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

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 LPS content, a miniaturized electrochemical cell sensor consisting of a screen-printed paper electrode, 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 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 ng/mL 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 ng/mL to 3 ng/mL. These results were confirmed by analysis of nitric oxide release from Raw264.7 via the Griess method. The miniaturized sensor was ultimately applied to detect LPS in fruit juice samples. The results indicated that the method exhibited high recovery and relative standard deviation lower than 2.65%, and LPS in samples contaminated with 102 CFU/mL - 105 CFU/mL bacteria could be detected, which proved the practical value of the sensor. Thus, a novel, low-cost and highly-sensitive approach to LPS detection was developed, providing a method to assess Gram-negative bacteria contamination in food.