Kinetics of GPIbα-A1 interaction. (A) Disassociation rate constants were estimated at wall shear stresses between 400 and 2000 s⁻¹ using a 2 μg/mL coating concentration of recombinant protein based on the duration of transient tether events. Kinetics of transient tethering between platelets and DC-D′A3 variant at a shear rates of 400, 600, and 1000 s⁻¹ is demonstrated as an example. The natural logarithm of the number of tethering events (∼ 150 individual interactions) was plotted against the duration of tethering and the negative slope of the lines is the k_off. Disassociation rate values at each shear rate are indicated (R² values were ∼ 0.9). (B) Rate of platelet tethering to either wtD-A3 or OLG variants was plotted a function of shear rate. The graph shows the number of platelets that transiently tethered to each substrate at a given shear rate, during 5 seconds at 3 different fields of view (0.4 × 0.3 mm). Values are shown as mean ± SEM of 3 independent experiments.