Validation of a new Custom Polyclonal Progesterone Receptor Antibody for Immunohistochemistry in the Female Mouse Brain

Abstract Immunohistochemical visualization of progesterone receptor (PR)–expressing cells in the brain is a powerful technique to investigate the role of progesterone in the neuroendocrine regulation of fertility. A major obstacle to the immunohistochemical visualization of progesterone-sensitive cells in the rodent brain has been the discontinuation of the commercially produced A0098 rabbit polyclonal PR antibody by DAKO. To address the unavailability of this widely used PR antibody, we optimized and evaluated 4 alternative commercial PR antibodies and found that each lacked the specificity and/or sensitivity to immunohistochemically label PR-expressing cells in paraformaldehyde-fixed female mouse brain sections. As a result, we developed and validated a new custom RC269 PR antibody, directed against the same 533-547 amino acid sequence of the human PR as the discontinued A0098 DAKO PR antibody. Immunohistochemical application of the RC269 PR antibody on paraformaldehyde-fixed mouse brain sections resulted in nuclear PR labeling that was highly distinguishable from background, specific to its antigen, highly regulated by estradiol, matched the known distribution of PR protein expression in the female mouse hypothalamus, and nearly identical to that of the discontinued A0098 DAKO PR antibody. In summary, the RC269 PR antibody is a specific and sensitive antibody to immunohistochemically visualize PR-expressing cells in the mouse brain.