Binding of FXII(a) to fibrin(ogen) analyzed by surface plasmon resonance. (A-B) The binding to fibrinogen (from top to bottom, 25, 12.5, 6.25, and 0nM FXII (A) and α-FXIIa (B), respectively). (C) The (negligible) binding of β-FXIIa to fibrinogen (from top to bottom, 312.5, 156.3, 78.1, and 0nM, respectively). (Bottom panels) Binding of FXII (D), α-FXIIa (E), and β-FXIIa (F) to fibrin at identical concentrations as in panels A to C, respectively. For reasons of clarity, not all tested concentrations are shown in this figure. Supplemental Figures 3 and 4 show the responses for all tested concentrations, including the ka and kd fitting, which were used to determine the Kd values as presented in Table 2. The graphs represent the mean of 3 experiments. (G-H) Western blots after immunoprecipitation of FXII from normal pooled plasma. (G) The blot was stained for FXII with polyclonal antihuman FXII. (H) The blot was stained for fibrinogen with polyclonal antihuman fibrinogen. (G) Lane 1 (0.05 μg nonreduced FXIIa) and lane 2 (0.05 μg reduced FXIIa) are controls, lane 3 is reduced immunoprecipitate, lane 4 is nonreduced immunoprecipitate, and lane 5 is a molecular weight marker. (H) Lane 1 is a molecular weight marker, lane 2 (0.25 μg reduced fibrinogen) is control, and lane 3 is reduced immunoprecipitate. The blots show the presence of zymogen FXII (80 kDa; G) and the Aα-chain (66 kDa), Bβ-chain (52 kDa), and γ-chain (46 kDa) of fibrinogen (H).