Type I diabetes mellitus results from the autoimmune destruction of insulin producing beta cells in the pancreas. Certain viral infections, especially those caused by coxsackie B viruses and related enteroviruses, have been associated with the development of type I diabetes. The sequence homology between the coxsackie B4 virus nonstructural protein 2C (CVB4 p2C) and the major diabetes autoantigen glutamic acid decarboxylase (GAD 65 ) provides a basis for the hypothesis of molecular mimicry. In this study, we investigated the prevalence of antibodies directed against nonstructural enterovirus proteins. In addition, a correlation of antibodies against CVB4 p2C and GAD 65 was studied in diabetes patients and in healthy controls. Antibody reactivity against CVB proteins was detected by immunoprecipitation of [ 35 S]-methionine-labelled viral proteins and GAD 65 antibodies were measured in a quantitative radio-immunoassay. It was shown that antibodies raised against the nonstructural proteins of CVB4 are very common in the population and a high degree of heterotypic cross-reactivity exists between different enterovirus types. CVB4 p2Cspecific antibodies were not only detectable in GAD 65 antibody-positive diabetes patients but also in GAD 65 antibody-negative healthy blood donors. Furthermore, GAD 65 antibodies could not be detected in p2C-positive subjects who had various enterovirus infections, indicating that an antibody response to CVB4 p2C does not necessarily induce a cross-reactive immune response against GAD 65 . A correlation was not found between antibodies against GAD 65 and p2C.