IKK activity is important for platelet secretion. (A) Mouse platelets were prepared as described in our “Methods” section. After adding 0.7 mM CaCl₂, platelets were incubated with (open symbols) or without (●) 5 μM BMS-345541 for 3 minutes, then stimulated with thrombin (0.05 U/mL) for the indicated times. (B) IKKβ^−/− platelets (○) and platelets from Cre⁻ littermate controls (●) were stimulated with thrombin (0.05 U/mL) under similar conditions. Release of [³H]5-HT from dense granules (Dense), PF4 from α-granules (Alpha), and β-hexosaminidase from lysosomes (Lysosome) was measured and percent secretion was calculated. Each data point represents the mean of triplicates and the standard deviation is indicated. (C) IKK-β^+/+ and IKK-β^−/− platelets were subjected to immunoblotting with anti-IKK-β antibody. (D) IKK-β^+/+ and IKK-β^−/− platelets were stimulated with 0.1 U/mL thrombin for 3 minutes and subjected to immunoblotting with anti-phospho-Ser95 (pSer-95) and anti-SNAP-23 antibodies.