期刊
BIOTECHNIQUES
卷 71, 期 2, 页码 403-415出版社
FUTURE SCI LTD
DOI: 10.2144/btn-2021-0017
关键词
androgen; autobioluminescence; bioreporter; endocrine disrupting chemical (EDC); HEK293; lux
An autobioluminescent HEK293(ARE/Gal4-Lux) assay was developed to screen compounds for their ability to induce androgen receptor (AR)-mediated transcriptional activation. When validated against a group of 40 recommended chemicals, the assay achieved an overall 87.5% accuracy in qualitatively classifying positive and negative AR agonists. This assay was demonstrated as a suitable tool for Tier 1 AR agonist screening, offering higher-throughput screening with reduced testing costs and maintained detection accuracy by eliminating exogenous substrate.
METHOD SUMMARY A human optimized version of the bacterial luciferase gene cassette was developed such that bioluminescence is controlled by exposure to androgen-disruptor chemicals. This cassette, along with the androgen receptor gene, was co-transfected into an HEK293 human cell host that naturally lacks hormone receptors. The resulting reporter cell line was used to screen compounds for androgenic activity in a low cost, high throughput format. Due to the public health concerns of endocrine-disrupting chemicals, there is an increasing demand to develop improved high-throughput detection assays for enhanced exposure control and risk assessment. A substrate-free, autobioluminescent HEK293(ARE/Gal4-Lux) assay was developed to screen compounds for their ability to induce androgen receptor (AR)-mediated transcriptional activation. The assay was validated against a group of 40 recommended chemicals and achieved an overall 87.5% accuracy in qualitatively classifying positive and negative AR agonists. The HEK293(ARE/Gal4-Lux) assay was demonstrated as a suitable tool for Tier 1 AR agonist screening. By eliminating exogenous substrate, this assay provided a significant advantage over traditional reporter assays by enabling higher-throughput screening with reduced testing costs while maintaining detection accuracy.
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