Hepatitis C virus (HCV) infection has been found to be strikingly associated with autoimmune phenomena. The aim of the present study was to investigate the presence of various autoantibodies in patients with HCV infection. Anti-neutrophil cytoplamic antibody (ANCA), anti-dihydrolipoamide dehydrogenase (anti-E3), rheumatoid factor (RF), anti-dihydrolipoamide acetyltransferase (anti-E2), anti-SS-A/Ro (60 kD), anti-SS-A/Ro (52 kD), anti-SS-B/La, anti-topoisomerase II (anti-topo II), anti-cardiolipin (aCL), anti-dsDNA, anti-ssDNA, anti-nuclear antibodies (ANA), anti-proteinase 3 (anti-Pr3) and anti-myeloperoxidase (anti-MPO) were determined in sera from 516 patients with HCV infection, 11 with primary biliary cirrhosis (PBC) and 44 healthy controls. Assays employed were indirect immunofluoresence, the particle latex agglutination test, enzyme-linked immunosorbent assay (ELISA) and immunoblotting. ANCA, anti-E3 antibody and RF were positive in 278/516 (55.6%), 276/516 (53.3%) and 288/516 (56%) patients with HCV infection, respectively. Positivity for ANA was present in 15.8%, anti-ssDNA in 15.6%, anti-dsDNA in 8.5%, aCL in 5%, anti-SS-B/La in 4.1%, anti-SS-A/Ro (60 kD) in 3.9%, anti-E2 in 3.3% and anti-SSA/Ro (52 kD) in 1.2%, anti-MPO in 4.8%, anti-Topo II and anti-actinin in 0%. All sera with ANCA showed c-ANCA patterns and contained anti-PR3 specificity. HCV patients with ANCA showed a higher prevalence of skin involvement, anaemia, abnormal liver function and alpha-Fetoprotein (alpha-FP). HCV patients with anti-E3 antibodies showed a higher prevalence of liver cirrhosis, arthritis, abnormal liver function and elevated alpha-FP levels. The prevalence of autoantibodies was not affected by treatment with interferon-alpha (IFN-alpha). In conclusion, autoantibodies are commonly found in patients with HCV infection. There is a high prevalence of anti-E3, ANCA and RF in these patients. Proteinase 3 and E3 are the major target antigens in HCV infection. HCV may be regarded as a possible causative factor in ANCA-related vasculitis.