Anti-histone antibodies (AHA) have been demonstrated frequently in the sera of patients with systemic lupus erythematosus (SLE). In the present studies, we found that peripheral blood mononuclear leukocytes (PBL) from a large subset of SLE patients spontaneously produce elevated levels of AHA in culture. In contrast, detectable production by normal mononuclear cells was extremely rare. Spontaneous production by patients' PBL correlated with both disease activity and elevated serum AHA levels, and thus appeared to reflect in vivo production. Interestingly, spontaneous AHA production was independent of polyclonal B-cell activation as measured by total Ig synthesis in culture. Production also appeared to be T-cell-independent in that cultures depleted of T cells produced AHA levels similar to those of cultures with unseparated PBL. Although PBL from normal individuals rarely produce AHA spontaneously, the presence of histone-specific B cells in normal peripheral blood could be detected after pokeweed mitogen stimulation. The present studies provide a basis for a further understanding of the mechanisms responsible for autoantibody production in SLE.