Endothelial cells secrete prothrombotic ultralarge von Willebrand factor (VWF) multimers, and the metalloprotease ADAMTS13 cleaves them into smaller, less dangerous multimers. This reaction is stimulated by tensile force applied to the VWF substrate, which may occur on cell surfaces or in the circulating blood. The cleavage of soluble VWF by ADAMTS13 was accelerated dramatically by a combination of platelets and fluid shear stress applied in a cone-plate viscometer. Platelet-dependent cleavage of VWF was blocked by an anti-GPIbα monoclonal antibody or by a recombinant soluble fragment of GPIbα that prevents platelet-VWF binding. Multimeric gel analysis showed that shear and platelet-dependent cleavage consumed large VWF multimers. Therefore, ADAMTS13 preferentially acts on platelet-VWF complexes under fluid shear stress. This reaction is likely to account for a majority of VWF proteolysis after secretion and to determine the steady-state size distribution of circulating VWF multimers in vivo.