Antibodies to double-stranded DNA (dsDNA), generally regarded as highly specific for systemic lupus erythematosus (SLE), have also been reported in chronic active hepatitis (CAH). Using the Farr assay and E. coli DNA, fractionated by benzoylated-naphthoylated-DEAE cellulose chromatography into dsDNA and dsDNA containing single-stranded regions, we compared the serum DNA binding of CAH and SLE patients. Although CAH sera were found to have dsDNA binding significantly above the normal control group such binding was of low level and we could find no evidence of markedly elevated dsDNA binding in CAH. However 12 of the 20 CAH sera studied did bind preferentially to dsDNA containing single-stranded regions suggesting the presence of anti-single-stranded DNA antibodies. We conclude that the description of elevated anti-dsDNA antibodies, as measured by the Farr assay, in CAH is due to the interaction of anti-single-stranded DNA antibodies or other serum components with single-stranded DNA contaminating dsDNA preparations.