Electrochemical Aptasensor Based On Hybridization Chain Reaction For Ultrasensitive Detection Of Ochratoxin A

Ochratoxin is a toxic foodborne mycotoxin produced mainly by Aspergillus ochraceus,Aspergillus carbonarius and Penicillium verrucosum.They frequently contaminate grain and agricultural products,and close with human and animal health.Among them,Ochratoxin A(OTA)is endowed with the highest toxicity,distribution of the most extensive and the highest level of pollution.We designed an electrochemical aptasensor based on hybridization chain reaction with ultra-low detection limit for the direct detection of OTA.First,a capture probe was immobilized on an electrode and then hybridized with an aptamer to form an aptamer/DNA duplex.In the presence of OTA,the formation of an aptamer-OTA complex results in the dissociation of the aptamer from the electrode.Next,the detection probe and two hairpin-helper DNAs were added,which combine with the remaining capture probe and led to the formation of extended ds DNA polymers through HCR on the electrode surface.Finally,a streptavidin-alkaline phosphatase(ST-AP)conjugate that binds to the remaining biotinylated hairpin DNAs was added.The electrochemical signal was produced through the catalytic hydrolysis between ST-AP and the substrate-naphthyl phosphate(-NP),DPV current that increases with increasing OTA concentration.Under optimal conditions,this method could detect OTA from 0.005 to 100 ng m L-1,with a detection limit of 2 pg m L-1.The assay was successfully applied for the determination of OTA in cereal samples.And the results showed satisfactory recovery and good agreement with those of HPLC.