Electrokinetic stringency control in self-assembled monolayer-based biosensors for multiplex urinary tract infection diagnosis

Abstract

Rapid detection of bacterial pathogens is critical toward judicious management of infectious diseases. Herein, we demonstrate an in situ electrokinetic stringency control approach for a self-assembled monolayer-based electrochemical biosensor toward urinary tract infection diagnosis. The in situ electrokinetic stringency control technique generates Joule heating induced temperature rise and electrothermal fluid motion directly on the sensor to improve its performance for detecting bacterial 16S rRNA, a phylogenetic biomarker. The dependence of the hybridization efficiency reveals that in situ electrokinetic stringency control is capable of discriminating single-base mismatches. With electrokinetic stringency control, the background noise due to the matrix effects of clinical urine samples can be reduced by 60%. The applicability of the system is demonstrated by multiplex detection of three uropathogenic clinical isolates with similar 16S rRNA sequences. The results demonstrate that electrokinetic stringency control can significantly improve the signal-to-noise ratio of the biosensor for multiplex urinary tract infection diagnosis.

From the clinical editor: Urinary tract infections remain a significant cause of mortality and morbidity as secondary conditions often related to chronic diseases or to immunosuppression. Rapid and sensitive identification of the causative organisms is critical in the appropriate management of this condition. These investigators demonstrate an in situ electrokinetic stringency control approach for a self-assembled monolayer-based electrochemical biosensor toward urinary tract infection diagnosis, establishing that such an approach significantly improves the biosensor's signal-to-noise ratio.

Keywords: Matrix effects; Multiplex detection; Self-assembled monolayers; Stringency control; Urinary tract infection.

Figure 1

(A) Self-assembled monolayer based electrochemical pathogen sensor. (B) Non-specific binding is removed by the temperature rise and fluid flow for washing based on electrokinetic control. (C) Pseudomonas aeruginosa, Staphylococcus saprophyticus, Enterococcus faecalis, and Escherichia coli are detected in the conditions of diffusion and electrokinetics. (D) The concentration of E. coli was measured with respect to diffusion and electrokinetics. A square wave with voltage of 6 Vpp at 200 kHz was applied for 8 minutes.

Figure 2

Fraction of species (perfect match and single-base mismatch) by applying a 200 kHz square wave. The signal was normalized by the signal of the perfect match target at room temperature.

Figure 3

Comparison of (A) experimental fraction (perfect match and single-base mismatch) results with different temperatures driven by electrokinetics, (B) predicted fraction (perfect match and single-base mismatch) of nearest-neighbor model and (C) experimental fraction results in temperature control with an incubator.

Figure 4

Voltage dependence of the background noise in human urine samples.

Figure 5

Multiplex detection of bacterial clinical isolates (P. mirabilis 1.11×10⁷ CFU/ml, P. aeruginosa 1.0×10⁷ CFU/ml, E. coli 3.2×10⁶ CFU/ml) using (A) E. coli capture probe, (B) P. mirabilis capture probe and (C) P. aeruginosa capture probe and cocktail detector probe with and without in situ electrokinetic stringency control.