Microfluidics-based optical biosensing method for rapid detection of Escherichia coli O157:H7.

摘要

Microbial contamination of food products causes foodborne diseases and related costs, presenting an urgent need for the rapid detection of bacterial pathogens in food samples. In this study, a microfluidics-based optical biosensing method was evaluated for the rapid detection of Escherichia coli O157:H7, specifically the performance of the microfluidic channels. The biosensing system consisted of a syringe pump for sample/reagent delivery, an antibody-modified microfluidic channel as a bioseparator/bioreactor for bacterial capture and enzymatic amplification, and an optical detector for signal measurement. Affinity-purified anti-E. coli O157:H7 antibodies were chemically immobilized onto the inner wall of microfluidic channels. The immobilized antibodies captured the target bacteria in a sample being pumped through the microfluidic channel. The sandwich complexes of immobilized antibody-target bacteria-labelled antibody were formed in the channel after the alkaline phosphatase-labelled antibodies passed through. Then, p-nitrophenyl phosphate was injected into the channel as the substrate of the alkaline phosphatase conjugates, and the absorbance of the enzymatic product, 4-nitrophenol, was measured at its maximum absorption wavelength of 402 nm. The results indicated that the microfluidics-based optical biosensing method was able to detect E. coli O157:H7 with a detection limit of 10-100 cfu/ml in 2 h without any enrichment. The absorbance was not significantly different among the microfluidic channels with different lengths from 10 to 60 cm. The interference from generic E. coli and other bacterial species was negligible. The microfluidic channel designed as the bioseparator/bioreactor showed its advantages in improved immunoassay kinetics and minimized reagent consumption and its potentials for automated operation and portable biosensors..