A sensitive assay for quantitating bovine activated Factor XI (Factor XIa) in vitro was developed by measuring the amidolytic activity of thrombin generated in a mixture of Factors XIa, IX and X and prothrombin prepared from bovine source and washed bovine platelets. In this system, the rate of thrombin generation increased linearly with increasing amounts (fmoles) of Factor XIa. The assay system for Factor XIa was not significantly affected by the presence of plasma kallikrein, Factor XIIa, high-molecular-weight kininogen, amylose sulfate or sulfatide within the range of the amounts used for surface-mediated activation of Factor XII, prekallikrein and Factor XI. Following surface-mediated activation of Factor XI, further generation of Factor XIa was blocked by adding freeze-thawed platelets that contain cationic proteins which bind to negatively-charged surfaces (J. Biochem. 97, 139-151, 1985). The method is useful to the kinetic analysis of the surface-mediated activation of Factors XII and XI, although it is not applicable to the activation in plasma.