Detection of platelet activation.
One hundred microliters platelets at 10 × 106 cells/mL in citrated PPP were sheared at 9600/s in a cone-plate viscometer with a 0.5° cone for 300 seconds. Function-blocking antibodies against GpIb (AN-51) and GpIIb-IIIa (BLE-6 or 7E3) were added in some runs. CD61-PerCP fluorescence was used to gate on platelets and microparticles. (A) Flow cytometric detection of activated platelets using Annexin V–FITC at t = 0 and 300 seconds. (B) SIPAct as a function of shearing time in the absence or presence of antibodies. Significant binding was observed at the earliest sampling time point, 10 seconds. (C) Percentage increase in microparticle number at t = 300 seconds after shear compared with the number at t = 0. Data are mean ± SEM for n ≥ 3. *P < .05 with respect to t = 0 sample. §P < .05 with respect to anti–GpIIb-IIIa treatment.
