The purpose of this study was to determine the frequency of antinuclear antibody (ANA) in a normal pediatric population. Sera from 241 children (age range 4 months to 16 years) were tested for IgG ANA by use of indirect immunofluorescence and two commercially available substrates, mouse kidney and human epithelial cells. Patients' sera were tested at three different dilutions. The positivity of the ANA was related to the substrate used. When a screening dilution of 1:5 or 1:10 was used, 2.0% and 1.6% were positive with the mouse kidney or human epithelial cell substrates, respectively, but the positivity dropped to 0.4% when the dilution was 1:20 and 1:40, respectively. Only one serum was positive with both substrates.