Background: Oestrogen receptor beta (ERbeta) is highly homologous with the classical ER (known now as ERalpha). The exact role of ERbeta in breast cancer and its contribution in influencing patient response to endocrine therapy remains unclear. The aim of this study was to develop and evaluate a flow cytometric method for the detection of ERbeta in breast cancer cells using the DAKO monoclonal anti-ERbeta 8D5-1 antibody.
Methods: MCF7 cells were used as a positive control and U937 as a negative control for titration of the antibody. Cell lines and tumour samples were fixed with 1% paraformaldehyde and permeabilised with 0.5% saponin prior to flow cytometric analysis.
Results: A ten fold difference in expression of ERbeta within the different breast cell lines studied was found. Confirmation of antibody specificity against ERbeta protein by Western blot analysis detected a single band at approximately 65kDa. ERbeta immunopositive nuclei were identified in MCF7 cells by immunohistochemistry.
Conclusions: DAKO ERbeta 8D5-1 antibody is specific for ERbeta protein and does not cross react with ERalpha protein. Using this antibody, ERbeta can be detected and accurately quantified in cell lines and solid breast tumours by flow cytometry.
Copyright 2001 Wiley-Liss, Inc.