The mechanism ofin vitrohyposensitization was examined in isolated rat mast cells. Surface distribution of immunoglobulin was examined by immunofluorescence technique. Hyposensitization could be ascribed neither to changes in the distribution of surface immunoglobulins nor to capping or shedding phenomena. The distribution of cell‐bound antigen and the number of antigen‐positive cells (patch‐bearing cells) was not changed by hyposensitization. The binding of antigen to surface Ig was quantitated by I125‐labelled antigen. In rats sensitized to two different antigens, hyposensitization of the mast cells with one of these inhibited the subsequent histamine release induced by either of the two antigens, whereas no changes were observed in the antigen binding capacity.The present investigation indicates that the mechanism ofin vitrohyposensitization is the result of events secondary to the initial binding of antigen to the mast cell surface immunog