A Label-Free Impedance Immunosensor Using Screen-Printed Interdigitated Electrodes and Magnetic Nanobeads for the Detection of E. coli O157:H7

Abstract

Escherichia coli O157:H7 is one of the leading bacterial pathogens causing foodborne illness. In this study, an impedance immunosensor based on the use of magnetic nanobeads and screen-printed interdigitated electrodes was developed for the rapid detection of E. coli O157:H7. Magnetic nanobeads coated with anti-E. coli antibody were mixed with an E. coli sample and used to isolate and concentrate the bacterial cells. The sample was suspended in redox probe solution and placed onto a screen-printed interdigitated electrode. A magnetic field was applied to concentrate the cells on the surface of the electrode and the impedance was measured. The impedance immunosensor could detect E. coli O157:H7 at a concentration of 10(4.45) cfu·mL(-1) (~1400 bacterial cells in the applied volume of 25 μL) in less than 1 h without pre-enrichment. A linear relationship between bacteria concentration and impedance value was obtained between 10(4.45) cfu·mL(-1) and 10(7) cfu·mL(-1). Though impedance measurement was carried out in the presence of a redox probe, analysis of the equivalent circuit model showed that the impedance change was primarily due to two elements: Double layer capacitance and resistance due to electrode surface roughness. The magnetic field and impedance were simulated using COMSOL Multiphysics software.

Keywords: E. coli O157:H7; immunosensor; impedance; magnetic nanobeads; rapid detection; screen-printed interdigitated electrode.

Figure 1

Immunomagnetic separation of E. coli O157:H7 from media using the antibody-coated nanobeads and the concentration of bacteria on the electrode surface using a magnetic field.

Figure 2

A typical Bode plot of the measured impedance data of the control and E. coli O157:H7 at a concentration of 10⁷ cfu/mL. (a) Impedance magnitude and (b) Phase angle. The frequency range was 10 Hz to 100 kHz. The amplitude of voltage applied was 50 mV.

Figure 3

Equivalent circuit used for data analysis. The equivalent circuit components were bulk electrolyte (R_sol), electron transfer resistance (R_et), resistance due to surface roughness (R_sur), double layer capacitance (C_dl), capacitance of bacterial cells (C_mem), and a Warburg impedance element (Z_w).

Figure 4

(a) Average impedance change between the control and bacteria measurements at 100 Hz for E. coli O157:H7. Error bars were based on standard deviation of triplicate tests; (b) ESEM photograph of the antibody-coated nanobeads; and (c) ESEM photograph of an E. coli O157:H7 cell attached with antibody-coated nanobeads.

Figure 5

Average impedance change (a) between a control sample and nanobeads captured E. coli O157:H7 with a magnet underneath; (b) pure E. coli O157:H7; (c) between a control sample and nanobeads captured E. coli O157:H7 with no magnet underneath. E. coli O157:H7 concentration was 10⁷ cfu·mL⁻¹. Error bars were based on standard deviation of triplicate tests. LDL line was determined by signal/noise ratio of 3, where noise was defined as the standard deviation of the pure redox probe measurements.

Figure 6

The comparison of impedance signals between the pure culture of E. coli O157:H7 and the ground beef with E. coli O157:H7 at concentrations of 10⁵ and 10⁶ cfu·mL⁻¹ using the immunosensor.

Figure 7

Impedance percent change due to simulated E. coli cells at varying distances from the electrode surface.