Platelet adhesion to fibrillar collagen under physiological flow conditions. (A) Whole blood from WT (WT, black lines and bars), CalDAG-GEFI^−/− (knockout [KO], red), clopidogrel-treated WT (WT + clop., blue), or clopidogrel-treated KO (KO + clop., green) mice was perfused over collagen at arterial (2000 s⁻¹, left) or venous (400 s⁻¹, right) shear conditions. Platelets in whole blood were labeled with Alexa488-labeled antibodies to GPIX before perfusion. The top graphs represent time traces of the mean fluorescence intensity ± SEM expressed as a percentage of the maximal fluorescence observed (WT blood, 400 s⁻¹). The bar graphs show the area coverage by fluorescent platelets after 5 minutes of blood perfusion, expressed as percentage of the collagen-coated area. Data are shown as mean ± SEM (n = 4-6, 3 independent experiments). *P < .05, **P < .01, ***P < .001. See supplemental Videos 1 to 4 for a better visualization of the differences in thrombus growth and stability observed in the respective study groups. (B-C) Effect of exogenous ADP and TxA₂ (U46619) on the adhesion of CalDAG-GEFI^−/− platelets. WT and CalDAG-GEFI^−/− (KO) whole blood was perfused over collagen at 400 s⁻¹ or 2000 s⁻¹ in the presence (KO + ADP/U46) or absence (KO) of exogenous ADP (25μM) and U46619 (5μM). (B) Bar graphs for area coverage (top) and fluorescence intensity (bottom) measured after 5 minutes of perfusion with the following blood samples: WT (black bar), KO (red bar), and KO reconstituted with 25μM ADP and 5μM U46619 (KO + ADP/U46, red checkered bar). Data are shown as mean ± SEM (n = 5, 3 independent experiments). *P < .05; **P < .01; ***P < .001. (C) Representative images. Images were obtained after 5 minutes of perfusion on a Nikon Eclipse Ti-U inverted microscope (equipped with a Retiga EXL monochrome camera [QImaging] and Nikon NIS Elements software [NIS-Elements Advanced Research]).