Sera from 102 patients with chronic Chagas' disease were studied for the presence of autoantibodies to intracellular proteins and nucleic acids by three different methods. Only four sera had autoantibodies detected by indirect immunofluorescence on HEp-2 cells. All of the sera were negative for anti-dsDNA, anti-Ro/SSA, anti-La/SSB, anti-Sm and anti-RNP autoantibodies but 12 (12%) of the sera had low to moderate levels of anti-histone antibodies. When Chagas sera were tested for autoantibodies to a total HeLa cell extract by Western blotting, weak reactivity was observed in 31 sera. Despite significant heterogeneity in the protein Ag targeted by these sera, seven recognized a 23-kDa protein. Strong binding to this 23 kDa protein was observed in one-third of the sera when isolated ribosomes were used as source of Ag. In contrast, no autoreactivity was detected with ribosomal proteins P0, P1, and P2. These findings confirm the presence of autoantibodies in chronic Chagas' disease and indicate a remarkable restricted humoral immune response to human ribosomal proteins. Furthermore, affinity-isolated anti-23-kDa antibody cross-reacted with a Trypanosoma cruzi ribosomal protein of similar molecular weight. This molecular mimicry may be responsible for the apparent breakdown of self-tolerance resulting in tissue damage. Indeed, experiments demonstrating that immunization of mice and rabbit with T. cruzi ribosomes have been reported to induce myocarditis.