Myelin basic protein (MBP)-reactive T cells may play an important role in the autoimmune pathogenesis of multiple sclerosis (MS). MBP-reactive T cells can be specifically targeted by T cell vaccination, a procedure whereby MS patients are immunized with attenuated autologous MBP reactive T cells. T cell vaccination induces immune responses to the vaccine cells together with a depletion of MBP reactive T cells. Forty-nine MS patients were treated with T cell vaccination in an extended phase I trial to study the safety, immune responses and clinical effects of T cell vaccination. In the present paper the immune responses towards the vaccine cells were characterized. Substantial long-term in vitro proliferative responses were observed in all treated patients. Some patients, immunized with different clones, displayed distinct proliferative reactivity against the various vaccine clones, suggesting unequal immunogenic properties of these clones. Reactive TCRαβ⁺, CD8⁺and CD4⁺T cells, and to a lesser extent, γδ T cells and NK cells were observed toin vitro stimulation with the vaccine cells. A small fraction only of CD8⁺T cells expressed cytolytic and inhibitory anti-clonotypic reactivity against the vaccine cells. Stimulation with the vaccine clones predominantly induced expression of pro-inflammatory cytokines in these mixed cultures, although one vaccine clone consistently induced production of IL-4. CD4⁺T cells are the major cytokine-producing cells in these anti-vaccine lines. We could not detect upregulated antibody responses to the vaccine cells in most patients, although a temporary antibody response was observed in one patient. In conclusion, immunization with attenuated autoreactive T cells induces a complex cellular response specifically targeted at the vaccine cells, but no antibody responses. These data provide further insights into the mechanisms of T cell vaccination and improve our understanding of the complex regulatory networks of autoreactive T cells.