Ha Thi Minh Pham, Martin Giersberg, Steffen Uhlig, Gerold Hanke, Kirsten Simon, Kirstin Kunath, Keith Baronian, Gotthard Kunze
The EstraMonitor has been developed to provide a device suitable for monitoring estrogens in sewage treatment plants, i.e. provide rapid detection in semi-online operation. This monitor is based on recombinant Arxula adeninivorans yeast cells as the biocomponent. The cells were engineered to co-express the human estrogen receptor ˛ (hER˛) gene and the inducible phytase (phyK, derived from Klebsiella sp. ASR1) reporter gene under the control of a promoter with estrogen response elements (EREs). The monitor comprises two measuring chambers containing immobilized transgenic yeast cells – a measuring strain with receptor and reporter gene cassettes and a control strain with only the reporter gene cassette. To measure the estrogenic activity, the measuring and control strains are incubated with the samples. In the presence of estrogenic substances, such as 17 estradiol (E2), the phyK gene is expressed and recombinant phytase is secreted into the sample. Any signal in the control strain is due to background activity and is used to correct the measuring strain signal. Phytase dephosphorylates p-aminophenyl phosphate (pAPP) and the level of phytase was quantified by electrochemically reducing the intermediate product, p-aminophenol (pAP). The electrons are detected as current and are proportional to the level of phytase activity. Since E2 concentration is correlated with phytase activity, the estrogenic activity can be calculated. The EstraMonitor can detect the total estrogenic activity in about 4 h with a measurement range between 5 and 100 ng L−1, and limit of detection of 5.3 ng L−1.
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