Homogeneous Electrochemical Method for Ochratoxin A Determination Based on Target Triggered Aptamer Hairpin Switch and Exonuclease III-Assisted Recycling Amplification

A simple, fast, sensitive, and homogeneous electrochemical method has been developed for ochratoxin A (OTA) determination, which combines the advantage of the high selectivity of aptamer to OTA and high efficiency of exonuclease (Exo) III-assisted recycling amplification mechanism. The metastable hairpin probe (HP) was designed into three parts: regions I is the aptamer of OTA as the target recognition; regions II is poly[dA] sequence as extended DNA (E-DNA); regions III as the target DNA (T-DNA) is complementary to a part of the aptamer to stabilize the hairpin-sharp form of the HP in the absence of OTA. The DNA probe with a methylene blue tag at its 3′ terminus (MB-DNA) was designed to be complementary with T-DNA. Introduction of OTA into the assay leads to the formation change of HP from hairpin shape to open form, thus facilitating the hybridization between T-DNA and MB-DNA. The electrochemical signal is amplified through continuous Exo III cleavage. Under the optimal conditions, the differential pulse voltammetric (DPV) response had a linear relationship with the logarithm of OTA concentration in the range of 0.001~0.5 ng mL−1. In addition, the developed method has been successfully applied to detect OTA in wheat standard quality control sample. This homogeneous electrochemical sensor may have a potential prospect in detecting other molecules or proteins, which possesses the corresponding aptamer, through easily designing the hairpin probe.