CD4(+)CD25(+)Foxp3(+) regulatory T cells (CD25(+) Tregs) direct the maintenance of immunological self-tolerance by active suppression of autoaggressive T cell populations. However, the molecules mediating the anergic state and regulatory function of CD25(+) Tregs are still elusive. Using differential proteomics, we identified galectin-10, a member of the lectin family, as constitutively expressed in human CD25(+) Tregs while nearly absent in resting and activated CD4(+) T cells. These data were confirmed on mRNA and protein level. Single cell staining and flow cytometry showed a strictly intracellular expression of galectin-10 in CD25(+) Tregs. Specific inhibition of galectin-10 restored the proliferative capacity of CD25(+) Tregs and abrogated their suppressive function. Notably, first identified here as expressed in human T lymphocytes, galectin-10 is essential for the functional properties of CD25(+) Tregs.