To identify whether sera containing antinuclear antibody (ANA) from patients with juvenile rheumatoid arthritis (JRA) recognize a common antigen, extracts of HeLa nuclei were prepared by incubating the nuclei with buffers of sequentially increasing salt concentrations (0.1 M, 0.3 M, and 0.5 M NaCl). Western blot analysis using sera from 82 patients with JRA revealed that 63% of ANA positive JRA sera reacted with 2 bands (50-40 kDa) in the 0.3 M extract whereas only 14% of ANA negative JRA sera had antibody to these bands. Antibody to the 50-40 kDa antigen(s) strongly correlated with the presence of ANA by immunofluorescence and was found in 70% of patients with iritis; however, it did not correlate with disease subgroup or disease activity.