Shiga toxin-producing E. coli (STEC) is a food-borne pathogen of significant concern due to the severity of the disease it can cause. Herein we report the development of a highly sensitive, label-free, electrochemical DNA-based sensor for detection of stx1 gene using interdigitated gold microelectrodes (IDEs) on fully integrated silicon chips. Each IDE comprised a working IDE, used for DNA probe immobilisation and generator IDE used for accumulation of methylene blue. First, the working IDE was modified with gold nanoparticles (Au NPs) and chitosan gold nanocomposite. Afterwards, amine-modified probe DNA was immobilised on the chitosan modified electrode using glutaraldehyde as a linker. The label-free electrochemical detection was undertaken using methylene blue as a redox molecule, which intercalated into the double-strand DNA after applying an open potential circuit at the generator IDEs. Reduction of methylene blue was recorded using square wave voltammetry (SWV). Using this label-free detection, we have achieved linear response between 10-16 and 10-6 M synthetic target strand with the lowest limit of detection of 100 aM after 20 minutes hybridisation time. The chromosomal DNA from four different E. coli strains (two stx1 positives and two stx1 negatives) was used to confirm the selectivity of the presented method. This novel on-chip biosensor for the detection of STEC has the potential to be used in point-of-use detection, for example, on the farm.
Highly sensitive electrochemical sensor for the detection of Shiga toxin-producing E. coli using interdigitated micro-electrodes selectively modified with a chitosan-gold nanocomposite