5,6-trans-Prostaglandin E2 (trans-PG E2) was found to accelerate the fibrinolysis in a cell-free system. trans-PG E2 decreased the lysis time by 15% in the fibrin clot lysis time method and increased the lysis area by 22% in the fibrin plate method. On the contrary, 5,6-cis-prostaglandin E2 did not have such effects. trans-PG E2 did not exert the effect in the absence of either tissue-plasminogen activator or plasminogen. These results suggest that trans-PG E2 enhances the generation of plasmin from plasminogen by tissue-plasminogen activator.