The parent cell line was transfected with pGL2.TATA.Inr.luc.neo which contains three estrogen responsive elements upstream of a luc reporter gene.xa0

The cells were selected for responsiveness to 17-beta-estradiol. They can be used to screen chemicals for estrogenic or anti-estrogenic activity.

1. Remove and discard culture medium.
2. Briefly rinse the cell layer with either Hanks Balanced Salt Solution (HBSS) or 0.25% (w/v) Trypsin - 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.
3. Add 1.0 to 2.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
   Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
4. Add 10 to 15 mL of complete growth medium and aspirate cells by gently pipetting.
5. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 2 x 10⁴ to 4 x 10⁴ viable cells/cm² is recommended.
6. Incubate cultures at 37C. We recommend that you maintain cultures at a cell concentration between 3 x 10⁴ and 2 x 10⁵ cells/cm².

Wilson VS, et al. Development and characterization of a cell line that stably expresses an estrogen-responsive luciferase reporter for the detection of estrogen receptor agonist and antagonists. Toxicol. Sci. 81: 69-77, 2004. PubMed: 15166400