Dissolution of a blood clot in thrombolytic procedure is affected by permeation of a thrombolytic agent into a clot, and permeation of plasminogen activator into a clot is dependent on the permeation pressure. A controlled experimental study on the effects of permeation pressure on clot lysis was carried out. The effects of intra-thrombus perfusion on thrombolyis were also explored. In vitro clot lysis tests were performed for perfusion pressures of 0, 1, and 10 kPa. Two perfusion media were used: buffer and urokinase solution. Clot lysis by diffusion was enhanced in urokinase solution compared with buffer solution, but the effects of perfusion medium were not significant in pressurized perfusion. Clot lysis was accelerated with the perfusion pressure in the early stage of perfusion, but the pressure effect on thrombolysis was not significant in the later stage (after 2 h). Intra-thrombus perfusion was more efficient in clot lysis than surface perfusion was. It is suggested that fluid flow inside the thrombus should be the most important factor in thrombolytic therapy.