An enzyme-free fluorescence biosensor for UO ₂ ²⁺ detection using a four-leaf clover-shaped quadruple walker as a signal amplifier

Given the potential threat of uranium contamination to ecosystems and human health, there is a pressing need to assess UO₂²⁺ uptake by <i>Macleaya cordata</i> (<i>M. cordata</i>) in uranium mine environments. To this end, an enzyme-free fluorescence biosensor was designed for ultrasensitive detection of UO₂²⁺ employing a DNAzyme as the recognition element and four-leaf clover-shaped quadruple walking as the amplification strategy. In the presence of UO₂²⁺, the four-leaf clover-shaped walker could be activated and move autonomously along pre-assembled tracks on MBs through DNAzyme-induced cleavage. As a result, the Cyanine 5 (Cys)/Black Hole Quencher-2 (BHQ2)-labelled tracks were cleaved and a high fluorescence signal was obtained for UO₂²⁺ detection. Benefiting from magnetic separation and the high amplification efficiency of four-leaf clover-shaped quadruple walking, the MB platform demonstrated high sensitivity and specificity for UO₂²⁺ detection, with a detection limit of 1.9 pM. This sensing system was robust and accurate, achieving the monitoring of trace UO₂²⁺ levels in <i>M. cordata</i>. Owing to its unique advantages of enzyme-free format, high sensitivity, excellent selectivity, and fluorescence readout, this sensing system holds great promise for the ultrasensitive sensing of UO₂²⁺ in real samples.