IL-10-producing regulatory B cells have been identified in mice and shown to downregulate inflammation, making them potentially important for maintenance of tolerance. In this study, we isolated B cells from human blood and spleen, and showed that after a short period of ex vivo stimulation a number of these cells produced IL-10. The IL-10-producing B cells did not fall within a single clearly defined subpopulation, but were enriched in both the memory (CD27(+)) and the transitional (CD38(high)) B-cell compartments. Combined CpG-B+anti-Ig stimulation was the most potent IL-10 stimulus tested. B cells stimulated in this way inhibited CD4(+)CD25(-) T-cell proliferation in vitro by a partially IL-10-dependent mechanism. These findings imply that manipulating IL-10 production by human B cells could be a useful therapeutic strategy for modulating immune responses in humans.