Shear-resistant arrest of circulating tumor cells is required for metastasis from the blood stream. Arrest during blood flow can be supported by tumor cell interaction with attached, activated platelets. This is mediated by tumor cell integrin alpha(v)beta3 and cross-linking plasma protein ligands. To analyze the mechanism of tumor cell ligand interactions under dynamic flow conditions, we used real-time video microscopy and tested human melanoma cell binding to fibrinogen, von Willebrand Factor, or fibronectin matrices in a buffer perfusion system. When perfused at venous flow, melanoma cells arrested abruptly and began to spread immediately. This was uniquely mediated by integrin alpha(v)beta3 on all tested ligands, and required alpha(v)beta3 activation and actin polymerization. Under static conditions, alpha(v)beta3 cooperated with alpha(v)beta1 and alpha5beta1 in supporting melanoma cell adhesion to fibronectin. But even when activated, beta1 integrins did not contribute to melanoma cell arrest during flow. Soluble ligand served as a cross-linker between attached and circulating tumor cells and enhanced melanoma cell arrest. Cohesion of activated melanoma cells was restricted to the matrix surface and did not occur in suspension. We conclude that the presence of alpha(v)beta3 in a functionally activated state provides a unique advantage for circulating tumor cells by promoting tumor cell arrest in the presence of flow-dependent shear forces.