A sensitive fluorescence anisotropy method for detection of lead (II) ion by a G-quadruplex-inducible DNA aptamer.

Sensitive and selective detection of Pb(2+) is of great importance to both human health and environmental protection. Here we propose a novel fluorescence anisotropy (FA) approach for sensing Pb(2+) in homogeneous solution by a G-rich thrombin binding aptamer (TBA). The TBA labeled with 6-carboxytetramethylrhodamine (TMR) at the seventh thymine nucleotide was used as a fluorescent probe for signaling Pb(2+). It was found that the aptamer probe had a high FA in the absence of Pb(2+). This is because the rotation of TMR is restricted by intramolecular interaction with the adjacent guanine bases, which results in photoinduced electron transfer (PET). When the aptamer probe binds to Pb(2+) to form G-quadruplex, the intramolecular interaction should be eliminated, resulting in faster rotation of the fluorophore TMR in solution. Therefore, FA of aptamer probe is expected to decrease significantly upon binding to Pb(2+). Indeed, we observed a decrease in FA of aptamer probe upon Pb(2+) binding. Circular dichroism, fluorescence spectra, and fluorescence lifetime measurement were used to verify the reliability and reasonability of the sensing mechanism. By monitoring the FA change of the aptamer probe, we were able to real-time detect binding between the TBA probe and Pb(2+). Moreover, the aptamer probe was exploited as a recognition element for quantification of Pb(2+) in homogeneous solution. The change in FA showed a linear response to Pb(2+) from 10 nM to 2.0 μM, with 1.0 nM limit of detection. In addition, this sensing system exhibited good selectivity for Pb(2+) over other metal ions. The method is simple, quick and inherits the advantages of aptamer and FA.