Aptamer-Quantum Dot Lateral Flow Test Strip Development for Rapid and Sensitive Detection of Pathogenic Escherichia coli via Intimin, O157- Specific LPS and Shiga Toxin 1 Aptamers

Background: The use of DNA aptamers to replace antibodies in lateral flow (LF) test strip assays coupled to quantum dots (QDs) has demonstrated enhanced sensitivity for the rapid and portable detection of several foodborne pathogenic bacterial species with simple ultraviolet (UV) illumination and visual assessment (Bruno, Pathogens, 2014; 3: 341-355). Objectives: The present report extends and focuses development on detection of O157- specific lipopolysaccharide (LPS), intimin protein and Shiga toxin 1 (Stx 1) using the same aptamer-QD LF assay approach to aid in rapid and sensitive detection of E. coli O157:H7 and other pathogenic serotypes of E. coli. Methods: Numerous anti-O157 LPS, intimin and anti-Stx 1 aptamer DNA sequence candidates were developed and screened by enzyme-linked aptamer assay (ELASA) followed by further screening of the best candidates in less expensive aptamer- latex particle or aptamer-colloidal gold (CG) LF formats. The highest affinity and most specific aptamer candidates were incorporated into the aptamer-QD LF format. Results: Several high affinity and specific aptamers against intimin, Stx1, and O157 LPS were developed and found to work well in various capture-reporter conjugate combinations using latex particles, CG and QDs with detection limits as low as 100 E. coli O157:H7 bacterial cells and 10 ng of Stx 1 in buffer by visual assessment. Conclusion: The seminal work in this aptamer-QD LF area has been extended to sensitive detection of E. coli O157 cells as well as detection of other pathogenic E. coli serotypes by targeting intimin and Stx 1.