Engineered aptamer-confined hydrogel for label-free detection of heavy-metal ions with enhanced interference...

Aptamer misfolding induced by complex sample matrices often compromises target recognition. Herein, we developed a hydrogel-confinement strategy to stabilize aptamer conformation and effectively suppress matrix interference. Using Pb²⁺ as a model analyte, aptamers were immobilized during in situ hydrogel polymerization, which helped maintain key binding-related structural features and promoted a G-quadruplex-like pre-organized conformation favorable for target recognition. The confined aptamer acted as a Tb³⁺ fluorescence sensitizer, enabling label-free Pb²⁺ detection with an apparent K_d of 0.25 μM and an LOD of 0.4 nM. The optimized aptamer hydrogel, featuring moderate interfacial cationicity and a three-site modified aptamer hydrogel showed 3.1-fold higher affinity than the free aptamer. It maintained stable responses under pH 3, salt concentrations up to 0.5 M, methanol content up to 30%, and nuclease exposure (0.16 U/mL). This strategy was further validated for Cd²⁺ sensing, demonstrating its potential for aptamer-based detection in complex food and environmental matrices.