Journal
ANALYTICAL CHEMISTRY
Volume 82, Issue 7, Pages 2844-2849Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ac100323k
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Funding
- National Institutes of Health [1R33-CA09924601]
- National Science Foundation [EPS-0346411]
- State of Louisiana Board of Regents Support Fund
- Texas Parks and Wildlife Department
- U.S. Department of Energy
- Texas Sea Grant [NA06OAR4170076]
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Low abundant (<100 cells mL(-1)) E. coli O157:H7 cells were isolated and enriched from environmental water samples using a microfluidic chip. The poly(methylmethacrylate), PMMA, chip contained 8 devices, each equipped with 16 curvilinear high aspect ratio channels that were covalently decorated with polyclonal anti-O157 antibodies (pAb) and could search for rare cells through a pAb mediated process. The chip could process independently 8 different samples or one sample using 8 different parallel inputs to increase volume processing throughput. After cell enrichment, cells were released and enumerated using benchtop real-time quantitative polymerase chain reaction (PCR), targeting genes which effectively discriminated the 0157:H7 serotype from other nonpathogenic bacteria. The recovery of target cells from water samples was determined to be similar to 72%, and the limit-of-detection was found to be 6 colony forming units (cfu) using the slt1 gene as a reporter. We subsequently performed analysis of lake and wastewater samples. The simplicity in manufacturing and ease of operation makes this device attractive for the selection of pathogenic species from a variety of water supplies suspected of containing bacterial pathogens at extremely low frequencies.
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