Rheumatoid arthritis (RA) is a T cell-mediated autoimmune disease, but target antigens (autoantigens) responsible for T cell activation remain unclear. Type II collagen (CII) is a candidate autoantigen that is largely confined to the articular cartilage. To investigate whether CII is an important antigen in patients with RA, we examined peripheral blood T cell reactivity to CII in HLA-DRB1*0101 and DRB1*0405-positive RA patients. Reactivities to candidate T cell epitopes of CII were also examined. Peripheral blood T cell reactivity to CII and CII peptides (256-271, 429-442, 593-610, 1064-1081) were detected by measurement of IL-2, IFN-gamma, and IL-4 in culture supernatant of PBMC after in vitro antigen stimulation. Cytokine concentration was measured by ELISA. In DRB1*0101-positive patients, T cell reactivity to CII as detected by measurement of IL-2 production in culture supernatant, was present in 4 out of 9 patients. IL-2 production upon stimulation with CII 256-271 peptide was found in all of these 4 patients. In DRB1*0405-positive patients, high frequency of positive T cell response to CII was detected in 9 out of 11 patients. IFN-gamma production was also detected in 4 out of 6 patients producing IL-2 by stimulation with CII. T cell response to CII 256-271 and/or CII 1064-1081 was detected in these patients. In DRB1*0101-positive RA patients, CII 256-271 peptide might function as a T cell epitope, whereas either CII 256-271 or CII 1064-1081 peptide may be a major T cell epitope in DRB1*0405-positive RA patients. In DRB1*0405-positive RA patients, CII reactive T cells might play a crucial role in the development of RA through IFN-gamma production.