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

The Method relates to
  • Animal health
  • Environment
  • Human health
The Method is situated in
  • Translational - Applied Research
Type of method
  • In vitro - Ex vivo
This method makes use of
  • Animal derived cells / tissues / organs
Species from which cells/tissues/organs are derived
Human
Type of cells/tissues/organs
Breast cancer cells (transfected)

Description

Method keywords
  • hER transactivation assay
  • MELN cell line
  • cytotoxicity assessment
  • luciferase activity
Scientific area keywords
  • endocrine disruption of chemicals
  • estrogen activity
Method description

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.

Lab equipment
  • Laminar flow ;
  • Cell incubator ;
  • Microplate reader for fluorescence and luminescence detection.
Method status
  • History of use
  • Internally validated
  • Published in peer reviewed journal

Pros, cons & Future potential

Advantages
  • High-throughput method for screening of estrogen activity ;
  • Agonist or antagonist mode.

References, associated documents and other information

References

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.

Associated documents
Witters et al Repr Tox 30-2010, 60-72.pdf

Organisations

VITO
Health
Belgium
Flemish Region