A nanozyme-based cascade colorimetric aptasensor for amplified detection of ochratoxin A

Colorimetric assays have been widely developed for the detection of toxin ochratoxin A (OTA), but most of them suffer from moderate sensitivity when they are adopted for the detection of trace OTA in a complicated food matrix. For the purpose of overcoming this issue, an innovative cascade reaction-based colorimetric aptasensor was developed for the achievement of high sensitivity. The biotin-labelled OTA aptamer was immobilized onto streptavidin magnetic beads by means of the biotin-streptavidin reaction. With OTA binding to its aptamer, the structural switching of the aptamer results in the release of the alkaline phosphatase-labelled oligonucleotide, which is partially complementary to the aptamer. Following the magnetic separation, the cascade reaction is initiated through the enzymatic conversion of ascorbic acid-2-phosphate into ascorbic acid. Subsequent to that, the generated ascorbic acid reduces MnO2 nanosheets to Mn2+ ions, accordingly destroying the oxidase-mimicking activity of MnO2 nanosheets. In consequence, it is not possible to oxidize 3,3,5,5-tetramethylbenzidine (TMB), a substrate for oxidase, with Mn2+ for the production of the blue colour product (TMB Ox). With the increasing amount of OTA, a colour change occurs from blue to colourless. The cascade reaction has the potential of greatly amplifying the detection signal, together with remarkably improving the sensitivity, making this colorimetric sensor a universal and promising platform for the highly sensitive detection of mycotoxins in the field of public food safety monitoring.