Acute ischemic stroke (AIS) is one of the most common causes of morbidity and mortality globally [1]. AIS is treated with reperfusion therapies, including intravenous thrombolytic therapy (IVT), endovascular thrombectomy (EVT), or a combination of the two. IVT targets the occlusive thrombus by systemic infusion of tissue plasminogen activator (tPA). Thrombolytic therapy intends to dissolve the thrombus structure and restore normal physiological flow to the affected region by breaking down the fibrin fiber matrix, whereas EVT relies on mechanical extraction of the thrombus material. In both cases, recanalization success is dependent on clot properties, including the histological composition and mechanical properties [2,3]. Therefore, models aimed at predicting IVT and EVT outcomes must take account of these inherent clot properties. Here, the effects of thrombolytics on thrombus mechanical properties are studied. Blood clots are formed in vitro with 40% hematocrit reconstituted human blood in a Chandler loop and aged in a plasma-like medium to replicate physiological thrombosis. The aged clots are treated for one hour in either human plasma only (negative control) or plasma with 1.3 µg/mL tPA (physiological IVT dose) and mechanical properties were quantified using a uniaxial tensile test. The clots maintain linear elastic properties in tension regardless of treatment condition. Application of tPA results in microstructural changes that reduce the average bulk elastic modulus from 5.78 ± 0.67 kPa to 2.70 ± 0.46 kPa (p < 0.01) and the average toughness (area under the stress-strain curve to fracture) from 1.69 ± 0.27 kPa to 0.78 ± 0.14 kPa (p < 0.01). However, the tPA has no significant effect on the fracture strain (0.75 ± 0.06 vs. 0.77 ± 0.05, p = 0.79). These results suggest IVT contributes to clot stiffness reduction without increasing fracture risk, which could make them more susceptible to removal by aspiration thrombectomy [4].