Immunoglobulins can serve as tolerogenic carriers for antigens, and B cells can function as tolerogenic antigen-presenting cells. We used this principle to design a strategy for gene therapy of experimental autoimmune uveitis, a cell-mediated autoimmune disease model for human uveitis induced with the uveitogenic interphotoreceptor retinoid-binding protein (IRBP). A retroviral vector was constructed containing a major uveitogenic IRBP epitope in frame with mouse IgG1 heavy chain. This construct was used to transduce peripheral B cells, which were infused into syngeneic recipients. A single infusion of transduced cells, 10 days before uveitogenic challenge, protected mice from clinical disease induced with the epitope or with the native IRBP protein. Protected mice had reduced antigen-specific responses, but showed no evidence for a classic Th1/Th2 response shift or for generalized anergy. Protection was not transferable, arguing against a mechanism dependent on regulatory cells. Importantly, the treatment was protective when initiated 7 days after uveitogenic immunization or concurrently with adoptive transfer of primed uveitogenic T cells. We suggest that this form of gene therapy can induce epitope-specific protection not only in naive, but also in already primed recipients, thus providing a protocol for treatment of established autoimmunity.