Highly sensitive and selective antibody microarrays based on a Cy5-antibody complexes coupling ES-biochip for E. coli and Salmonella detection

Abstract

Foodborne pathogens are threats in food and a cause of major health issues globally. Microbial safety has become a key concern to eliminate disease-causing pathogens from the food supply. For this purpose, the Cy5 dye conjugated with a double-biotin DNA linkage and a detection antibody (Cy5-Ab complexes) was developed to amplify a foodborne detection signal on a microarray. Additionally, the ES-biochip was designed to attain a visual screening of an antibody microarray for the simultaneous threat detection of Salmonella and Escherichia coli (E. coli). Quantification was also performed by fluorescence. After optimizing the Cy5-Ab complex appendage and enhancing the detection signal from a sandwich immunoassay, high sensitivity and selectivity were observed. The limits of detection for both pathogens in buffer and food samples were 10³ CFU mL⁻¹ and less than 9 CFU mL⁻¹ by visual screening and fluorescent intensity quantification, respectively. Mono and duplex responses were not significantly different which means that no cross-reactivity occurred. Uniquely, the assays hold great potential to be used in several fields, such as clinical diagnosis of foodborne microbes, food hygiene screening, and pathogen detection.