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Detection Of Escherichia Coli O157:H7 In 10- And 25-Gram Ground Beef Samples With An Evanescent-Wave Biosensor With Silica And Polystyrene Waveguides

DANIEL R. DeMARCO, DANIEL V. LIM

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A portable evanescent-wave fiber-optic biosensor was used to detect Escherichia coli O157:H7 in seeded 10- and 25-g ground beef samples. The biosensor works by launching light from a 635-nm laser diode into specially designed optical fiber probes, generating an evanescent field that extends approximately 1,000 nm from the fiber surface. Fluorescent molecules within the evanescent field are excited, and a portion of their emission recouples into the fiber probe. The return path emission is transported by an optical fiber to a photodiode within the biosensor that detects and quantifies the fluorescent signal. A sandwich immunoassay was performed on the fiber probes with cyanine 5 dye–labeled polyclonal anti–E. coli O157:H7 antibodies for generation of the specific fluorescent signal. Biotin-streptavidin interactions were used to attach polyclonal antiE. coli O157:H7 antibodies to the surface of the fiber probe. A centrifugation method was developed to obtain samples suitable for biosensor analysis from 10- and 25-g ground beef samples. The assay was shown to be sensitive and repeatable. One hundred percent correct identification of positive samples was demonstrated at 9.0 × 103 CFU/g for 25-g ground beef samples with silica waveguides and at 5.2 × 102 CFU/g for 10-g ground beef samples with polystyrene waveguides. The reaction was highly specific. No false positives were observed for 10-g ground beef samples not spiked with the pathogen. In addition, when samples were spiked with high concentrations of a variety of non–E. coli O157:H7 organisms, no false positives were observed. The method was rapid, with results being obtained within 25 min of sample processing.