Immobilized-free miniaturized electrochemical sensing system for Pb2+ detection based on dual Pb2+-DNAzyme assistant feedback amplification strategy

Publication date: 15 October 2018
Source:Biosensors and Bioelectronics, Volume 117
Author(s): Wei Cai, Shunbi Xie, Jin Zhang, Dianyong Tang, Ying Tang
We presented a novel dual-DNAzyme feedback amplification (DDFA) strategy for Pb²⁺ detection based on a micropipette tip-based miniaturized homogeneous electrochemical device. The DDFA system involves two rolling circle amplification (RCA) processes in which two circular DNA templates (C1 and C2) have been designed with a Pb²⁺-DNAzyme sequence (8–17 DNAzyme, anti-GR-5 DNAzyme) and an antisense sequence of G-quadruplex. And a linear DNA (L-DNA), which consists of a primer sequence and a Pb²⁺-DNAzyme substrate sequence, could hybridize with C1 and C2 to form two DNA complexes. In presence of Pb²⁺, the Pb²⁺-DNAzyme exhibited excellent cleavage specificity toward the substrate sequence in L-DNA, leaving primer sequence to trigger two paths of RCA process and finally resulting in massive long nanosolo DNA strands with reduplicated G-quadruplex sequences. And then, methylene blue (MB) could selectively intercalate into G-quadruplex to reduce the free MB concentration in the solution. Thereafter, a carbon fiber microelectrode-based miniaturized electrochemical device was constructed to record the decrease of electrochemical signal due to the much lower diffusion rate of MB/G-quadruplex complex than that of free MB. Therefore, the concentration of Pb²⁺ could be correctively and sensitively determined in a homogeneous solution by combining DDFA with miniaturized electrochemical device. This protocol not only exhibited high selectivity and sensitivity toward Pb²⁺ with a detection limit of 0.048 pM, but also reduced sample volume to 10 µL. In addition, this sensing system has been successfully applied to Pb²⁺ detection in Yangtze River with desirable quantitative manners, which matched well with the atomic absorption spectrometry (AAS).



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