Peripheral immune cells may participate to a lesser extent in delayed tPA–exacerbated brain injury during the early phase of ischemic stroke. Mice were subjected to 3-hour MCAO followed by vehicle, tPA, or tPA plus IFNβ administration (n = 6-7 per group) or subjected to 40-minute MCAO as positive control (n = 3). Three-hour MCAO mice administered vehicle, tPA, or tPA plus IFNβ were euthanized at 13 hours postinjury, and 40-minute MCAO mice were euthanized at 48 hours postinjury. The ischemic brains were harvested and subjected to mononuclear cells isolation. The isolated mononuclear cells were then subjected to staining with antibodies against CD45, CD11b, Ly6G, and CD3 followed by FACS analysis. (A) The infiltrating myeloid immune cells were determined based on their expression of CD45^hiCD11b⁺, and neutrophils were determined based on their positive expression of Ly6G within the population of CD45^hiCD11b⁺ cells. T cells were determined based on their positive expression of CD3 within the population of CD45^hiCD11b⁻ cells. The numbers of total infiltrating myeloid immune cells (B), neutrophils (C), and T cells (D) in the contralateral and ipsilateral hemispheres of ischemic brains were analyzed. C, contralateral hemisphere; I, ipsilateral hemisphere.