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Abstract

Water is a precious good which in good quality we need essentially to survive. In this work a novel method for the detection of bioactive pollutants in aqueous media will be presented. It is based on a sensor system, which uses mammalian cells, RLC-18 (rat liver cells) or MCF-7 (breast cancer cell line) as the detection layer for harmful substances. With these mammalian cells as the sensing layer a metabolically active sensor interface will become available reflecting the physiology of living organisms. In this work the cell layer is cultured on a multi-electrode silicon chip and the response of the cells to toxic substances is monitored by recording morphological changes, oxygen consumption, as well as the acidification properties of the cells.

Until now there is no easy-to-use system that can detect the overall toxicity of a water sample. Analytical chemistry can be used to detect chemical substances at very low levels, down to few picograms. However, only substances known to be harmful will be analyzed in dedicated protocols. New, unknown chemicals or derivatives can be present unnoticed. For this work, four known organic pollutants were chosen as model substances to test the sensitivity and reliability of the developed system: 2-Aminoanthracene (2-AA), a heterocyclic amine used in dyes, drugs, inks and plastics; chlorpyrifos, an organophosphate insecticide; 3,3‘,5,5‘-Tetrabromobisphenol A (TBBPA), a flame retardant; and 4-methylbenzylidene camphor (4-MBC), an ultraviolet filter. RLC-18 cells were exposed to these chemicals in single and combinatorial measurements. These measurements revealed, which physiological parameters were targeted, and at which time point during treatment the toxic effect occurred. Cotreatment with various combinations showed how the chemicals interacted with each other either by reinforcing their toxicity or mitigating it. Moreover, also cell promoting effects of certain substances could be detected using this system. Proliferating effects could indicate hormonal activities of the pollution. In this context, the focus was on compounds mimicking the effect of estrogen, which can be detected using MCF-7 cells as the detection layer since this cell line expresses the estrogen receptor and shows hormone dependent growth.

Due to the different options for the interaction of pollutions, a more dedicated analysis was required. In order to predict the presence of a pollutant in a sample, even in the presence of another compound that could suppress the expected physiological effect, an algorithm using functional data analysis was designed and an analytical protocol including analysis software was developed.

A major problem was the maintenance of the cell layer, to make it suitable for application in the field. For this a gelatinous medium was developed to protect the cells against impacts, dehydration and temperature variations during storage and transportation. This gel medium permitted the storage of cell loaded chips at 4°C for up to one week. Moreover, it could be demonstrated that, with some optimization, this system can be used to analyze natural water samples from different sources.

Taken together, the cell-based detection system developed in this work could be used for the detection of pollutants with acute toxicity and other biological activity. With this work, it could be shown that a future cell based sensor system should be an ideal toxicity monitor, to continuously record, in “real time”, the quality of inshore waters as a reliable toxicity meter.