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A Cantilever Biosensor-Based Assay for Toxin-Producing Cyanobacteria Microcystis aeruginosa using 16S rRNA

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Department of Chemical and Biological Engineering, Drexel University, Philadelphia, Pennsylvania 19104, United States
*(R.M.) Phone: (215) 895-2236; fax: (215) 895-5837; e-mail: [email protected]
Cite this: Environ. Sci. Technol. 2013, 47, 21, 12333–12341
Publication Date (Web):September 26, 2013
https://doi.org/10.1021/es402925k
Copyright © 2013 American Chemical Society

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    Abstract

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    Monitoring of cyanotoxins in source waters is currently done through toxin-targeting assays which suffer from low sensitivity due to poor antibody avidity. We present a biosensor-based method as an alternative for detecting toxin-producing cyanobacteria M. aeruginosa via species-selective region of 16S rRNA at concentrations as low as 50 cells/mL, and over a five-log dynamic range. The cantilever biosensor was immobilized with a 27-base DNA strand that is complementary to the target variable region of 16S rRNA of M. aeruginosa. The cantilever sensor detects mass-changes through shifts in its resonant frequency. Increase in the biosensor’s effective mass, caused by hybridization of target strand with the biosensor-immobilized complementary strand, showed consistent and proportional frequency shift to M. aeruginosa concentrations. The sensor hybridization response was verified in situ by two techniques: (a) presence of duplex DNA structure postdetection via fluorescence measurements, and (b) secondary hybridization of nanogold-labeled DNA strands to the captured 16S rRNA strands. The biosensor-based assay, conducted in a flow format (∼ 0.5 mL/min), is relatively short, and requires a postextraction analysis time of less than two hours. The two-step detection protocol (primary and secondary hybridization) is less prone to false negatives, and the technique as a whole can potentially provide an early warning for toxin presence in source waters.

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