Real-time imaging analysis of the effect of the thrombomodulin (TM)/thrombin-activatable fibrinolysis inhibitor (TAFI) system on the accumulation of labeled plasminogen. (A) Representative sequential overlaid images of Alexa Fluor 488-labeled fibrinogen (fbg-488; green) and Alexa Fluor 568-labeled plasminogen (plg-568; red). The dense fibrin network formed at the activated platelets indicates the heterogeneity of the fibrin network structure. Tissue-type plasminogen activator, at 1.5 nM, was added to initiate plg-568 accumulation at the activated platelets and on the fibrin fibers. (Upper) Control (2, 15, 30, 45, and 60 minutes from the start of video capture). (Middle) TAFIa inhibitor (TAFIaI; 2, 10, 15, 20, and 25 minutes from the start of video capture). (Lower) Neutralizing antibody of TM (TM-Ab; 2, 10, 15, 20, and 25 minutes from the start of video capture). (B) Average fluorescence intensities of fbg-488 (blue circles) and plg-568 (red circles) in the dense fibrin region, in 30-pixel (∼34-μm) circular regions of interest, are plotted over time for the control. Plasminogen accumulation (Plg. accum.) time (double-headed arrow) was measured as the period from the appearance of the fibrin fiber structure to the achievement of maximal intensity for plg-568 fluorescence. (C) Plg. accum. time in control (n = 31, median 45 [minutes], interquartile range [IQR] 30.5-48), TAFIaI (n = 22, median 16 [minutes], IQR 14.3-16), and TM-Ab (n = 22, median 17 [minutes], IQR 12.3-17). Statistical analysis was performed using the Mann-Whitney U test (^#P < .001 vs control).