When human W25I-labeled Factor Xa is incubated with washed platelets, prothrombin, and Ca2+, a small amount of thrombin is formed which causes the platelet release reaction after a period of time that decreases as the Xa concentration is increased from 0.9 to 19 ng/ml. After a further lag period, the Xa binds reversibly to receptors on the platelet surface and rapid thrombin formation follows (3 units The results suggest that specific receptors for Xa appear on the platelet surface after the release reaction occurs. The bound Xa catalyzes thrombin formation 1000-fold faster than does Xa added to reactions in which phospholipids are substituted for platelets.The first stage in hemostasis is thought to be adhesion of platelets to subendothelial collagen or other substances at sites of injury. The platelet release reaction then occurs and secretion of numerous substances, including ADP and products of arachidonic acid transformation, is believed to promote further platelet aggregation with the eventual formation of a primary hemostatic plug. Later, fibrin accumulates to provide a more permanent arrest of bleeding. Understanding of the interactions between platelets and plasma coagulation factors in generating a fibrin clot is incomplete. Numerous investigators have observed that platelets accelerate the clotting of plasma (for review, see ref. 1). The term "platelet factor 3" historically has been used to describe this procoagulant activity, although neither its subcellular localization nor its chemical nature has been clearly defined (2).Factor Xa catalyzes the conversion of prothrombin to thrombin. Factor V, Ca2+, and phospholipid increase the rate of the reaction by organizing the enzyme and substrate into a lipoprotein complex (3). Milstone (4) showed that phospholipid and Factor V can replace platelets in prothrombin activation. Phospholipid can substitute for platelets in various assays that measure the clotting time of plasma as well, and it has been proposed that platelets may promote clotting by providing a phospholipid surface (5). Marcus et al. (6) suggested that more than phospholipid is involved because 20 times more extracted platelet phospholipid is required to achieve the same clotpromoting activity as that present in the intact membranes.Experiments to measure the interaction of individual plasma coagulation factors with platelets require purified factors andThe costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact. platelets washed free of plasma. We have previously described the interaction of thrombin with washed human platelets (7).Numerous experiments suggest that the first step in the action of thrombin on platelets is binding of thrombin to a specific cell surface receptor (7-10). We now report the first of our studies to determine if thrombin is actually formed on the platelet surface. We have measured the interaction ...