Antibodies of the immunoglobulin E isotype sensitize mast cells and basophils for antigen-induced mediator release by binding through the Fc portion to a high-affinity receptor (Fc^ɛRl, K^a = 10⁹ M⁻¹) on the cell surface^1,2 causing the clinical manifestations of type I hypersensitivity. As the amino acid sequence of the human epsilon chain is now known³, attempts have been made to map the Fc^ɛRl binding site on IgE to a fragment smaller than Fc^ɛ using proteolytic cleavage products, none of which proved to be active³. Cleavage between the C^ɛ2 and C^ɛ3 domains released two inactive fragments, suggesting that the junction between these segments could be important in receptor binding³. This region is protected against protease digestion in the rat IgE complex with the receptor of rat basophilic leukaemia cells⁴. Here we report the mapping of the mast cell receptor binding site on human IgE to a sequence of 76 amino acids at the C^ɛ2/C^ɛ 3 junction. Recombinant peptides containing this sequence inhibit passive sensitization of skin mast cells in vivo and sensitize mast cells to degranulation by anti-IgE in vitro almost as efficiently as a myeloma IgE. Fragments containing the separate domains are inactive. Additional sequences are required for rapid assembly of fragments into disulphide-linked dimers, suggesting that a single chain can form the active site. In a three-dimensional model of the human Fc^ɛ, the two identical segments are far apart. Each folds to generate a cleft between the C^ɛ2 and C^ɛ3 domains on the surface of the Fc^ɛ. The docking of IgE on to mast cells could take place within this cleft.