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A monoclonal antibody (anti-estrogen receptor [ER] from the ER-immunocytochemical assay [ICA] [Abbott Laboratories, Berkshire, England]), which binds to epitopes of the estrogen receptor in surgical biopsy specimens of breast carcinomas, was used in an improved alkaline phosphatase anti-alkaline phosphatase (APAAP) technique for in-situ receptor detection. This method was compared with a standard radioligand binding biochemical assay in assessing the ER status of 48 human breast carcinomas. The monoclonal antibody allowed detection of ER in the presence of estrogens or anti-estrogens and avoided problems of nonspecific intrinsic reactions, due to endogenous steroid hormones. The immunohistologic results (positive and negative) correlated well with the radioligand binding results (rs = 0.760; P less than 0.001) and allowed the additional assessment of cell morphology and tumor heterogeneity in the cryostat sections. The APAAP method provides diagnostic rapidity (approximately 4 hours) combined with sensitivity as compared with the radioligand assay (an approximately 2-day assay). The vivid red color associated with APAAP staining also was a distinct advantage compared with the brown reaction product of ER-ICA in interpreting stained sections. The results of this study indicate that the APAAP technique is an improved method for detection of ER and is applicable in routine hospital practice, thereby contributing to the immediate clinical management of patients with breast tumors.