Miniaturized Paper-Supported 3D Cell-Based Electrochemical Sensor for Bacterial Lipopolysaccharide Detection

Abstract

Sensitive detection of lipopolysaccharides (LPSs), which are present on the outer wall of Gram-negative bacteria, is important to reflect the degree of bacterial contamination in food. For indirect assessment of the LPS content, a miniaturized electrochemical cell sensor consisting of a screen-printed paper electrode, a three-dimensional cells-in-gels-in-paper culture system, and a conductive jacket device was developed for in situ detection of nitric oxide released from LPS-treated mouse macrophage cells (Raw264.7). Nafion/polypyrrole/graphene oxide with excellent selectivity, high conductivity, and good biocompatibility functionalized on the working electrode via electrochemical polymerization could enhance sensing. Raw264.7 cells encapsulated in the alginate hydrogel were immobilized on a Nafion/polypyrrole/graphene oxide/screen-printed carbon electrode in paper fibers as a biorecognition element. Differential impulse voltammetry was employed to record the current signal as-influenced by LPS. Results indicated that LPS from Salmonella enterica serotype Enteritidis caused a significant increase in peak current, varying from 1 × 10–2 to 1 × 104 ng/mL, dose-dependently. This assay had a detection limit of 3.5 × 10–3 ng/mL with a linear detection range of 1 × 10–2 to 3 ng/mL. These results were confirmed by analysis of nitric oxide released from Raw264.7 via the Griess method. The miniaturized sensor was ultimately applied to detect LPSs in fruit juice samples. The results indicated that the method exhibited high recovery and relative standard deviation lower than 2.65% and LPSs in samples contaminated with 102–105 CFU/mL bacteria could be detected, which proved the practical value of the sensor. Thus, a novel, low-cost, and highly sensitive approach for LPS detection was developed, providing a method to assess Gram-negative bacteria contamination in food.

Advances in Nanomaterials-Based Electrochemical Biosensors for Foodborne Pathogen Detection

“…GO combined with chitosan has been demonstrated to be an excellent means for electron transfer for the detection of short DNA sequences achieving the detection limit of 3.584 × 10 −15 M [94]. Paper-based sensors with screen-printed electrodes modified by a Nafion/PPy/GO composite were proposed for the detection of lipopolysaccharides (LPSs), which are a marker for Gram-negative bacteria [117]. Raw264.7 macrophage cells were used as a recognition element.…”

Current State of Development of Biosensors and Their Application in Foodborne Pathogen Detection

“…The limitation of previously mentioned sensors is that none of these tests can confirm the functionality of the targeted toxin and pathogen, as antibody only picks up a specific region on the surface, and aptamers only recognize a partial sequence of the analyte. (13,14,43,96).…”

Electrochemical Cell-Based Sensor for Detection of Food Hazards

“…Nafion/polypyrrole/graphene oxide composites have excellent selectivity, high conductivity and good biocompatibility, which can be enhanced by electrochemical polymerization on the working electrode. Mouse macrophage cells encapsulated in alginate hydrogel were used as biological cognitive elements to immobilize Nafion/polypyrrole/graphene oxide screen-printed carbon electrodes in paper fibers [ 33 ].…”

“…Compared with Ponsonnet’s design [ 60 ], the cells can have a better physiological state to detect lipopolysaccharide. The sensor has a good detection effect on lipopolysaccharides; the detection limit of the sensor is 3.5 × 10 −3 ng/mL and the linear detection range is 1 × 10 −2 to 3 ng/mL [ 33 ].…”

“… Schematic illustration of the construction and working principle of the miniaturized paper-supported 3D cell-based electrochemical sensor [ 33 ]. Copyright © 2020, American Chemical Society.…”