Two different subsets of CD8⁺, CD57⁺ cells have been defined, one expressing high levels (CD8_high⁺(CD57⁺)), the other expressing low levels of surface CD8 (CD8_low⁺(CD57⁺)). Increased numbers of CD8_high⁺(CD57⁺) cells correlated with previous HCMV infection. By three-colour fluorescence analysis, the CD8_high⁺(CD57 ⁺) population expressed T cell markers such as CD3 and CD5, and most were αβ T cell receptor (αβ TCR)-positive. A significant proportion also expressed CD71 (transferrin receptor) and MHC class II, although little if any CD25 (IL-2R-p55). Some (≥ 40%) co-expressed CD45RA and CD45RO. The CD8_low⁺ (CD57⁺) population expressed classical natural killer (NK) cell markers—CD2, CD16 and CD56. The two subsets were also functionally distinct; CD8_high⁺ (CD57⁺) cells suppressed pokeweed mitogen (PWM)-driven, but not phytohaem-agglutinin (PHA)-driven proliferation and immunoglobulin production; CD8_low⁺ (CD57⁺) cells exhibited NK cytotoxic activity which was not increased by interferon-alpha (IFN-α). Supernatant from cultured CD8_high⁺ (CD57⁺) cells suppressed PWM-driven immunoglobulin production, but not proliferation, and this effect was abrogated by physical separation with tissue culture inserts. Thus, a T cell subset expressing activation and memory T cell markers with direct non-specific suppressor activity was present in peripheral blood mononuclear cells (PBMC) of healthy subjects with asymptomatic HCMV infection.