Assessment of multiple different estrogen receptor-beta antibodies for their ability to immunoprecipitate under chromatin immunoprecipitation conditions.

Several different antibodies to total estrogen receptor (ER)beta, ERbeta1 and ERbeta2/cx have been tested and compared for their ability to immunoprecipitate ERbeta specific isoforms under chromatin immunoprecipitation conditions (ChIP). The rabbit polyclonal antibodies AP-ERbeta1 and AP-ERbeta2/cx, specific for ERbeta1 and ERbeta2/cx isoforms, respectively, were the most efficient for ChIP. The monoclonal antibody MCA1974/PPG5/10 was also able to ChIP ERbeta1, but less efficiently than AP-ERbeta1. All other antibodies tested were not suitable for ChIP analyses although most antibodies tested immunoprecipitated the appropriate ERbeta isoforms under standard conditions. To identify antibodies that can also be used to verify in-vivo expression profiles, a comparison of the antibodies to detect ERbeta isoforms by western blotting and immunohistochemistry was also undertaken. Under the tissue processing and autostaining conditions used at the Manitoba Breast Tumor Bank 385P/GC17, MCA1974/PPG5/10, Ab288/14C8 and MCA2279S/57/3 were found to be the best for IHC of ERbeta isoforms in human breast tissue biopsy sections, while Ab14021, AP-ERbeta1 and AP-ERbeta2/cx were best for western blot detection of ERbeta isoforms.