A colorimetric assay is described for determination of cytosine-rich ssDNA at physiological pH values. The working principle is based on (a) Ag(I) ion-induced formation of an i-motif structure, and (b) glucose oxidase-controlled growth of gold nanoparticles (AuNPs). The combination between Ag+ and cytosine-rich DNA can modulate the generation of H2O2 resulting from enzyme catalyzed glucose oxidation. Depending on the amount of H2O2 formed, the solution containing the AuNPs will turn red in the presence of cytosine-rich ssDNA but blue in the absence of such DNA if Ag+ is added before the formation of the red AuNPs. Upon addition of C-DNA at different concentrations, the peak shift (Δλ) of the AuNP solution relative to the SPR peak position (560 nm) in the absence of C-DNA is taken as the signal readout. The method shows a good linear response toward C-DNA over the range 10-200 nM with a detection limit of 2.7 nM. It may also be performed visually. The photometric assay is highly sensitive, specific, and rapid. The method is particularly attractive in terms of applications such as in human serum analysis, a colorimetric logic gate, and the calculation of binding constants for the interaction between Ag+ and glucose oxidase (GOx), and between Ag+ and cytosine-rich ssDNAs. Graphical abstract Schematic presentation of colorimetric detection of cytosine (C)-rich ssDNA (C-DNA) based on the modulation of the glucose oxidase (GOx)-catalyzed growth of gold nanoparticles (AuNPs) with Ag+ as the enzyme inhibitor.
Keywords: Binding constants; Dixon plot; Enzymatic activity; H2O2; Human serum; I-motif structure; Logic gate; Peak shift; Surface plasmon resonance; Transmission electron microscopy.