Diabetogenic strains of Coxsackievirus B4 (CB4) produce a diabetes syndrome in susceptible mice that resembles insulin-dependent diabetes mellitus. To assess the possible role of autoimmunity, the expression of a 64,000-M_r islet antigen in SJL/J and CD1 mice infected with a diabetogenic strain of CB4 was monitored in early and late infection. Additionally, virus-induced abnormalities in glucose metabolism were correlated with several changes in purified islets to assess β-cell physiology. Over 80% of the mice exhibited subnormal blood glucose at 72 h postinfection (p.i.) and were hyperglycemie at 6 and 8 wk p.i. Islet yield in infected mice decreased by 29–47% at 72 h and 6 wk p.i. compared to noninfected mice. Insulin release stimulated by 16.7 mM glucose increased greater than twofold at 72 h p.i. but declined at 6 wk well below the level of noninfected mice. Likewise, residual islet insulin content after release also increased at 72 h and then declined. Total protein synthesis in the islets decreased by 30% at 72 h and by 60% at 6 wk p.i. Although the synthesis of five proteins of heterogeneous molecular weights, including tubulin, was severely depressed in the infected islets at 72 h p.i. compared with control islets or islets at 6 wk p.i., synthesis of the 64,000-M_r component and another protein of 36,000 M_r increased by two- to threefold. It is possible that CB4 infection may initiate or enhance an autoimmune reaction by increased expression of the 64,000-M_r antigen.