Harnessing soft DNA nanospheres as a superior alternative to hard nanoparticles for boosting DNAzyme-based colorimetric sensing of lead ions

Lead ions (Pb 2+ ), as a persistent and bioaccumulative environmental toxin, pose a significant threat to public health, demanding ultrasensitive and field-deployable detection methods. Conventional sensors often struggle with a trade-off between sensitivity and stability, especially when employing hard nanomaterial enhancers like gold nanoparticles. However, the practical utility of these inorganic nanoprobes is often compromised by their susceptibility to surface passivation (biofouling) and aggregation in salt-rich or protein-rich environments. Herein, we introduce a novel concept of a soft-matter DNA nanoreactor for engineering a high-performance colorimetric biosensor. This nanoreactor is self-assembled from a single palindromic DNA strand into a DNA nanosphere (DS), creating a spatially confined microenvironment. By anchoring a Pb 2+ -specific DNAzyme system onto this platform, we developed the DS-S-Dz probe. The nanoreactor architecture dramatically enhances the catalytic efficiency of the DNAzyme through a profound spatial confinement effect, which accelerates the Pb 2+ -triggered cleavage of a substrate strand and subsequent release of a G-rich sequence. This liberated sequence then folds into a G-quadruplex/hemin DNAzyme, catalyzing a robust colorimetric reaction. This strategy effectively suppresses background noise while amplifying the target signal, achieving a remarkable limit of detection of 0.96 nM, a 50-fold improvement over the non-amplified system (48 nM) and well below the EPA safety limit (72 nM). The sensor demonstrated exceptional selectivity against a panel of interfering metal ions and proved its robustness and accuracy in complex real-world samples like tap water and milk. This work presents a paradigm shift from simple nanoparticle enhancement to the rational design of programmable DNA nanoreactors, offering a powerful and elegant platform for sensitive and reliable environmental monitoring.