Sublingual (s.l.) immunotherapy has in the last decade emerged as an effective approach to desensitize patients with pollen, food and insect sting allergies. This treatment has recently also attracted interest as a potential modality to control self-reactive T-cell responses associated with autoimmune disorders. Here, we show that s.l. administration of ovalbumin (OVA) conjugated to cholera toxin B subunit (CTB) (OVA/CTB) can efficiently suppress peripheral effector T (Teff) cell responses to OVA in mice that had adoptively received OVA-specific T-cell receptor (TCR) transgenic CD4(+) T cells, and that the suppression was associated with the development of OVA-specific Foxp3(+)CD25(+)CD4(+) regulatory T (Treg) cells as well as with apoptosis (Annexin V(+)) and depletion of OVA-specific Teff cells in peripheral lymph nodes. The induction of Teff cell apoptosis by s.l. OVA/CTB administration was found to be critically dependent on CD25(+) Treg cells but independent of IL-10 production. Our results suggest that s.l administration of a CTB-conjugated antigen can efficiently induce peripheral Teff cell tolerance through the induction of antigen-specific Treg cells that both inhibit Teff cell proliferation and cytokine production and induce Teff cell apoptosis and depletion.