The use of HLA class II‐transgenic (Tg) mice has facilitated identification of antigenic T cell epitopes that may contribute to inflammation in T cell‐mediated diseases such as rheumatoid arthritis and multiple sclerosis (MS). In this study, we compared the encephalitogenic activity of three DR2‐restricted myelin determinants [mouse (m) myelin oligodendrocyte glycoprotein (MOG)‐35–55, human (h)MOG‐35–55 and myelin basic protein (MBP)‐87–99] in Tg mice expressing the MS‐associated DR2 allele, DRB1*1501. We found that mMOG‐35–55 peptide was strongly immunogenic and induced moderatelysevere chronic experimental autoimmune encephalomyelitis (EAE) with white matter lesions after a single injection in Freund's complete adjuvant followed by pertussis toxin. hMOG‐35–55 peptide,which differs from mMOG‐35–55 peptide by a proline for serine substitution at position 42, was also immunogenic, but not encephalitogenic, and was only partially cross‐reactive with mMOG‐35–55. In contrast, MBP‐87–99, which can induce EAE in double‐Tg mice expressing both HLA‐DR2 and a human MBP‐specific TCR, was completely non‐encephalitogenic in HLA‐DR2‐Tg mice lacking the human TCR transgene. These findings demonstrate potent encephalitogenic activity of the mMOG‐35–55 peptide in association with HLA‐DR2, thus providing a strong rationale for further study of hMOG‐35–55 peptide as a potential pathogenic determinant in humans.