Endogenous FXIa generation cleaves the K1 domain of TFPI and increases fibrin generation on HUVEC. (A) FXIIa and kallikrein generation in citrated plasma pretreated with vehicle or aprotinin (50 µM) and CTI (50 µg/mL) in the presence or absence of HUVECs was determined using S-2302 hydrolysis. (B) HUVECs were pretreated with citrated plasma for 30 minutes; reactions were stopped with aprotinin (50 µM) and CTI (50 µg/mL). Cell surface detection of TFPI was measured by an anti-TFPI K1 antibody or a polyclonal anti-TFPI antibody. P < .05 with respect to vehicle in the presence of the anti-K1 antibody. (C) HUVECs were pretreated with citrated plasma for 30 minutes in the presence or absence of anti-TFPI K1 and K2 antibodies (10 µg/mL), 14E11 (20 µg/mL), 1A6 (20 µg/mL), 10C9 (50 µg/mL), or aprotinin (50 µM); reactions were stopped with aprotinin (50 µM) and CTI (50 µg/mL). Subsequently, recalcified plasma pretreated with 20 µg/mL 1A6 was added to HUVECs, and the time for fibrin generation to reach half-maximal (T_half-max) was measured. Data are mean ± SE (n = 3). P < .05 with respect to vehicle in the presence of plasma. Mann-Whitney U test was used for statistical comparisons.