The development of T cell–mediated autoimmune diseases hinges on the balance between effector and regulatory mechanisms. Using two transgenic mouse lines expressing identical myelin basic protein (MBP)–specific T cell receptor (TCR) genes, we have previously shown that mice bearing exclusively MBP-specific T cells (designated T/R⁻) spontaneously develop experimental autoimmune encephalomyelitis (EAE), whereas mice bearing MBP-specific T cells as well as other lymphocytes (designated T/R⁺) did not. Here we demonstrate that T/R⁻ mice can be protected from EAE by the early transfer of total splenocytes or purified CD4⁺ T cells from normal donors. Moreover, whereas T/R⁺ mice crossed with B cell–deficient, γ/δ T cell–deficient, or major histocompatibility complex class I–deficient mice did not develop EAE spontaneously, T/R⁺ mice crossed with TCR-α and -β knockout mice developed EAE with the same incidence and severity as T/R⁻ mice. In addition, MBP-specific transgenic mice that lack only endogenous TCR-α chains developed EAE with high incidence but reduced severity. Surprisingly, two-thirds of MBP-specific transgenic mice lacking only endogenous TCR-β chains also developed EAE, suggesting that in T/R⁺ mice, cells with high protective activity escape TCR-β chain allelic exclusion. Our study identifies CD4⁺ T cells bearing endogenous α and β TCR chains as the lymphocytes that prevent spontaneous EAE in T/R⁺ mice.