Detection of the α₂β₁ conformation on platelets using flow cytometry. (A) Left panel: interaction of IAC-1-FITC (70 μg/mL) with resting PRP (thin line) or PRP stimulated with 50 μM ADP (dotted line) or 5 μM U46619 (bold line). Right panel: binding of IAC-1-FITC to resting washed platelets (thin line) or after stimulation with 1 U/mL thrombin (bold line). Tracings are representatives of at least 3 experiments. (B) Binding of IAC-1-FITC to platelets stimulated with indicated concentrations of convulxin in the absence (▦) or the presence (□) of 5 U/mL apyrase, expressed as the percentage of binding induced by 200 ng/mL convulxin. Error bars indicate the SEMs of 3 independent experiments. (C) Binding of either FITC-labeled Gi9 (20 μg/mL), IAC-1 (70 μg/mL), or soluble collagen (100 μg/mL) was measured to PRP with 50 ng/mL convulxin (left) or 5 μM U46619 (right). Data are represented as the ratio of MF for stimulated versus unstimulated platelets, and are means ± SEMs of at least 3 experiments. (D) Representative tracings of the binding of 100 μg/mL soluble collagen-FITC to unstimulated platelets in PRP (dotted line) and platelets stimulated (thin line) with 50 ng/mL CVX (left) or 5 μM U46619 (right); the enhanced collagen binding was reversed by 20 μg/mL blocking mAb Gi9 (bold line).