An electrochemical biosensor for detection of copper(ii) based on Fe3O4@Au magnetic nanoparticles and Cu2+-dependent DNAzyme assisted nicking endonuclease signal amplification

Abstract

Copper ions are essential elements in the human body and participate in various physiological activities in the bodies of organisms. Herein, an ultrasensitive electrochemical biosensor was developed for detection of copper ions (Cu²⁺) based on Fe₃O₄@Au magnetic nanoparticles (Fe₃O₄@Au MNPs) and a Cu²⁺-dependent DNAzyme assisted nicking endonuclease signal amplification (NESA) strategy. dsDNA is formed by a hybridization reaction between DNA S2 and S1 immobilized on the surface of Fe₃O₄@Au MNPs. Fe₃O₄@Au MNPs not only manifested the construction of a magnetically controlled electrochemically responsive interface but also served as promising scaffolds to carry DNA. In the presence of Cu²⁺, DNAzyme was activated and it catalyzed the cleavage of the dsDNA. The left fragment of the dsDNA on the Fe₃O₄@Au MNPs hybridized with DNA S3, initiating a polymerization reaction in the presence of DNA polymerase and dNTP and forming a long dsDNA product with specific recognition sites for the nicking endonuclease Nb.BbvCI. With the help of Nb.BbvCI, the long dsDNA product was cleaved to release the free ssDNA probe, which hybridized with Fc-labeled hairpin DNA S4 immobilized on a gold electrode and resulted in a decrease of the electrochemical signal. Subsequently, the remaining DNA probe on Fe₃O₄@Au MNPs triggered the next cycle of the “hybridization–cleavage–dissociation” reaction and more ssDNA probes dissociated, producing significant signal amplification. The developed Cu²⁺ electrochemical biosensor exhibited a reliable linear range of 1 pM to 10 μM with a detection limit of 0.4 pM (S/N = 3). Furthermore, the proposed assay platform can be easily extended to monitor other metal ions by using specific DNAzymes and it shows potential for promising application in environmental monitoring.