Details zur Publikation

Referenztyp Zeitschriften
DOI / URL Link
Titel (primär) Downscaling procedures reduce chemical use in androgen receptor reporter gene assay
Autor Di Paolo, C.; Kirchner, K.; Balk, F.G.P.; Muschket, M.; Brack, W.; Hollert, H.; Seiler, T.-B.;
Journal / Serie Science of the Total Environment
Erscheinungsjahr 2016
Department WANA;
Band/Volume 571
Sprache englisch;
POF III (gesamt) T32;
Keywords Downscaling; Miniaturization; Low-volume; Androgen receptor binders; Reporter gene bioassay; (Anti)androgens
UFZ Querschnittsthemen RU2;
Abstract Bioactivity screening studies often face sample amount limitation with respect to the need for reliable, reproducible and quantitative results. Therefore approaches that minimize sample use are needed. Low-volume exposure and chemical dilution procedures were applied in an androgen receptor reporter gene human cell line assay to evaluate environmental contaminants and androgen receptor modulators, which were the agonist 5α-dihydrotestosterone (DHT); and the antagonists flutamide, bisphenol A, 1-hydroxypyrene and triclosan. Cells were exposed in around 1/3 of the medium volume recommended by the protocol (70 μL/well). Further, chemical losses during pipetting steps were minimized by applying a low-volume method for compound dilution in medium (250 μL for triplicate wells) inside microvolume glass inserts. Simultaneously, compounds were evaluated following conventional procedures (200 μL/well, dilution in 24-well plates) for comparison of results. Low-volume exposure tests produced DHT EC50 (3.4–3.7 × 10− 10 M) and flutamide IC50 (2.2–3.3 × 10− 7 M) values very similar to those from regular assays (3.1–4.2 × 10− 10 and 2.1–3.3 × 10− 7 M respectively) and previous studies. Also, results were within assay acceptance criteria, supporting the relevance of the downscaling setup for agonistic and antagonistic tests. The low-volume exposure was also successful in determining IC50 values for 1-hydroxypyrene (2.1–2.8 × 10− 6 M), bisphenol A (2.6–3.3 × 10− 6 M), and triclosan (1.2–1.9 × 10− 6 M) in agreement with values obtained through high-volume exposure (2.3–2.8, 2.5–3.4 and 1.0–1.3 × 10− 6 M respectively). Finally, experiments following both low-volume dosing and exposure produced flutamide and triclosan IC50 values similar to those from regular tests. The low-volume experimental procedures provide a simple and effective solution for studies that need to minimize bioassay sample use while maintaining method reliability. The downscaling methods can be applied for the evaluation of samples, fractions or chemicals which require minimal losses during the steps of pipetting, transference to medium and exposure in bioassays.
ID 17926
dauerhafte UFZ-Verlinkung http://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=17926
Di Paolo, C., Kirchner, K., Balk, F.G.P., Muschket, M., Brack, W., Hollert, H., Seiler, T.-B. (2016):
Downscaling procedures reduce chemical use in androgen receptor reporter gene assay
Sci. Total Environ. 571 , 826 - 833