CD100⁻CD8⁺ T cells are functionally impaired after stimulation. (A) Representative ELISPOT wells (left column) illustrating IFN-γ production in response to PMA/ionomycin in FACS sorted CD100⁺ or CD100⁻ CD8⁺ T cells from HCs (n = 3). Control wells containing cells in media only are shown on the right column of wells. (B) Summary of 3 independent ELISPOT experiments. (C) Assessment of IFNγ, MIP-1β IL-2, TNF-α, perforin, and granzyme B production by Luminex assay after PMA/ionomycin stimulation representing cytokine production per 1000 cells (n = 9). D) Assessment of IL-2, IFN-γ, perforin production by Luminex assay after CMV antigen-specific stimulation with CMV peptide-pulsed autologous PBMCs representing cytokine production per 1000 antigen-specific T cells (n = 6). Statistical analysis was performed using (B) Student t tests (***P < .001), and (C-D) 2-tailed paired t tests.