ABT-263 depletes intracellular calcium stores. (A) Washed platelets were incubated in HBS with ABT-263 (1μM) and caspase inhibitors (z-VAD.fmk, 100μM, Q-VD-POh, 50μM) for 2 hours before exposure to the platelet agonists CRP-XL (2 μg/mL), A23187 (10μM), or TRAP (100μM) for 10 minutes. Platelet activation was assessed by exposure of P-selectin and flow cytometry. (B) Whole blood was incubated with ABT-737 (10μM) and caspase inhibitors (z-VAD.fmk, 100μM, Q-VD-POh, 50μM) for 2 hours. PRP count was adjusted with autologous filtered PPP to 1.5 × 10⁸/mL, and aggregation was measured using an aggregometer. Data are mean ± SD (n = 3). (C) Washed human platelets were stained with Fluo4/AM in HBS for 30 minutes. Intracellular calcium levels were continuously monitored by flow cytometry. After establishing a baseline fluorescence signal for 1 minute, ABT-263 (10μM) was added and the calcium response was monitored for an additional 4 minutes. (Left panel) Representative graph. (Right panel) Mean ± SD of the geometric mean fluorescence (MFI) (n = 3). (D) Washed human platelets were incubated with or without ABT-263 (1μM) and z-VAD.fmk (100μM) in HBS for 2 hours before staining with Fluo4/AM for an additional 30 minutes. After establishing a baseline fluorescence signal for 1 minute, A23187 (5μM) was added and the calcium response was monitored for an additional 4 minutes. Data are mean ± SD of the geometric mean fluorescence (n = 3).