Interferon (IFN)—γ synthesis of CD45RO+ (memory) and CD45RA+ (naive) CD8+ cytotoxic T lymphocytes (CTLs) and the role of interleukin (IL)—12 in the regulation of human CTL functions in virus-specific immunity were investigated. After culture with influenza virus, CD45RO+ CD8+ T cells from human peripheral blood mononuclear cells increased in frequency and exhibited significant major histocompatibility complex class I—mediated CTL activity, whereas CD45RA+ CD8+ T cells did not. Influenza virus—stimulated CD45RO+ CD8+ T cells contained significantly higher levels of IFN-γ—producing cells and IFN-γ—specific mRNA than did CD45RA+ CD8+ T cells. Recombinant human IL-12 further enhanced CTL activity and IFN-γ production by CD45RO+ CD8+ T cells. These data clearly show that human virus-specific CTL activity and coproduction of IFN-γ are associated with the CD45RO+ CD8+ T cells that are modulated by the cell-mediated, immunity-inducible cytokine IL-12 in humans.