Fluorometric detection of copper ions using click chemistry and the target-induced conjunction of split DNAzyme fragments

This study aimed to develop a fluorescence assay for Cu2+ monitoring with cascade signal amplification using split Mg2+-dependent DNAzyme subunits as the sensing system and fluorescence as the signal reporter. A subunit of DNAzyme can be formed through a Cu2+-catalysed click reaction between azide- and alkyne-modified short sequences immobilized on magnetic beads, eliminating the washing steps of conventional assays. The immobilized subunit recruits the assisting oligonucleotides to form an autocatalytic DNAzyme as the signal amplifier and a caged fluorescent molecule as the signal reporter. This assay is ultrasensitive, enabling the detection of trace levels of Cu2+ as low as 2 nM, which enhances the detection sensitivity by approximately 100-fold. In addition, the whole reaction process does not require a long manipulation time or a complicated procedure, making the system simple and cost-effective. This method is robust and can be applied to the determination of spiked Cu2+ in water samples with satisfactory recovery. The combination of click chemistry with fluorescent DNAzyme for a simple on-site detection assay can be used for the rapid and sensitive screening of Cu2+.