Mice treated with caADAMTS13 1 hour after distal thrombotic MCAo exhibit reduced neutrophil recruitment to the brain parenchyma and reduced peri-infarct microglial activation at 24 hours postocclusion. (A) Recruitment of neutrophils to the ischemic tissue was visualized by IF staining for the neutrophil marker Ly-6G. Images are a representative field of view, chosen to visualize the main site of thrombosis (#), isolated from whole-brain imaging performed using Slidescanner. Scale bars, 200 μm. (B) The number of Ly-6G⁺ particles, with an area greater than 80 μm², was measured using automated particle counting applied to thresholded images in ImageJ. (C) Higher magnification images of regions 1 to 3 (indicated in panel A) of a brain section from a vehicle-treated animal. Neutrophils recruited to a VWF-lined vessel (∞) and 4′,6-diamidino-2-phenylindole/Ly-6G–stained cells (presumed to be neutrophils) in the brain parenchyma (white arrows) were observed. Scale bars, 20 μm. (D) The total number of Iba1⁺ cells and the number of Iba1⁺ cells with a circularity >0.5 were counted by automated particle analysis of thresholded single-channel images and used to quantify the percentage of total microglia with an activated morphology. (E) Representative Iba1 images from the peri-infarct cortical region of the ipsilateral hemisphere and a corresponding region of the contralateral hemisphere for comparison. Scale bars, 100 μm. Inset images are the ipsilateral field of view at higher magnification to show microglial morphology used to determine activation status. Scale bars, 50 μm. Error bars represent mean ± standard deviation. Single comparisons were performed using an unpaired Student t test. *P < .05, ***P < .001.