Ligation-Driven Electrochemical Magneto-Genoassay Platform Based on PNA Probes for the Multiple Detection of Soy and Mustard DNA in Wheat Flour

Food allergies are one of the most critical food safety issues, with epidemiological studies confirming a global increase. In this context, effective and sensitive analytical methods play a crucial role in ensuring allergen-free food products. To face this issue, electrochemical biosensors offer powerful, sensitive, selective, and cost-effective alternatives to conventional methods for food allergen analysis while enabling rapid on-site detection. In this study, we developed a sandwich electrochemical magneto-genoassay aimed at the parallel detection of soy (Glycine max) and mustard (Sinapis alba) allergens, suitable for implementation on multichannel instrumentation. The assay involves the functionalization of magnetic microbeads functionalized with peptide nucleic acid-based (PNA) capture probes, capable of undergoing target-induced bio-orthogonal ligation with biotin-labelled signalling probes. Carbon nanotubes-modified screen-printed carbon electrodes were exploited for the voltammetric readout. We demonstrated the effectiveness of functional PNA probes by comparing their performance with those achieved using analogous DNA probes. The developed method exhibited excellent selectivity in terms of cross-reactivity, sensitivity, and precision, achieving detection limits of 16 and 19 pM for soy and mustard, respectively. Finally, by successfully applying the biosensor platform to genomic DNA extracted from plant-based food ingredients, we demonstrated its potential as a valuable tool in food safety risk management.