Theiler's murine encephalomyelitis virus (TMEV)-induced demyelinating disease is a relevant mouse model of multiple sclerosis. Demyelination is linked to persistent TMEV infection of the central nervous system and characterized by perivascular inflammatory mononuclear infiltrates and primary demyelination. Myelin damage is a T cell-dependent process and susceptibility correlates with the temporal development of chronic virus-specific delayed-type hypersensitivity (DTH) responses. Our previous results have shown that inflammatory processes mediated by Th1 cells specific for a determinant(s) on virus capsid protein 2 (VP2) play a major immunopathologic role in SJL/J mice. This study identifies a 13 amino acid peptide on VP2 (VP2(74-86)) as the immunodominant T cell epitope in TMEV-infected and -immunized SJL/J mice, and demonstrates the ability of that sequence to prime for the majority of the SJL/J DTH T cell response to intact TMEV. The importance of T cell responses to this epitope in the demyelinating process was illustrated by experiments in which SJL/J mice displayed an increased incidence and accelerated onset of clinical disease after peripheral immunization with a fusion protein containing VP2(74-84) before intracerebral infection with a suboptimal dose of the BeAn strain of TMEV. Identification of this immunopathologic TMEV T cell epitope will be critically important for delineation of the mechanisms of T cell-mediated myelin damage and for potential use to prevent and/or treat TMEV-induced demyelinating disease via the induction of epitope-specific tolerance.