This Test Guideline describes an in vitro assay, which  provides mechanistical information, and can be used for screening and prioritization purposes. The test system utilises the hERalpha-HeLa-9903 cell line derived from a human cervical tumor and stably transfected. This cell line can measure the ability of a test chemical to induce hERalpha-mediated transactivation of luciferase gene expression. The cells are exposed to 7 non-cytotoxic concentrations of the test chemical for 20-24 hours to induce the reporter gene products. Four reference chemicals should be included in each experiment: a strong estrogen (17beta-estradiol), a weak estrogen (17alpha-estradiol), a very weak estrogen (17alpha-methyltestosterone) and a negative control (corticosterone). The activity of the luciferase enzyme is measured in a luminometer. A test chemical is considered to be positive if the maximum response induced is equal to or exceeds 10% of the response of the positive control (1 nM 17alpha-estradiol) in at least two of two or two of three runs. Software to be used with TG 425, 432, 455. Click here. Software not part of the Mutual Acceptance of Data.