AbstractTolerance of macrophages to endotoxin (lipopolysaccharide, LPS) can be induced in vitro by LPS itself. We show here that one of the mechanisms of tolerance to LPS can be mediated via an autocrine process. Continuous exposure to LPS is not required to induce macrophage desensitization. Refractoriness to production of tumor necrosis factor (TNF) in response to LPS can be transferred from tolerant to naive macrophage populations by incubation of the latter with the culture supernatant of the former, in the absence of endotoxin. The active factor present in this macrophage‐desensitizing culture supernatant (MD‐Sup) is more efficiently removed by incubation with tolerant macrophages than by incubation with naive macrophages. The refractoriness elicited by treatment with MD‐Sup is restricted to a decreased TNF response to LPS; interleukin‐1 (IL‐1) and IL‐6 responses are not affected.