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A new autoantibody which reacted with anterior-pituitary tissue was deteected by immunofluorescent staining. Of 287 patients having one or more autoimmune endocrine diseases, sera from 19 reacted with pituitary glands obtained at hypophysectomy for breast cancer. Using specific rabbit antibodies to each of the six pituitary hormones and rhodamine-labelled goat-anti-rabbit-immunoglobulin (Ig) followed on the same unfixed cryostat section by a patients's serum counterstained with fluorescein-conjugated sheep-anti-human-Ig, it was posssible to show that the autoantibodies reacted specifically with the hypertrophied prolactin cells in these glands. The antibodies, belonging to the 3 main Ig classes, were complement fixing, with titres varying from 1 to 80. Double staining of prolactin cells and analogy with other autoimmune systems suggested that the antigen is in cytoplasmic organelles involved in the synthesis or delivery of the hormone. No pituitary antibodies were found in panhypopituitary cases, but there are indications that antibodies to other pituitary cells exist in some sera and that an autoimmune process may account for some cases of isolated pituitary hormone defects occurring in adult life.