Nucleosome (NS)-reactive autoantibodies appear to be involved in the pathogenesis of systemic lupus erythematosus (SLE), but the mechanisms responsible for their production remain elusive. To study whether antibodies specific to NS are present prior to anti-double-stranded (ds) DNA antibodies in lupus mice, ELISA was carried out using the plates which had been confirmed to be coated with comparable amounts of oligo-NS and dsDNA. IgG antibodies reactive with NS or dsDNA were both detected as early as 6 weeks and 10 weeks of age in MRL/lpr and MRL/+ mice, respectively. Their titers increased with age. The specificity of dsDNA binding activity in these young mice was confirmed by the finding that their sera reacted not only with mammalian dsDNA but also with dsDNA from different sources including synthetic polynucleotides. On the other hand, NS binding activity in young mice could be ascribed to cross-reactivity of the anti-dsDNA antibodies, because it was completely inhibited by free dsDNA. A role for NS or chromatin in triggering the production of anti-dsDNA antibodies has recently been suggested, but the present results argue that, at least in these strains of mice, antibodies specifically recognizing conformational epitopes on NS are not produced before those specific to dsDNA.