Bitiscetin-induced binding of ¹²⁵I-vWF to platelets and its inhibition by MoAb directed against the A1 domain of vWF or against the GPIb-binding site of vWF. (A) The binding of¹²⁵I-vWF (0.1 to 25 μg/mL) to fixed-platelets (10⁸/mL) in the presence of 2 μg/mL of bitiscetin was quantified as described in Materials and Methods, using a labeled preparation of vWF with a specific radioactivity of 1.4 × 10⁶ cpm/μg. Nonspecific binding estimated in the absence of bitiscetin (⧫) was substracted from total binding in the presence of stimulus (•) to derive the specific binding (○). Binding isotherm and binding parameters were estimated by fitting the data of the specific binding with a nonlinear least-square, regression analysis. (B) Inhibition of the bitiscetin (2 μg/mL)-induced binding of ¹²⁵I-vWF (1 μg/mL) to fixed platelets (10⁸/mL) by MoAbs (0.01 to 20 μg/mL). Nonspecific binding estimated in the absence of inducer was substracted. Results are expressed as percent of the specific binding measured in the absence of competitor (100%). This value of specific binding represents a bound radioactivity of ≠30% of the total. The added MoAb were MoAb 701 (▴), 710 (◊), 724 (○), 322 (▪) and 6D1 (□). MoAb 454 (•) was used as a control.