Development and assessment of a novel Arxula adeninivorans androgen screen (A-YAS) assay and its application in analysis of cattle urine

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Development and assessment of a novel Arxula adeninivorans androgen screen (A-YAS) assay and its application in analysis of cattle urine

Gerlach T, Knaust J, Kaiser C, Körner M, Hettwer K, Uhlig S, Simon K, Baronian K, Kunze G

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 platformXplor®2 was used to select stablemitotic 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 themost sensitive yeast-based androgen screenwith 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.

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