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Applied and Environmental Microbiology, October 2004, p. 6138-6146, Vol. 70, No. 10
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.10.6138-6146.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Detection of Low Levels of Listeria monocytogenes Cells by Using a Fiber-Optic Immunosensor

Tao Geng,¹ Mark T. Morgan,² and Arun K. Bhunia^1*

Molecular Food Microbiology Laboratory,¹ Sensors and Controls Laboratory, Department of Food Science, Purdue University, West Lafayette, Indiana²

Received 26 November 2003/ Accepted 22 June 2004

Biosensor technology has a great potential to meet the need for sensitive and nearly real-time microbial detection from foods. An antibody-based fiber-optic biosensor to detect low levels of Listeria monocytogenes cells following an enrichment step was developed. The principle of the sensor is a sandwich immunoassay where a rabbit polyclonal antibody was first immobilized on polystyrene fiber waveguides through a biotin-streptavidin reaction to capture Listeria cells on the fiber. Capture of cells on the fibers was confirmed by scanning electron microscopy. A cyanine 5-labeled murine monoclonal antibody, C11E9, was used to generate a specific fluorescent signal, which was acquired by launching a 635-nm laser light from an Analyte 2000 and collected by a photodetector at 670 to 710 nm. This immunosensor was specific for L. monocytogenes and showed a significantly higher signal strength than for other Listeria species or other microorganisms, including Escherichia coli, Enterococcus faecalis, Salmonella enterica, Lactobacillus plantarum, Carnobacterium gallinarum, Hafnia alvei, Corynebacterium glutamicum, Enterobacter aerogenes, Pseudomonas aeruginosa, and Serratia marcescens, in pure or in mixed-culture setup. Fiber-optic results could be obtained within 2.5 h of sampling. The sensitivity threshold was about 4.3 x 10³ CFU/ml for a pure culture of L. monocytogenes grown at 37°C. When L. monocytogenes was mixed with lactic acid bacteria or grown at 10°C with 3.5% NaCl, the detection threshold was 4.1 x 10⁴ or 2.8 x 10⁷ CFU/ml, respectively. In less than 24 h, this method could detect L. monocytogenes in hot dog or bologna naturally contaminated or artificially inoculated with 10 to 1,000 CFU/g after enrichment in buffered Listeria enrichment broth.

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* Corresponding author. Mailing address: Molecular Food Microbiology Laboratory, Department of Food Science, Purdue University, 745 Agriculture Mall Dr., West Lafayette, IN 47906-2009. Phone: (765) 494-5443. Fax: (765) 494-7953. E-mail: bhunia{at}purdue.edu.

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Applied and Environmental Microbiology, October 2004, p. 6138-6146, Vol. 70, No. 10
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.10.6138-6146.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

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