In this study, we aimed to develop a thrombus-targeting delivery system of collagenase bound to a monoclonal antibody, and to investigate the thrombolysis of an immune-conjugate in vitro and in vivo as well as the targeting effect. We prepared the immunizing conjugation of collagenase by the 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDCI) method. In order to conjugate collagenase and a monoclonal antibody, bovine serum albumin was used as a linker, increasing the number of collagenase molecules carried and keeping collagenase and the monoclonal antibody active. In vitro thrombolysis experiments showed that collagenase had a strong dissolving effect on collagen-embolus within 24 hours. We established a rabbit pulmonary embolism model to investigate the thrombolysis effect of collagenase and collagenase immunizing conjugation in vivo. Our results revealed a significant difference between collagenase and collagenase immunizing conjugation (P < 0.05). We also established a rabbit ear edge vein model to investigate the active target of collagenase immunizing conjugation. We found that collagenase immunizing conjugation had active targets, and had a strong ability to dissolve organized thrombi. In conclusion, the thrombus-targeting delivery system of collagenase we developed has active targeting effects on thrombi.