Subtilisin QK, a new fibrinolytic enzyme, could cleave directly cross-linked fibrin in vitro. To verify the thrombolytic function of Subtilisin QK in vivo, the thrombolytic effect of purified Subtilisin QK on tail-thrombus of mouse was investigated. After injected with carrageenan, the tail-thrombus of Subtilisin QK treated group were shorter than the physiological saline treated group. Moreover, the tail-thrombus decreased correlate with Subtilisin QK in a dose-dependent manner. Thrombus nearly disappeared while the mice were treated with 12000 IU Subtilisin QK. The result indicated that Subtilisin QK significantly inhibited thrombus formation in mouse tail. This study made more foundation for further development of Subtilisin QK as a novel bifunctional thrombolytic agent.