title: Freeze-drying as suitable method to achieve ready-to-use yeast biosensors for androgenic and estrogenic compounds.
abstract: Recombinant yeast assays (RYAs) have been proved to be a suitable tool for the fast screening of compounds with endocrine disrupting activities. However, ready-to-use versions more accessible to less equipped laboratories and field studies are scarce and far from optimal throughputs. Here, we have applied freeze-drying technology to optimize RYA for the fast assessment of environmental compounds with estrogenic and androgenic potencies. The effects of different cryoprotectants, initial optical density and long-term storage were evaluated. The study included detailed characterization of sensitivity, robustness and reproducibility of the new ready-to-use versions, as well as comparison with the standard assays. Freeze-dried RYAs showed similar dose-responses curves to their homolog standard assays, with Lowest Observed Effect Concentration (LOEC) and Median effective Concentration (EC50) of 1 nM and 7.5 nM for testosterone, and 0.05 nM and 0.5 nM for 17β-estradiol, respectively. Freeze-dried cells stored at 4 °C retained maximum sensitivity up to 2 months, while cells stored at -18 °C showed no decrease in sensitivity throughout the study (10 months). This ready-to-use RYA is easily accessible and may be potentially used for on-site applications.
authors.affiliation: Masaryk University, Faculty of Science, RECETOX, Kamenice 5/753, Brno CZ62500, Czech Republic.
authors.forename: Sergio
authors.initials: S
authors.lastname: Jarque
authors.affiliation: Masaryk University, Faculty of Science, RECETOX, Kamenice 5/753, Brno CZ62500, Czech Republic.
authors.forename: Michal
authors.initials: M
authors.lastname: Bittner
authors.affiliation: Masaryk University, Faculty of Science, RECETOX, Kamenice 5/753, Brno CZ62500, Czech Republic. Electronic address: hilscherova@recetox.muni.cz.
authors.forename: Klára
authors.initials: K
authors.lastname: Hilscherová
mesh_terms: D000728:Androgens / Q000494:pharmacology*; D001681:Biological Assay / Q000379:methods; D015374:Biosensing Techniques / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D052244:Endocrine Disruptors / Q000494:pharmacology*; D004784:Environmental Monitoring / Q000379:methods*; D065171:Estrogen Receptor Antagonists; D047628:Estrogen Receptor alpha / Q000235:genetics; D004967:Estrogens / Q000494:pharmacology*; D005612:Freeze Drying / Q000379:methods*; D050296:Microbial Viability / Q000187:drug effects; D011944:Receptors, Androgen / Q000235:genetics; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics
chemical_list: C410798:AR protein, human; D000728:Androgens; D003451:Cryoprotective Agents; C506487:ESR1 protein, human; D052244:Endocrine Disruptors; D065171:Estrogen Receptor Antagonists; D047628:Estrogen Receptor alpha; D004967:Estrogens; D011944:Receptors, Androgen
keywords: Androgen; Estrogen; Freeze-drying; Long-term; Ready-to-use; Recombinant yeast assay