title: Comprehensive assessment of estrogen receptor beta antibodies in cancer cell line models and tissue reveals critical limitations in reagent specificity.
abstract: Estrogen Receptor-β (ERβ) has been implicated in many cancers. In prostate and breast cancer its function is controversial, but genetic studies implicate a role in cancer progression. Much of the confusion around ERβ stems from antibodies that are inadequately validated, yet have become standard tools for deciphering its role. Using an ERβ-inducible cell system we assessed commonly utilized ERβ antibodies and show that one of the most commonly used antibodies, NCL-ER-BETA, is non-specific for ERβ. Other antibodies have limited ERβ specificity or are only specific in one experimental modality. ERβ is commonly studied in MCF-7 (breast) and LNCaP (prostate) cancer cell lines, but we found no ERβ expression in either, using validated antibodies and independent mass spectrometry-based approaches. Our findings question conclusions made about ERβ using the NCL-ER-BETA antibody, or LNCaP and MCF-7 cell lines. We describe robust reagents, which detect ERβ across multiple experimental approaches and in clinical samples.
authors.affiliation: Cancer Research UK Cambridge Institute, University of Cambridge, Robinson Way, Cambridge, CB2 ORE, UK; Academic Urology Group, Department of Surgery, University of Cambridge, Cambridge, CB2 0QQ, UK; Department of Urology, Addenbrooke's Hospital, Cambridge University Hospitals NHS Foundation Trust, Hills Road, Cambridge, CB2 0QQ, UK.
authors.forename: Adam W
authors.initials: AW
authors.lastname: Nelson
authors.affiliation: Cancer Research UK Cambridge Institute, University of Cambridge, Robinson Way, Cambridge, CB2 ORE, UK.
authors.forename: Arnoud J
authors.initials: AJ
authors.lastname: Groen
authors.affiliation: Cancer Research UK Cambridge Institute, University of Cambridge, Robinson Way, Cambridge, CB2 ORE, UK.
authors.forename: Jodi L
authors.initials: JL
authors.lastname: Miller
authors.affiliation: Department of Histopathology, Cambridge University Hospitals NHS Foundation Trust, Hills Road, Cambridge, CB2 0QQ, UK.
authors.forename: Anne Y
authors.initials: AY
authors.lastname: Warren
authors.affiliation: Cancer Research UK Cambridge Institute, University of Cambridge, Robinson Way, Cambridge, CB2 ORE, UK.
authors.forename: Kelly A
authors.initials: KA
authors.lastname: Holmes
authors.affiliation: Dame Roma Mitchell Cancer Research Laboratories, Hanson Institute Building, School of Medicine, Faculty of Health Sciences, The University of Adelaide, SA 5005, Australia.
authors.forename: Gerard A
authors.initials: GA
authors.lastname: Tarulli
authors.affiliation: Dame Roma Mitchell Cancer Research Laboratories, Hanson Institute Building, School of Medicine, Faculty of Health Sciences, The University of Adelaide, SA 5005, Australia.
authors.forename: Wayne D
authors.initials: WD
authors.lastname: Tilley
authors.affiliation: Departments of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
authors.forename: Benita S
authors.initials: BS
authors.lastname: Katzenellenbogen
authors.affiliation: Department of Biochemistry and Molecular Biology, Mayo Clinic, 200 1st Street SW, Rochester, MN 55905 USA.
authors.forename: John R
authors.initials: JR
authors.lastname: Hawse
authors.affiliation: Academic Urology Group, Department of Surgery, University of Cambridge, Cambridge, CB2 0QQ, UK; Department of Urology, Addenbrooke's Hospital, Cambridge University Hospitals NHS Foundation Trust, Hills Road, Cambridge, CB2 0QQ, UK.
authors.forename: Vincent J
authors.initials: VJ
authors.lastname: Gnanapragasam
authors.affiliation: Cancer Research UK Cambridge Institute, University of Cambridge, Robinson Way, Cambridge, CB2 ORE, UK. Electronic address: Jason.carroll@cruk.cam.ac.uk.
authors.forename: Jason S
authors.initials: JS
authors.lastname: Carroll
mesh_terms: D000912:Antibodies, Neoplasm / Q000494:pharmacology*; D001940:Breast / Q000187:drug effects / Q000378:metabolism; D045744:Cell Line, Tumor; D004318:Doxycycline / Q000494:pharmacology; D047629:Estrogen Receptor beta / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D007202:Indicators and Reagents; D008297:Male; D010455:Peptides; D011467:Prostate / Q000187:drug effects / Q000378:metabolism; D015203:Reproducibility of Results
chemical_list: D000912:Antibodies, Neoplasm; D047629:Estrogen Receptor beta; D007202:Indicators and Reagents; D010455:Peptides; D004318:Doxycycline
keywords: Antibody; Breast; Cancer; Estrogen receptor beta; Prostate