Figure 3.

CLEC‐2‐dependent tyrosine phosphorylation is partially reduced by cyclic nucleotide‐elevation. Washed platelets (4 × 10⁸ per mL) incubated with PGI₂ (1 μm) or GSNO (100 μm) in the presence of apyrase (2 U mL⁻¹), indomethacin (10 μm) and lotrafiban (10 μm) were stimulated with 10 μg mL⁻¹ CLEC‐2 mAb for 3 min and lysed. Aliquots were analyzed by SDS‐PAGE (A) and immunoprecipitation of PLCγ2, Syk and CLEC‐2 (B) (n = 3–4). Blots were probed with anti phospho‐tyrosine mAb (A & B) and reprobed for equal loading. Densitometry of PLCγ2, Syk and CLEC‐2 bands was expressed as fold increase over basal and plotted as mean ± SEM. Statistical differences were evaluated by one‐way anova test and Bonferroni post‐test (*P < 0.05, **P < 0.01) (C).