Abstract
Fiber optic biosensor has a great potential to meet the need for rapid, sensitive,and real-time microbial detection systems. We developed an antibody-based fiber-opticbiosensor to rapidly detect low levels of Escherichia coli O157:H7 cells in ground beef. Theprinciple of the sensor is a sandwich immunoassay using an antibody which is specific forE. coli O157:H7. A polyclonal antibody was first immobilized on polystyrene fiberwaveguides through a biotin-streptavidin reaction that served as a capture antibody. AnAlexa Fluor 647 dye-labeled antibody to E. coli O157:H7 was used to detect cells andgenerate a specific fluorescent signal, which was acquired by launching a 635 nm laser-lightfrom an Analyte-2000. Fluorescent molecules within several hundred nanometers of thefiber were excited by an evanescent wave, and a portion of the emission light fromfluorescent dye transmitted by the fiber and collected by a photodetector at wavelengths of670 to 710 nm quantitatively. This immunosensor was specific for E. coli O157:H7compared with multiple other foodborne bacteria. In addition, the biosensor was able todetect as low as 103 CFU/ml pure cultured E. coli O157:H7 cells grown in culture broth.Artificially inoculated E. coli O157:H7 at concentration of 1 CFU/ml in ground beef couldbe detected by this method after only 4 hours of enrichment.
Highlights
Escherichia coli O157:H7 is a Gram-negative rod-shaped Shiga toxin(s) producing bacterium
Determination of optimal concentrations of capture and detection antibodies In order to determine the optimum concentrations of detection antibody required for the assay development, we used a constant numbers of E. coli O157:H7 (108 CFU/ml) cells and a constant amount of capture antibody (100 μg/ml) with variable concentrations of Alexa-Fluor- labeled detection antibody
Detection limit for fiber optic biosensor was determined by testing with different concentrations of E. coli O157:H7 cells
Summary
Escherichia coli O157:H7 is a Gram-negative rod-shaped Shiga toxin(s) producing bacterium. In this experiment, using the Analyte 2000 (Research International), we employed a modified antibody immobilization procedure and a more sensitive fluorescent dye Detection limit for fiber optic biosensor was determined by testing with different concentrations of E. coli O157:H7 cells.
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