Assessment of estrogenic or anti-estrogenic activity of chemicals by the human stably transfected estrogen sensitive MELN cell line
Commonly used acronym: hER assay with MELN cell line
Scope of the method
- Animal health
- Human health
- Translational - Applied Research
- In vitro - Ex vivo
- Animal derived cells / tissues / organs
- hER transactivation assay
- MELN cell line
- cytotoxicity assessment
- luciferase activity
- endocrine disruption of chemicals
- estrogen activity
MELN cells [provided by INSERM, Montpellier, FR; Balaguer et al. (1999)] are oestrogen-sensitive human breast cancer cells (MCF-7) stably transfected with the oestrogen-responsive gene (ERE-βGlo-Luc-SVNeo) carried by integrated plasmids. A standard set-up has been developed to expose MELN cells and measure ER-transactivation for xeno-oestrogenic compounds. Cells are exposed for 19-20 hours to dilutions of test compounds (dissolved at 0.1% DMSO) in comparison to a dilution series of the positive control, 17b-estradiol. At the end of the incubation period, the remaining medium is removed for analysis of cell damage using the CytoTox-ONE Homogenous Membrane Integrity Assay (Promega) as described by Berckmans et al. (2007). Next, cells are lysed and luciferase assay is applied. The hERα activation is expressed as percentage of luciferase induction by the vehicle control, and concentration response curves are generated (Graphpad Prism software). The estrogenic potency of chemicals is assessed through comparison of EC50 values relative to the EC50 of 17b-estradiol.
- Laminar flow ;
- Cell incubator ;
- Microplate reader for fluorescence and luminescence detection.
- History of use
- Internally validated
- Published in peer reviewed journal
Pros, cons & Future potential
- High-throughput method for screening of estrogen activity ;
- Agonist or antagonist mode.
References, associated documents and other information
Witters H, Freyberger A, Smits K, Vangenechten C, Lofink W, Weimer M, Bremer S, Ahr PH, Berckmans P. (2010). The assessment of estrogenic or anti-estrogenic activity of chemicals by the human stably transfected estrogen sensitive MELN cell line: Results of test performance and transferability. Reprod Toxicol. 30 (1), 60-72.