Effect of anti-GPVI Nbs in whole blood microfluidics. (A) Representative images of whole blood perfused at an arterial shear rate (1000/s) over Horm collagen I in the presence of either PBS or 500 nM Nb 2, 21, or 35. Adhered platelets and platelet aggregates were imaged in brightfield after 3.5 minutes of flow. Platelets were labeled with Annexin V AF568 to assess PS exposure. All images were taken on an EVOS AMF4300 using a 60×, 1.42 NA oil objective. Data are representative of 2 runs for each of 3 donors per treatment. Scale bars, 50 µm. (B-D) Quantitative analysis of the images assessed the effect of the Nbs on percentage of total surface area covered by platelets (B), multilayered thrombi (C), and platelets exposing PS (D). Data points are individual runs for each of 3 donors (shown in different colors) per treatment (mean ± SD). Unpaired Student t test with Mann-Whitney correction was used to test for statistical significance: **P < .005. SAC, surface area coverage.