HHV6B antigen-specific CTLs are polyfunctional. (A) IFN-γ cytokine production, as evaluated by ICS, from CD3⁺/CD8⁻ (CD4 helper; bottom quadrant) and CD3⁺/CD8⁺ (cytotoxic; top quadrant) CTLs on day 9 after stimulation in 1 representative donor (dot plots shown were gated on CD3⁺ cells; left panel). Right panel: Summary results from 5 donors (mean ± SEM). (B) Polycytokine production by CTLs as assessed using ICS in 1 representative donor (left panel) and the summary results from 5 donors showing double-cytokine (IFN-γ and TNF-α) ■ or single cytokine-producing T cells IFN-γ only or TNF-α only with specificity for U11, U14, U54, and U90 (right panel). U71 was omitted from the analysis because of the low frequency of reactive T cells detected. (C) HHV6-specific T-cell lines generated from 4 donors (IFN-γ ELISPOT; top panel) evaluated 9 days after pepmix stimulation were also cytotoxic (bottom panel), as evaluated by standard 4-6 hours Cr⁵¹ release assay using pepmix-pulsed monocytes as targets. Results are presented as percentage specific lysis (mean ± SD, n = 4).