Peripheral immune tolerance following iv administration of Ag has been shown to occur in the absence of B cells. Because different mechanisms have been identified for iv vs low dose oral tolerance and B cells are a predominant component of the gut-associated lymphoid tissue (GALT) they may play a role in tolerance induction following oral Ag. To examine the role of B cells in oral tolerance we fed low doses of OVA or myelin oligodendrocyte glycoprotein to B cell-deficient (μMT) and wild-type C57BL/6 mice. Results showed that the GALT of naive wild-type and μMT mice was characterized by major differences in the cytokine microenvironment. Feeding low doses of 0.5 mg OVA or 250 μg myelin oligodendrocyte glycoprotein resulted in up-regulation of IL-4, IL-10, and TGF-β in the GALT of wild-type but not μMT mice. Upon stimulation of popliteal node cells, in vitro induction of regulatory cytokines TGF-β and IL-10 was observed in wild-type but not μMT mice. Greater protection against experimental autoimmune encephalomyelitis was found in wild-type mice. Oral tolerance in μMT and wild-type mice was found to proceed by different mechanisms. Anergy was observed from 0.5 mg to 250 ng in μMT mice but not in wild-type mice. Increased Ag was detected in the lymph of μMT mice. No cytokine-mediated suppression was found following lower doses from 100 ng to 500 pg in either group. These results demonstrate the importance of the B cell for the induction of cytokine-mediated suppression associated with low doses of Ag.