Antibodies to native (n-) DNA and denatured (d-) DNA were detected simultaneously and quantitatively in the sera of patients with systemic lupus erythematosus (SLE) and other rheumatic diseases by a filter method using (14)C•n-DNA or (14)C•d-DNA as the radiolabelled antigen. In a group of patients with SLE, most of the sera had antibodies to both n-DNA and d-DNA, only a few sera had antibody to d-DNA alone. In contrast, none of the sera reacted with n-DNA alone. On the other hand, sera from patients with progressive systemic scleroderma, dermatomyositis, polymyositis or SjΦgren's syndrome had antibody to d-DNA alone. In order to estimate the participation of anti-d-DNA antibody in lupus nephritis, patients with SLE were classified into two groups according to immunofluorescent glomerular staining. In the group of patients that had lumpy or granular staining, the sera reacted predominantly with n-DNA. In contrast, sera from the other group of patients that had mesangial or linear staining had antibodies that mainly reacted with d-DNA. The difference in reactivity between these two groups was statistically significant (p 0.02). In the other group of sera from patients with non-rheumatic diseases, anti-d-DNA antibodies were detected by the half saturated ammonium sulfate method using (125)I•d-DNA as the antigen. Positive incidences of anti-d-DNA antibody in these group were as follows: lung tuberculosis 21%, Graves' disease 55%, Hashimoto's thyroiditis 73%, chronic hepatitis 52%, cancer patients 25%, and patients undergoing hemodialysis 24%. Significant correlation between antinuclear antibody and anti-d-DNA antibody was seen in a group of patients with Hashimoto's thyroiditis, in patients undergoing hemodialysis, and in lung tuberculosis. In patients with chronic hepatitis, significant correlation was seen between HBsAg and anti-dDNA antibody. These results suggest that the anti-d-DNA antibody has an extremely wide distribution in rheumatic and non-rheumatic diseases with the highest titer and incidence in SLE. In SLE sera, however, anti-d-DNA antibody is less relevant to the severity of lupus nephritis and has a close correlation to anti-n-DNA antibody.