Copper-Mediated DNA-Scaffolded Silver Nanocluster On-Off Switch for Detection of Pyrophosphate and Alkaline Phosphatase.

ABSTRACT

We present a new copper-mediated on-off switch for detecting either pyrophosphate (PPi) or alkaline phosphatase (ALP) based on DNA-scaffolded silver nanoclusters (DNA/AgNCs) templated by a single-stranded sequence containing a 15-nt polythymine spacer between two different emitters. The switch is based on three favorable properties the quenching ability of Cu(2+) for DNA/AgNCs with excitation at 550 nm; the strong binding capacity of Cu(2+) and PPi; and the ability of ALP to transform PPi into orthophosphate (Pi). The change in fluorescence of DNA/AgNCs depends on the concentrations of Cu(2+), PPi, and ALP. Copper(II) acts as a mediator to interact specifically with the Probe, while PPi and ALP convert the signal of the Probe by removing and recovering Cu(2+), operating as an on-off switch. In the presence of Cu(2+) only, DNA/AgNCs exhibit low fluorescence because the combination of Cu(2+) and DNA template disturbs the precise formation of DNA/AgNCs. When PPi is added to the system containing Cu(2+), free DNA template is obtained due to the stronger interaction of PPi and Cu(2+), leading to a significant fluorescence increase (ON state) which depends on the concentration of PPi. Further addition of ALP results in the release of free Cu(2+) via ALP-catalysis of hydrolysis of PPi into Pi, thereby returning the system to the low fluorescence OFF state. The switch allows the analysis of either PPi or ALP by observation of the fluorescence status, with the detection limit of 112.69 nM and 0.005 U/mL for PPi and ALP, respectively. The AgNCs on-off switch provides the advantages of simple design, convenient operation, and low experimental cost without need of chemical modification, organic dyes, or separation procedures.