A digital aptasensor for the ultrasensitive detection of T-2 toxin by using single molecule array

[Display omitted] •A novel digital aptasensor is designed for ultrasensitive detection of T-2 toxin.•The detection limit is as low as 44.5 fM (approximately 20.8 fg/mL) for T-2 toxin.•This work combined aptamer and single molecule array system provides a new avenue for detecting T-2 toxin.•The digital aptasensor can be expanded to identify similar targets and multiple detection of mycotoxins. T-2 toxin is a low molecular weight, refractory mycotoxin, often contaminating cereal-based foods, and can cause detrimental effects on human health such as nausea, vomiting, and long-term immune suppression. Therefore, highly sensitive detection platform for T-2 toxin test at the molecular level is an urgent need for food safety and public health. In this work, a digital aptasensor is developed for the ultrasensitive detection of T-2 toxin by combining aptamer and single-molecule array technology. Initially, T-2 toxin competed with the biotin-labeled complementary sequence probe to bind the T-2 toxin aptamer. The free biotin-labeled probe was then hybridized with a capture probe modified on the magnetic beads to form a stable double-stranded structure. After combining with streptavidin-enzyme, the structure was used for fluorescence imaging analysis via single-molecule microwell arrays. By correlating the concentration of T-2 toxin with the number of fluorescent microtiter wells, the ultra-sensitive detection of the T-2 toxin was realized. The developed digital aptasensor has demonstrated remarkable sensitivity toward T-2 toxin, with a detection range between 0.16 and 300 pM (approximately 0.075–140 pg/mL) and a detection limit of 44.5 fM (approximately 20.8 fg/mL). Additionally, 87.0 %-103.1 % recoveries were obtained from spiked samples in corn. This work offers a highly effective approach to detecting trace amounts of T-2 toxin, which has the potential to be used for enhancing food safety protocols, providing a powerful tool for industries and regulatory agencies to protect public health.