Because of the demonstrated ability of fibronectin to mediate particle uptake by macrophages and the demonstrated affinity of plasma fibronectin for fibrin, we investigated the ability of plasma fibronectin to augment macrophage binding of fibrin. Fibronectin significantly increased fibrin binding by elicited peritoneal macrophages and isolated hepatic Kupffer cells. The binding of fibrinogen was not augmented in the presence of fibronectin. The small amount of macrophage-associated fibrin observed in the absence of fibronectin was primarily internalized, whereas the increment in fibrin binding in the presence of fibronectin remained primarily surface bound, as indicated by susceptibility to removal by trypsin. An amino terminal fibrin-binding fragment of plasma fibronectin could similarly support binding of fibrin by peritoneal macrophages. Greater quantities of fibrin were associated with the macrophages in the presence of protease inhibitors, which inhibited elastase activity, but not in the presence of those that inhibited cathepsin activity, suggesting that an elastase-like protease may degrade surface-bound fibrin. Uptake of both fibrin and fibronectin was inhibited by prior treatment of cells with trypsin. Competitive binding studies suggested the presence of a high-affinity fibronectin receptor on peritoneal macrophages. Data from the current study thus support the conclusion that fibronectin augments binding of fibrin to the surface of mononuclear phagocytes.