Production And Characterization Of An Estrogen Receptor Beta Subtype-Specific Mouse Monoclonal Antibody

An important step in differentiating the unique physiological roles of the alpha and beta forms of estrogen receptor is to determine the precise expression pattern of each of these receptors. We report the generation and characterization of a murine IgG1 monoclonal antibody (MAb), ER15.64A that is ER beta subtype-specific and capable of recognizing full-length human ER beta as well as all of its known protein isoforms. ER15.64A, raised against a ER beta peptide (aa2-18)-keyhole limpet hemocyanine conjugate, reacted to the immunizing peptide and the full-length E. coli expressed ER beta in ELISA and BIAcore assays. It also immunostained nuclei of Sf9 insect cells that were infected with an ER beta -baculovirus. In Western analysis, ER15.64A recognized ER beta1 and ER beta2 proteins from a reticulocyte in vitro transcription/translation preparation. This antibody did not crossreact with recombinant ER alpha in ELISA, BIAcore, immunocytochemistry, or Western blot analysis. The specificity of ER15.64A should make this antibody a useful tool for monitoring expression of ER beta and its isoforms at the protein level and should aid in distinguishing the pattern of ER beta receptor expression from that of ER alpha.