Cerebral infarction size

2,3,5-triphenyltetrazolium chloride (TTC) staining was used to assess the cerebral infarction size. Following cerebral ischemia/reperfusion injury for 24 hours, rats were euthanized under deep anesthesia with pentobarbital sodium (i.v. 100 mg/kg). The brains were quickly removed and frozen at -20°C for 10 min. Then, sections with a 2-mm thickness were cut using brain microtome and the slices were stained with 2% TTC solution (Sigma, USA) for 20 min at 37°C in a dark place followed by fixation with 4% paraformaldehyde overnight. The normal brain tissue was stained dark red, whereas the infracted area remained pale white. The infarct area was analyzed by an observer blinded to the experimental conditions using Image-Pro plus 6 and the percentage of infarction volume was calculated as the ratio of the infarcted area to the overall area.