PRODUCTION OF MONOCLONAL-ANTIBODIES TO HUMAN ESTROGEN-RECEPTOR PROTEIN (ER) USING RECOMBINANT ER (RER)

Two monoclonal antibodies (MAbs), IC5 and ID5, were produced using spl een cells from BALB/c mice immunized with recombinant estrogen-recepto r protein (RER). On immunoblotting, both MAbs reacted with the 67-kDa polypeptide chain obtained by transformation of E. coli and transfecti on of COS cells with plasmid vectors expressing ER. The epitopes of bo th MAbs were in the N-terminal domain (A/B region) of the receptor. In normal human tissues, IC5 and ID5 reacted with cells known to contain large amount of ER such as cells of the mammary gland and the uterus. Staining was localized predominantly in nuclei with little or no cyto plasmic reactivity. IC5 and ID5 were unreactive with tissues usually c onsidered to be negative for ER. The reactions of these 2 MAbs were fu rther tested on different tumor types, using immunohistochemical (IHC) method on frozen sections. In breast cancer, a good correlation was f ound between the results obtained on frozen sections and those using t he conventional radioligand dextran-coated charcoal (DCC) assay. Immun ostaining with IC5 and ID5 MAbs was also assessed on routinely process ed paraffin sections using the antigen-retrieval method. Staining was comparable to that obtained on frozen sections in virtually all the br east carcinomas. Negative reactions were consistently obtained with bo th antibodies on human neoplasms derived from other non-estrogen-depen dent organs. IC5 and ID5 MAbs may thus be of value in routine diagnost ic histopathology for assessment of the estrogen-receptor content in h uman carcinomas. (C) 1993 Wiley-Liss, Inc.