A net fishing enrichment strategy for colorimetric detection of E. coli O157:H7

Abstract

The strict regulatory requirements for pathogen monitoring in food systems to ensure safety demands that the detection method can recognize small numbers of pathogens. Although significant efforts on the development of biosensors have been reported with marked improvement in sensitivity, appropriate enrichment strategies are still needed in a detection scheme. Herein we proposed a unique strategy for the enrichment of pathogens, based on a concept termed as net fishing, to capture target bacteria with antibodies functionalized on porous substrates to fish the target from samples. Because of their large surface area porous nitrocellulose membranes (NC membranes) were used as substrates for target capture. By immersing the antibody modified NC membrane into liquid samples and upon mixing, we expect to anchor target pathogens on the membrane due to the specific interaction between antibody and target, where the nonspecific enrichment could be reduced compared to methods such as filtration. Meanwhile the net fishing enrichment requires 2h, less than common culture methods. An enzyme enhanced procedure was performed with the NC membrane after the capture to obtain colorimetric signal. Gold nanoparticle (GNP) probes conjugated with horseradish peroxidase (HRP) and antibody were used to label capture target bacteria, and the presence of pathogens was determined by the colorimetric signal generated from HRP catalyzed TMB reaction. With E. coli O157:H7 as target, we investigated the detection performance of the proposed net fishing strategy with an enzyme amplification step for colorimetric reporting. After optimization of detection conditions, we show that as low as 100CFU/ml E. coli O157:H7 could be detected. The feasibility was then demonstrated using pineapple juice as a food model.