Cd8 4sm15

Artery grafts were perfusion fixed with 4% paraformaldehyde. For histological examination of medial injury, 2 cross-sections per artery that were at least 100 μm apart were H&E stained and the average thickness of the media measured in Image J (National Institutes of Health). The medial thickness was normalized to arterial radius.
Immunohistochemistry was performed using antibodies toward myeloperoxidase (Abcam, Cambridge, United Kingdom) to visualize neutrophils, CD4 (4SM95; eBioscience, Waltham, Massachusetts), CD8 (4SM15, eBioscience), Foxp3 (FJK-16S, eBioscience), and Mac-3 (BD Biosciences, Franklin Lakes, New Jersey) to visualize macrophages as described previously. 29 Staining was visualized by aminoethyl carbazole chromagen (red; Vector Laboratories, Burlingame, CA) and counterstained with hematoxylin (blue). For quantification, the total number of cells in each cross-section was manually counted in a blinded manner and reported per mm 2 of media or adventitia. Two cross-sections that were at least 100 μm apart were evaluated for each artery and an average value from the cross-sections calculated and used as a single data point.