αFXIIa plasminogen activator activity is enhanced by polyP₇₀. (A) Clots were formed with 3.8 μM fibrinogen, 0.24 μM Glu-plasminogen, and 200 nM αFXIIa in the absence (gray) and presence (black) of 70 μM polyP₇₀. Clotting was initiated with 0.25 U/mL of thrombin and 10 mM CaCl₂, and subsequent lysis was monitored at 405 nm. Mean data ± SEM is expressed as percentage turbidity (n = 5; P < .0001). (B) Plasminogen activation was analyzed by incubating 200 nM αFXIIa and 200 nM plasminogen in the absence (light gray) or presence (black) of 70 μM polyP₇₀. Plasmin activity was detected using the chromogenic substrate S2251 at 405 nm. Data represent mean ± SEM (n = 3; P < .0001). (C) αFXIIa-mediated plasminogen activation was analyzed in the presence of various concentrations of polyP₇₀ by incubating 200 nM αFXIIa and 200 nM plasminogen in the absence (dashed line) or presence of 70, 35, 17.5, or 4.4 μM polyP₇₀, as indicated. Plasmin activity was detected using the chromogenic substrate S2251 at 405 nm. Data represent mean ± SEM (n = 3; P < .0001). (D) Similarly, direct effects of polyP on preformed plasmin (6.25 nM) were analyzed in the absence (gray line) and presence (black line) of 70 μM polyP₇₀ with S2251. Data represent mean ± SEM (n = 3; P = .93). (E) Activation of plasminogen (200 nM) by βFXIIa (200 nM) was monitored in the absence (gray) and presence (black) of 70 μM polyP₇₀ and was detected using S2251. Data represent mean ± SEM (n = 4; P = .71). (F) Plasminogen activation was analyzed by incubating 200 nM αFXIIa and 200 nM plasminogen in the absence (light gray) or presence (dark gray) of 70 μM platelet-derived polyP. Plasmin activity was detected using the chromogenic substrate S2251 at 405 nm. Data represent mean ± SEM (n = 3; P < .0001). A, absorbance; SEM, standard error of the mean.