Comparable kinetics of glycoprotein receptor shedding and PS exposure in ionomycin-stimulated platelets. Washed platelets, which were preincubated with vehicle or GW280264X (GW; 5 μM), were left unstimulated (Ctrl) or were stimulated for the indicated times (15 to 300 minutes) with ionomycin (37°C) and evaluated for glycoprotein expression by flow cytometry. (A-B) ADAM-dependent decrease in surface expression of GPIbα and GPVI over time. Mean fluorescence intensities were normalized to those of unstimulated platelets. (C-D) Analysis of platelet populations after dual staining for GPIbα (FITC mAb) or GPVI (PE mAb) and for PS exposure (AF647-annexin A5). Flow cytometric events were separated into 4 platelet populations: GPIbα/VI^high PS⁻, GPIbα/VI^high PS⁺, GPIbα/VI^low PS⁻, and GPIbα/VI^low PS⁺. Shown are representative dot plots and quantification of 4 quadrants. Data are mean ± SD, n = 3-5 (≥3 donors). *P < .05 vs control, ^#P < .05 vs vehicle.