Measurement of infarct volume and brain edema

Infarction volume was evaluated by 2,3,5-triphenyltetrazolium chloride (TTC) staining. Freshly removed brains were cut into 2-mm-thick coronal slices using a mice brain matrix. The slices were stained with 2% TTC (Sigma-Aldrich) for 30 min at 37°C in the dark, followed by fixation with 10% formalin. The stained slices were scanned and analyzed with Image J software (NIH, Bethesda, MD, USA).

Wet weight of removed brains was measured immediately after resection and dry weight was measured after brains were dried at 95°C for 24 hours in an oven. Brain Water Content (%) was calculated as [(wet weight-dry weight)/wet weight]×100 [22].