We aimed to determine the frequency of new microbleeds after intravenous thrombolysis using contiguous thin-slice 3T magnetic resonance imaging. We retrospectively examined clinical and imaging data from 121 consecutive acute ischemic stroke patients who underwent magnetic resonance imaging before and 24 hours after intravenous thrombolysis. Of the included patients, 44 (36.4%) were women, with a median age of 69 years (range, 35-94 years). A total of 363 baseline microbleeds were observed in 57 patients and 8 new microbleeds in 6 patients. Multiple regression analysis indicated that baseline infarct volume (odds ratio, 1.556/10 mL; 95% CI, 1.017-2.379; P = 0.04) and systolic blood pressure (odds ratio, 1.956/10 mm Hg; 95% CI, 1.056-3.622; P = 0.03), but not the presence of baseline microbleeds, were independently associated with new microbleeds. The frequency of neither symptomatic intracranial hemorrhage nor remote hemorrhage or any hemorrhagic transformation was different between patients with and without new microbleeds (0.0% vs 1.7%, P > 0.99; 0.0% vs 1.7%, P > 0.99; 50.0% vs 28.7%, P =0.36). New microbleeds developed rapidly 24 hours after intravenous thrombolysis. The significance of these new microbleeds and their effect on cognitive and functional outcome merits further investigation.