Pre-validation of choriogenin H transgenic medaka eleutheroembryos as a quantitative estrogenic activity test method

Xueping Chen, Shuk Han Cheng, Masato Kinoshita, Peter A. de Witte, Jianjun Liu, David Hinton, Thomas Braunbeck, Ian Cotgreave, Daniel Schlenk, Zhiyuan Gong, Hani El-Nezami, Kin Chung Ho, Kwok Fai Chan, Shisan Xu, Pui Ying Yiu, Huan Zhang, Desheng Wu, Yu Suen Chan, Annelii Ny, Jan Maes

Research output: Journal article publicationJournal articleAcademic researchpeer-review

3 Citations (Scopus)


The choriogenin H – EGFP transgenic medaka (Oryzias melastigma) has been used to test estrogenic substances and quantify estrogenic activity into 17β-estradiol (E2) equivalency (EEQ). The method uses 8 eleutheroembryos in 2 ml solution per well and 3 wells per treatment in 24-well plates at 26 ± 1 °C for 24 ± 2 h, with subsequent measurements of induced GFP signal intensity. EEQ measurements are calculated using a E2 probit regression model with a coefficient of determination (R2) > 0.90. The selectivity was confirmed evaluating 27 known estrogenic and 5 known non-estrogenic compounds. Limit of quantitation (LOQ), recovery rate and bias were calculated to be 1 ng/ml EEQ, 104% and 4% respectively. Robustness analysis revealed exposure temperature is a sensitive parameter that should be kept at 26 ± 1 °C. The repeatability of intra- and inter-laboratories achieved CV < 30% for most tested food and cosmetics samples. The lot-lot stability was confirmed by the stable EEQ qualitative control (QC, 1 ng/mL E2) and calibration curve results. The stability of standard reagents, samples and sample extracts was also investigated. These data demonstrated this method to be an accurate indicator of estrogenic activity for both chemicals and extracts.

Original languageEnglish
Article number114311
JournalAnalytical Biochemistry
Publication statusPublished - 15 Sept 2021
Externally publishedYes


  • Bioanalytical method
  • EEQ test
  • Estrogen equivalency
  • Estrogenic activity
  • Semi-quantitative
  • Transgenic medaka

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology


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