ICAM-2 exhibits differences from ICAM-1 and ICAM-3 in mediating perforin and granzyme release from CD56⁺CD8⁺ CTL subsets. (A) IL-2–activated PBMCs (200 ng/mL, 12 hours) were mock-treated (IgG) or treated with ICAM-1, ICAM-2, or ICAM-3 (10 μg/mL Fc fusion protein) 30 minutes prior to incubation with target HL60 cells at a 1:50 E/T ratio for 4 hours. Cells were then prepared for flow cytometry with CD8-Cy5PE, CD56-PE surface stains, and perforin-Cy5 and granzyme A–FITC intracellular stains. Cells were gated for CD56⁺CD8^low, CD56⁺CD8^med, CD56⁺CD8^high, CD56^–CD8^–, CD56^–CD8^high populations as shown in panel i and population frequencies within appropriate gate. Panels ii-vii are subset-gated populations displayed for perforin and granzyme A log fluorescent intensities. Results are representative of 3 independent experiments and were similar at 1:25 and 1:12.5 E/T ratios (data not shown). Numbers in each panel correspond to population frequencies. (B) Relative perforin changes in CD56CD8 subsets after target cell mixing. Median fluorescence values were computed for CD56CD8 subsets and made relative to median fluorescence values. Error bars show standard deviations of 3 experiments.