Image j system

Whole brain images were taken with a Nikon digital camera and the percent of damaged tissue was identified using the Image J^™ System (Media Cybernetics, Silver Spring, MD). The percentage of injured tissue for each rat was calculated by tracing the perimeter of the injured area and the determining its surface area, compared to the diameter of the whole cerebral cortex.

Like the previously tested males, at 1 month post-injury, the females were fatally anesthetized and their brains perfused with saline, then with 10% formalin, and then extracted for histological analysis. The brains were cut into 20-μm sections on a cryostat and stored at −80°C on 1% gelatin coated (subbed) slides. Slides were stained in 0.1% cresyl violet solution (0.1 g cresyl violet acetate and 0.3 ml of glacial acetic acid dissolved in 100 ml dH₂O) for 10 min at 45°C and then rinsed in distilled water. The percent of damaged issue was identified in six 20-micron Nissl-stained sections and analyzed from between 4.5 and -0.5 mm from β for lesion size using the Image J^™ System (Media Cybernetics, Silver Spring, MD).
Stained slides were scanned with Silverfast Pathscan software (PathScan Enabler IV, Meyer Instruments, Houston TX) and the scanned images were analyzed using ImageJ^™. The percentage of injured tissue from a single section was calculated by tracing the perimeter of the injury and determining its surface area, dividing this by an estimate of the total surface area of the section (taken by tracing both the remaining tissue and the estimated perimeter of the necrotic cavity), and multiplying by 100.