CCR4 on resting human platelets and P selectin after stimulation.
(A) Presence of CCR4 on resting human platelets detected by flow cytometry and (B) expression of P selectin after stimulation with SDF-1 (0.5 μg/mL) and MDC (0.5 μg/mL) and low doses of ADP (0.1 μM), compared with thrombin (1 U/mL). (A) Washed, resting platelets fixed with paraformaldehyde were stained with anti-CCR4 mAb or the same concentration of an isotype control IgG followed by the addition of FITC-labeled goat–anti-mouse IgG. The staining with control antibody is shown as the solid profile, and the staining with the anti-CCR4 antibody is shown as the open profile. Percentage positive expression is shown above. (B) Platelets were stimulated in the absence of fibrinogen with SDF-1 and ADP for 10 seconds, MDC and ADP for 10 seconds, thrombin for 10 seconds (middle distribution), and for 10 minutes (right distribution) at 37°C and immediately fixed. Fixed platelets were incubated with anti-P selectin mAb S12 or control nonspecific mouse IgG1 for 1 hour at room temperature, followed by the addition of FITC-labeled goat-antimouse IgG. Treated platelets are shown in the open profiles.
