Electrocatalytic assay of mercury(II) ions using a bifunctional oligonucleotide signal probe

Engineered nucleic acid probes containing recognition and signaling functions find growing interest in biosensor design. In this paper, we developed a novel electrochemical biosensor for sensitive and selective detecting of Hg(2+) based on a bifunctional oligonucleotide signal probe combining a mercury-specific sequence and a G-quadruplex (G4) sequence. For constructing the electrochemical Hg(2+) biosensor, a thiolated, mercury-specific oligonucleotide capture probe was first immobilized on gold electrode surface. In the presence of Hg(2+), a bifunctional oligonucleotide signal probe was hybridized with the immobilized capture probe through thymine-mercury(II)-thymine interaction-mediated surface hybridization. The further interaction between G4 sequence of the signal probe and hemin generated a G4-hemin complex, which catalyzed the electrochemical reduction of hydrogen peroxide, producing amplified readout signals for Hg(2+) interaction events. This electrochemical Hg(2+) biosensor was highly sensitive and selective to Hg(2+) in the concentration of 1.0 nM to 1 μM with a detection limit of 0.5 nM. The new design of bifunctional oligonucleotide signal probes also provides a potential alternative for developing simple and effective electrochemical biosensors capable of detecting other metal ions specific to natural or artificial bases.