DNA-Templated Silver Nanoclusters for Fluorescence Turn-on Assay of Acetylcholinesterase Activity

Abstract

We have developed a fluorescence turn-on assay using DNA-templated silver nanoclusters (Ag NCs) (i.e., 12 polycytosine-templated silver nanoclusters, dC12-Ag NCs), which is amenable to rapid, ultrasensitive assay of acetylcholinesterase (AChE). The detection mechanism is based on the concept, that is, AChE hydrolyzes the acetylthiocholine (ATCh) chloride to produce thiocholine (TCh). Subsequently, TCh sensitively and rapidly reacts with dC12-Ag NCs via Ag-S bond forming and enhances the fluorescence of dC12-Ag NCs. Using dC12-Ag NCs, detection of TCh has a linear concentration range of 2.0 nM to 16.0 nM and a detection limit of 0.3 nM. Due to the sensitive and rapid fluorescence turn-on response of dC12-Ag NCs to TCh, AChE with activity as low as 0.5 × 10(-4) U/mL (signal/noise = 3) can be analyzed with a dynamic range of 0.1 to 1.25 × 10(-3) U/mL. The promising application of the proposed method in AChE inhibitor screening was demonstrated. AChE concentrations were determined in human blood red cell (RBC) membranes from clinical specimens using dC12-Ag NCs, and the quantitative results were validated with Ellman's method. Aside from the ease of manufacture, reduction of matrix effect, and low background noise, the continuous detection format and detection sensitivity can open up to wider applications to AChE activity assay in neurobiology, toxicology, and pharmacology, among other fields.

Gold nanoclusters–Cu2+ ensemble-based fluorescence turn-on and real-time assay for acetylcholinesterase activity and inhibitor screening

“…4E). This value is in the same order of magnitude as those previously reported by other AChE assays (Lei et al, 2015;Zhang et al, 2013). It is essentially demonstrated that our ensemble-based fluorescence assay could be used to screen the potential AChE inhibitors.…”

“…Among them, fluorescent materials-based assays have several advantages and attractive potentials owing to their intrinsic high sensitivity and real-time monitoring with fast response time. Indeed, along with traditional organic fluorophores (Liu et al, 2012;Maeda et al, 2005), several novel fluorescent nanomaterials such as semiconductor quantum dots (Saa et al, 2010) and DNA templated metal nanoclusters Zhang et al, 2013) have already been utilized to assess AChE activity and screen its inhibitors.…”

DNA-stabilized, fluorescent, metal nanoclusters for biosensor development

“…They attributed this increase to either new charge transfer enabled by the Ag-S bond or by the change of the local environment brought by the thiols [130]. This enhancement was utilized to detect the acetylcholinesterase activity, where a free thiol was generated via the enzymatic reaction [131].…”

In situ induced metal-enhanced fluorescence: A new strategy for biosensing the total acetylcholinesterase activity in sub-microliter human whole blood

“…Several quantification techniques on AChE (Holas et al, 2012;Miao et al, 2010;Sabelle et al, 2002;Feng et al, 2007;Wang et al, 2009;Peng et al, 2009;Golub et al, 2013;Liao et al, 2013aLiao et al, , 2013bZhang et al, 2013;Garai-Ibabe et al, 2014) have been developed, each of which is suitable for different purposes and can be improved on user demand. The existing methods can solve problems in AChE activity determination.…”