 160 μL contained S-2222 (500μM), CaCl₂ (5mM), cephalin (35μM), FXa (200pM), and TFPI (0-16nM). For all panels, the concentrations of TFPI from top to bottom are 0, 0.5, 1, 2, 4, 8, and 16nM, except for TFPI_C2 where the highest concentration is 12nM and not 16nM. The data were fitted by nonlinear regression using the program Grafit (Version 3.01) from Erithacus software. Each experiment was repeated at least once. One set of experiments is shown.