We characterized three monoclonal antibodies with histone reactivity which were derived from spleen cells obtained from unmanipulated NZB/NZW or MRL/1 mice. By using an enzyme-linked immunosorbent assay, we noted that all three antibodies reacted with chromatin histones as well as with total histones extracted from chromatin. None of the antibodies appeared to require DNA as part of the antigen. One antibody (BWH-1) recognized a determinant present in the nucleosome core H2A-H2B complex but showed little reactivity with any of the individual histones (H1, H2A, H2B, H3, or H4). In contrast, the other two monoclonal antibodies each recognized multiple individual histones in a unique pattern. Antibody MH-1 reacted with H2A, H2B, and H3; antibody MH-2 reacted with H2A, H3, and H4. MH-1 demonstrated cross-reactivity with poly-1-lysine but not poly-1-arginine or protamine sulfate; the opposite pattern of cross-reactivity was observed with MH-2. The antigenic determinants recognized by MH-2 were all trypsin-sensitive, suggesting that these determinants were present on the N-terminal regions of the respective individual histones. These studies revealed markedly different specificities of anti-histone monoclonal antibodies derived from murine models of systemic lupus erythematosus. These and other similarly derived antibodies may provide interesting tools to understand the specificity and biologic importance of anti-histone autoantibodies in different diseases.