Oral administration of antigen induces a state of tolerance that is associated with activation of CD8+ T cells that can transfer unresponsiveness to naïve syngeneic hosts. These T cells are not lytic, but they inhibit development of antibody, CD4+ T helper cell, and CD8+ ytotoxic T lymphocyte (CTL) responses upon adoptive transfer into naïve, syngeneic mice. In addition, we have shown that depletion of γδ T cells by injection of the anti-δ chain antibody (GL3) down modulates the expression of γδ T-cell receptor (TCR) and inhibits the induction of oral tolerance to ovalbumin. Oral administration of antigen also fails to induce tolerance in TCR δ-chain knockout mice suggesting that γδ T cells play a critical, active role in tolerance induced by orally administered antigen. To further study the contribution of γδ T cells to tolerance, murine γδ T cells were isolated from intraepithelial lymphocytes (IEL) of the small intestine by stimulation with splenic filler cells, concanavalin A and growth factors. γδ IEL lines demonstrated lytic activity in a redirected lysis assay. γδ T-cell clones express different γδ TCR genes and secrete large amounts of interleukin (IL)-10, but little or no IL-2, IL-4, or interferon-γ. γδ IEL clones expressed transforming growth factor-β1 and macrophage migration inhibitory factor, as well as IL-10, mRNA. Moreover, γδ T-cell clones potently inhibited the generation of CTL responses by secreted molecules rather than by direct cell-to-cell contact.