Serum antipituitary antibodies were investigated by the immunoblotting method using human anterior pituitary membrane preparation as the antigen.Thirteen patients with autoimmune lymphocytic hypophysitis, two patients with infundibuloneurohypophysitis, four patients with isolated ACTH deficiency, 21 patients with diabetes mellitus (type 1 and type 2) and 38 healthy subjects were studied.Human pituitary membrane antigens were electrophoresed by sodium dodecylsulphate‐polyacrimide gel electrophoresis (SDS‐PAGE). The antigens were transferred to polyvinylidene difluoride membrane and reacted with the sera followed by incubation with biotinylated anti‐human IgG goat serum.Serum antibodies to 68, 49, 43 kD human pituitary membrane antigens were detected in five of 13, one of 12 patients with infundibuloneurohypophysitis and none of four patients with isolated ACTH deficiency. These antibodies were not detectable when human thyroid, liver or rat pituitary preparations were used as the antigen.These findings suggest that serum antibodies to 68, 49, 43 kD human anterior pituitary antigen are specific but not so frequently detected in autoimmune lymphocytic hypophysitis.