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

Abstract

ABSTRACT 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 × 10 3 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 × 10 4 or 2.8 × 10 7 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.