Homogeneous PDA/Fe₃O₄ Nanocomposites and 3D-HCR/AgNCs-Based One-Pot Biosensor for Triple-Modal Nucleic Acid Detection

The seamless and efficient integration of multiple detection modes─such as electrochemical, colorimetric, magnetic, thermal, and fluorescent signaling─into a single sensing platform remains a significant challenge in advanced sensing technology. In this work, we developed a trimodal biosensor based on homogeneous polydopamine (PDA)/Fe₃O₄/CP (CP: capture probe) nanocomposites and three-dimensional hybridization chain reaction-templated silver nanoclusters (3D-HCR/AgNCs) for the ultrasensitive and precise detection of nucleic acids. Briefly, the presence of the target KRAS gene segment initiates a hybridization chain reaction (HCR), which acts as a bridge linking capture probes (PDA/Fe₃O₄/CP) and signal probes (3D-HCR/AgNCs). The HCR bridge hybridizes with both components, forming a PDA/Fe₃O₄/CP/HCR-bridge/3D-HCR/AgNCs complex that can be magnetically separated. Since DNA-templated AgNCs generate measurable signals across three distinct modes─electrochemistry, fluorescence, and visible/UV-vis spectroscopy─a fully integrated trimodal sensing platform is realized. The results from these three modes can be cross-verified, significantly enhancing the reliability of the assay. Leveraging homogeneous PDA/Fe₃O₄/CP nanocomposites and a strategically designed blocker, this strategy enables straightforward, one-pot detection of target nucleic acids without the need for vibration, electrode modification, or multiple washing and separation steps. Importantly, beyond serving as a signal amplifier, the 3D-HCR structure also promotes aggregation-induced emission enhancement of AgNCs, thereby increasing the sensitivity of the fluorescence mode. This work introduces a facile and efficient approach that paves the way for the construction of integrated trimodal sensing platforms.