A novel biosensor for Escherichia coli O157:H7 based on fluorescein-releasable biolabels.

ABSTRACT

New techniques are required for the rapid and sensitive detection of Escherichia coli O157H7 (E. coli O157H7), a pathogenic bacterium responsible for serious and sometimes life-threatening diseases in humans. In this study, we developed a highly sensitive and efficient biosensor for the quantitative detection of E. coli O157H7 by integrating fluorescein-releasable biolabels with a magnetism-separable probe. Hollow silica nanospheres with a diameter of approximately 350 nm were synthesized, enriched with fluorescein, and surface-protected with macromolecule layers of poly (acrylic acid) and poly (dimethyldiallylammonium chloride). These fluorescein-enriched hollow silica nanospheres were characterized using scanning electron microscopy, transmission electron microscopy, and Fourier transform infrared spectroscopy. They were further functionalized as immune labels of E. coli O157H7 for a sandwich-type immune reaction between this bacterium and magnetic nanoparticles (Fe3O4@SiO2). Next, the E. coli O157H7 cells were captured, magnetically separated, and quantified based on the fluorescence intensity of the fluorescein released from the biolabels of the fluorescein-enriched hollow silica nanospheres. This analytic process can be completed within 75 min, and the biosensor showed a linear relationship ranging from 4 to 4.0 × 10(8)cfu/mL with a detection limit of 3 cfu/mL. These results show that the developed fluorescent sensor has excellent specificity, and good reproducibility and stability. This study used real spiked samples for detection, indicating that this technique has a wide range of potential applications and may be readily adapted for detecting other pathogens.