FVIIIa-dependent thrombus formation induced by the TF-FVIIa initiation complex in flowing blood. (A) Blood reconstituted with washed type 0 blood cells added to the original count into FVII-deficient citrated PPP with 200 pM WT or mutant FVIIa and without or with the inhibitory anti-FVIII MoAb C5 (25 μg/mL) was recalcified to 1.29 mM Ca²⁺ and perfused for 3.5 min at 300 s⁻¹ wall shear rate. Where indicated, FIXa (20 pM) was added to blood. Representative confocal images are shown with superimposed green (platelet aggregates and leukocytes) and red (fibrin) fluorescence channels. Image side = 312 μm. (B) Quantification of the volume of platelet aggregates and deposited fibrin after adding FVIIa WT, T99Y, or S195A without or with anti-FVIII MoAb (n = 4-6 for the different conditions). (C) As in panel B, but after adding FVIIa WT or E154A (n = 3-8 for the different conditions). Results in panels B and C (shown as 25th-75th percentile bars, min-to-max whiskers, line at the median; or min-to-max floating bars, line at the mean when n ≤ 3) were evaluated by the ANOVA/Tukey tests. *P < .05. **P < .01. ***P < .001.