Detection of Superfund chemicals is a first step towards eliminating them from the environment or studying their effect on biological processes. To that end, we propose to develop molecular tools for the highly sensitive and direct detection of Superfund chemicals such as PCBs. These molecular tools will include whole cell- and protein-based optical sensing systems. For that, we propose to utilize naturally available operons induced in the presence PCBs and the catabolic enzymes involved in their degradation. We plan to modify these operons by incorporating the gene for the reporter, which will allow us to monitor their induction in the presence of PCBs. Catabolic enzymes involved in the degradation of PCBs, in particular the enzyme biphenyl dioxygenase (BphAt), will be isolated and labeled with signal transduction moieties for the analysis of these compounds. In addition, to improve the selectivity of these catabolic enzymes towards PCB congeners, mutagenesis studies aimed at modifying the active site of the enzyme are proposed. In order to make these biosensing systems amenable to field studies and high throughput sample analysis, we propose to incorporate them into a microfluidics platform. This miniaturized analytical platform should find applications in the on-site monitoring of Superfund chemicals. Specifically, we plan to adapt our biosensing systems for PCBs to a compact disc centrifugal microfluidics platform. The advantages of our proposed biosensing systems include sensitivity, portability, and the feasibility of analysis of PCBs in different matrices, such as soil, water, and biological samples.
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