To determine whether endogenously expressed ERα was phosphorylated at sites S282, S294, and S559, ERα was first immunoprecipitated using phosphospecific antibodies directed towards p-S282, p-S294, and p-S559, followed by Western blotting with antibody to total ERα in a panel of ERα+ breast cancer cell lines. Here, ESR1 is linked to breast carcinoma.