Ultrasensitive detection of total copper with an electrochemical biosensor built on the in cis coupling of hexynyl CLICK-17 DNAzyme with azido self-assembled monolayers

Abstract

New methods to detect copper sensitively and specifically are of practical importance because of its presence in both ecological and biological systems. Herein, we report a newly selected DNAzyme, CLICK-17, which catalyzes the azide-alkyne cycloaddition reaction strictly dependent on the presence of copper, as a superior transducer for electrochemical copper quantitation. CLICK-17 offers the following notable advantages: it does not require the presence of an unstable reducing agent (detects both Cu(II) and Cu(I) in samples); it offers rapid detection owing to the fast in cis coupling kinetics; and, it generates no background because its catalytic activity strictly relies on the presence of copper (signal-on). For performing electrochemical sensing, the DNAzyme efficiently couples to a binary 11-azido-1-undecanethiolate/octanethiolate monolayer on gold (N3C11S-/C8S-Au), which is then accurately quantitated by the cyclic voltammetric response of electrostatically bound [Ru(NH3)6]3+. As the negative charges on the phosphate backbone of surface-bound 79nt-DNA strands (CLICK-17) are inclusively compensated by the multiply charged redox cations (e.g., [Ru(NH3)6]3+) at low ionic strength, the electrochemical signal is naturally amplified (~ 30 folds). Thus constructed biosensor detects Cu(II) with a LOD of 3.5 nM and Cu(I) with a LOD of 0.8 nM, which is also free of background signals and highly specific against other metal ions. More importantly, we have shown the successful application of this DNAzyme-based electrochemical sensor to determine copper concentrations in natural ores from the mining industry, with performance comparable to the gold standard flame atomic absorption spectrophotometry.