title: Development and assessment of a novel Arxula adeninivorans androgen screen (A-YAS) assay and its application in analysis of cattle urine.
abstract: The novel A-YAS assay for the detection of androgenic activity in liquid samples such as urine has been developed and assessed. The assay is based on transgenic Arxula adeninivorans yeast cells as the bio-component. The cells were engineered to co-express the human androgen receptor (hAR) gene and the inducible phytase reporter gene (phyK, derived from Klebsiella sp. ASR1), under the control of an Arxula derived glucoamylase (GAA) promoter, which had been modified by the insertion of hormone-responsive elements (HREs). The Arxula transformation/expression platform Xplor®2 was used to select stable mitotic resistance marker free transformants and the most suitable cells were characterized for performance as a sensor bio-component. The assay is easy-to-use, fast (6-25 h) and is currently the most sensitive yeast-based androgen screen with an EC50, limit of detection and of quantification values for 5α-dihydrotestosterone (DHT) of 277.1±53.0, 56.5±4.1 and 76.5±6.7 ng L(-1), respectively. Furthermore, the assay allows the determination of androgenic and anti-androgenic activity of various compounds such as naturally occurring androgens and estrogens, pharmaceuticals and biocides. The robustness of the A-YAS assay enables it to be used for analysis of complex samples such as urine. The results of the analysis of a number of cattle urine samples achieved by the A-YAS assay correlate well with GC-MS analysis of the same samples.
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany.
authors.forename: Torsten
authors.initials: T
authors.lastname: Gerlach
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Knaust
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Kaiser
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany.
authors.forename: Martina
authors.initials: M
authors.lastname: Körner
authors.affiliation: Quo Data GmbH, Kaitzer Str. 135, D-01187 Dresden, Germany.
authors.forename: Karina
authors.initials: K
authors.lastname: Hettwer
authors.affiliation: Quo Data GmbH, Kaitzer Str. 135, D-01187 Dresden, Germany.
authors.forename: Steffen
authors.initials: S
authors.lastname: Uhlig
authors.affiliation: New Diagnostics GmbH, Moosstr. 92c, D-85356 Freising, Germany.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Simon
authors.affiliation: School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand.
authors.forename: Keith
authors.initials: K
authors.lastname: Baronian
authors.affiliation: Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, D-06466 Gatersleben, Germany. Electronic address: kunzeg@ipk-gatersleben.de.
authors.forename: Gotthard
authors.initials: G
authors.lastname: Kunze
mesh_terms: D000726:Androgen Antagonists / Q000032:analysis / Q000652:urine*; D000728:Androgens / Q000032:analysis / Q000652:urine*; D000818:Animals; D001681:Biological Assay / Q000379:methods*; D002417:Cattle; D008401:Gas Chromatography-Mass Spectrometry
chemical_list: D000726:Androgen Antagonists; D000728:Androgens
keywords: Androgen receptor; Arxula adeninivorans; Endocrine disruption; Reporter assay; Urine; Yeast assay