OBJECTIVE To partially purify and characterize a specific inhibitor of interleukin 1 alpha (IL-1 alpha) activity from synovial fluids (SF) of patients with rheumatoid arthritis. METHODS SF inhibitor of IL-1 alpha (SFI alpha) was partially purified by ammonium sulfate precipitation, gel filtration, anion exchange chromatography, and chromatofocusing. Specificity of inhibition of IL-1 alpha activity was tested in T lymphocyte proliferation assays. The mechanism of this inhibition was investigated by binding assays and mobility shift on gel filtration. RESULTS The SFI alpha inhibited all in vitro activities of IL-1 alpha tested, but did not inhibit activities of IL-1 beta or IL-2. Inhibitor activity was not diminished by neutralizing antibodies against the IL-1 receptor antagonist, and was not removed by immunoadsorption with antibodies against IL-1 receptors. It was not depleted by monoclonal antibodies against human IgG subclasses, IgA, or IgM. The SFI alpha specifically inhibited binding of IL-1 alpha to cell surface receptors, and appeared to form a high molecular weight complex with IL-1 alpha. CONCLUSIONS SFI alpha appeared to block biological activities of IL-1 alpha by specific binding to this cytokine, thereby preventing receptor occupancy. The SFI alpha did not appear to be related to previously described inhibitors of IL-1.