Abstract
A rapid and reproducible fiber-optic immunosensor for Escherichia coli O157:H7 (E. coli O157:H7) was described. The biosensor consisted of a flow cell, an optical fiber with a thin Ni layer, and a PC linked fluorometer. First, the samples with E. coli O157:H7 were incubated with magnetic beads coated with anti-E. coli O157:H7 antibodies and anti-E. coli O157:H7 antibodies labeled cyanine 5 (Cy5) to make sandwich complexes. Then the Cy5-(E. coli O157:H7)-beads were injected into a flow cell and pulled to the magnetized Ni layer on the optical fiber set in the flow cell. An excitation light (λ = 635 nm) was used to illuminate the optical fiber, and the Cy5 florescent molecules facing the optical fiber were exposed to an evanescent wave from the optical fiber. The 670 nm fluorescent light was measured using a photodiode. Finally, the magnetic intensity of the Ni layer was removed and the Cy5-E. coli O157:H7-beads were washed out for the next immunoassay. E. coli O157:H7, diluted with phosphate buffer (PB), was measured from 1 × 105 to 1 × 107 cells/mL. The total time required for an assay was less than 15 min (except for the pretreatment process) and repeating immunoassay on one optical fiber was made possible.
Highlights
Bacteria detection is important in the food industry, clinical assay and environmental assessment.Conventional approaches for measuring bacteria cell counts employ a selective culture, serological and biochemical characterization
A commercially available polyclonal antibody was used as a capture antibody (No 01-95-90, Anti-E. coli O157:H7, Kierkegaard and Perry Laboratories Inc., Gaithersburg, MD, USA)
The antibody-based fiber-optic immunosensor waspromising a method applications developed to meet the need for separation, magnetic-beads with bioactive molecules immobilized on the surface are attracting interest
Summary
Conventional approaches for measuring bacteria cell counts employ a selective culture, serological and biochemical characterization. The achievement of sensitive and selective bacteria detection typically needs prolonged assay times Conventional direct assays are RIA (radio-immunoassay) and ELISA (enzyme-linked immunosorbent assay). In both approaches (RIA and ELISA), an enzyme or radioisotope is covalently bound to the antibody or antigen. The antibody binding is measured as fluorescence by the product of an enzyme reaction in ELISA or as the amount of radioactivity by the coated well in RIA. Even though those approaches reduce the assay time compared with conventional culture
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