Cationic Covalent Organic Polymer Thin Film for Label-free Electrochemical Bacterial Cell Detection

Abstract

Numerous species of bacteria pose a serious threat to human health and cause several million deaths annually. It is therefore essential to have quick, efficient, and easily operable methods of bacterial cell detection. Herein, we synthesize a novel cationic covalent organic polymer (COP) named CATN through the Menshutkin reaction and evaluate its potential as an impedance sensor for Escherichia coli cells. On account of its positive surface charge (ζ-potential = +21.0 mV) and pyridinium moieties, CATN is expected to interact favorably with bacteria that possess a negatively charged cell surface through electrostatic interactions. The interdigitated electrode arrays were coated with CATN using a simple yet non-traditional method of electrophoresis and then used in two-electrode electrochemical impedance spectroscopy (EIS) measurements. The impedance response showed a linear relationship with the increasing concentration of E. coli. The system was sensitive to bacterial concentrations as low as ∼30 CFU mL^-1, which is far below the concentration considered to cause illnesses. The calculated limit of detection was as low as 2 CFU mL^-1. This work is a rare example of a COP used in this type of bacteria sensing and is anticipated to stimulate further interest in the synthesis of organic polymers for EIS-based sensors.

Keywords: E. coli; covalent organic polymers; detection; electrochemical impedance spectroscopy; electrophoresis.

Figure 1

Design and characterization of CATN. (a) Synthetic scheme showing the preparation of CATN; (b) SEM micrograph of CATN showing spherical morphology; (c) cross-polarization magic-angle spinning (CP/MAS) solid-state ¹³C NMR spectrum of CATN with peaks assigned to C atoms in panel (a). Signals marked with * correspond to trapped solvents used in washing (DMF and CHCl₃); (d) FT-IR spectra of CATN and its constituent building blocks; (e) ζ-potential measurements of CATN in water showing a positive surface charge.

Figure 2

Preparation of the sensor electrode. (a) Schematic representation of the electrophoresis setup for CATN deposition onto a Au IDEA with Cu foil as a counter electrode; (b) optical microscopy images of the naked Au IDEA (top) and CATN-coated IDEA (bottom); (c) SEM micrograph showing a CATN-coated arm and a naked arm of the IDEA.

Figure 3

EIS detection of E. coli cells. (a) Schematic representation of the experimental setup. WE = working electrode, CE = counter electrode, S = sense, RE = reference electrode. (b) Nyquist plot showing experimental, and fitted real and imaginary components of impedance; (c) Bode plot showing experimental and fitted absolute impedance, and phase shifts as functions of frequency; (d) circuit diagram used in fitting the data shown in panels (b) and (c); (e) Bode plot showing a change in impedance as a function of frequency with increasing E. coli concentration; (f) linear relationship between the change of the impedance and the logarithm of the concentration of E. coli at 10 Hz. Line represents the linear regression curve: |Z| – |Z_PBS| = 1033 log(CFU mL^–1) – 1370; R² = 0.992.

Figure 4

Experiment with the porphyrin monomer-coated IDEA. (a) SEM micrograph showing a zoomed-in section with an uncoated gold and a 5,10,15,20-tetra(4-pyridyl)porphyrin-coated electrodes of an IDEA; (b) Bode plot showing the impedance signal as a function of frequency with increasing E. coli concentration.