Vascular diseases leading to thrombo-occlusion are the leading cause of morbidity and mortality worldwide. Revascularization and restoration of antegrade blood flow is critical for tissue survival and patient health. In this aspect, systemic administration of thrombolytics (e.g., streptokinase) to dissolve occlusive thrombi is a clinically established strategy. However, this strategy typically necessitates the administration of large doses, leading to a high incidence of hemorrhagic complications due to systemic side effects. To minimize this risk, liposomes specifically targeted to the site of thrombo-occlusion have been bioengineered by exploiting ligand–receptor relationships pertinent to thrombus-associated cell phenotypes. This study focuses on encapsulating streptokinase within these liposomes, specifically regarding the effect of liposome processing conditions on streptokinase encapsulation and activity. Theoretical calculations of encapsulation capacity agreed well with that reported in the literature. The experimental encapsulation efficiency values are 45.9 ± 34.0% (n = 9 ± SD) and 21.6 ± 30.0% (n = 6 ± SD), using two different methods. The liposome processing conditions are found to decrease streptokinase activity; however, over 30% remain active after processing, maintaining enough activity to be therapeutic especially when protected inside a vehicle targeted to the site of thrombo-occlusion. The insight gained from the research reported here would enable refining the design and the processing conditions of liposomal formulations of fibrinolytics to yield reduced variability in encapsulation efficiency and streptokinase activity. The design of a thrombus-targeted ‘stealth’ liposome reported earlier and the current findings of fibrinolytics' encapsulation and activity in such liposomes can be efficiently integrated to develop an efficient strategy for vascular nanomedicine.