Detachable nanoladders: A new method for signal identification and their application in the detection of ochratoxin A (OTA)

Abstract A highly sensitive fluorescence turn-off biosensor for the detection of ochratoxin A (OTA) was developed based on graphene oxide (GO) and an aptamer-induced detachable nanoladders. In this assay, two types of ssDNA (H1 and H2) were involved in the assembly of the DNA nanoladders, in which H1 was labeled with fluorophore, and H2 was the OTA binding aptamer. Self-assembly of the DNA nanoladders with the addition of GO weakened its adsorption and the fluorescence intensity remained strong. In the presence of OTA, the aptamer was specifically recognized and an aptamer-OTA complex was formed, leading to the detached of DNA nanoladders. With the addition of GO, the released H1 was adsorbed on the GO surface, thus efficiently quenching the fluorescence signal (turning off). The detection limit of OTA in this assay was 4.59 nM. To improve the sensitivity of this strategy, we creatively developed an alternative strategy to replace the sturdy nanoladders with frail nanoladders. More precisely, the sequences of H1 had mismatched bases, which, when hybridized with H2 could be used to create long non-perfect complementary nanoladders. For the mismatched bases-based frail nanoladders, it was easier for OTA to bind its aptamer sequence, thus enabling a more thorough and faster detachment of the nanoladders, along with a greater degree of fluorescence quenching. The detection limit for OTA was estimated to be 0.1 nM. The biosensors we developed were sensitive, specific, enzyme-free, cost-effective and can be applied in red wine samples spiked with known concentration of OTA.