A DNAzyme assay coupled with effective magnetic separation and rolling circle amplification for detection of lead...

Rapid, portable, and efficient detection of lead cations (Pb²⁺) is of great significance for monitoring and evaluating environmental toxicity and human healthcare. In this work, we developed a simple and low-cost homogenous fluorescence DNAzyme assay for Pb²⁺ determination based on Pb²⁺-dependent cleaving and rolling circle amplification (RCA). DNAzyme and its substrate (5'-biotin) formed double-stranded hybrids in solution which could thoroughly react with Pb²⁺ in aqueous phase. Then, the unreacted DNAzyme/substrate hybrids as well as cleaved parts with biotin labeling of substrate strand would be captured by magnetic beads through biotin-streptavidin interactions and removed from the reaction solution. Meanwhile, the other parts of the substrate strand remained in solution and subsequently acted as the primer and triggered RCA. The concentration of the cleaved substrate strand correlated to that of Pb²⁺ and non-specific amplification was effectively minimized through biotin-streptavidin isolation. With a smartphone camera, the fluorescence intensity was recorded and quantified after 30-90 min amplification, making it a portable method with minimum instrumentation. A dynamic range of 1-100 nM of Pb²⁺ was achieved under optimized conditions and it was successfully employed for Pb²⁺ detection in spiked lake water. Graphical abstract.