Three detection modes with one electrode: A multifunctional electrochemical sensing platform for thrombin, p53 gene, and cholesterol detection

Abstract

The development of diverse biomarker detection platforms is crucial to efficient and low-cost assays. Unlike most conventional multi-target methods requiring complex probe labeling and prone to signal cross-talk, a novel multifunctional sensor incorporating host-guest chemistry in electrochemical assays was developed to detect protein, nucleic acid, and small molecule with a single electrode in this work. This sensor operates in three distinct modes: protein detection, nucleic acid detection, and small molecule detection. In the protein detection mode, the methylene blue (MB)-labeled probe recognized the thrombin and formed a probe/thrombin complex, inducing fewer probes to be captured by the β-cyclodextrin (β-CD) and enabling a measurable electrochemical signal change. In the nucleic acid detection mode, the p53 gene interacted with the corresponding probe and formed a rigid double-stranded DNA (dsDNA) structure, reducing the electron transfer rate constant (k_s) and inhibiting MB signal transmission. In the small molecule detection mode, cholesterol competitively bonded with the host molecules and removed MB from the β-CD cavity, leading to an electrochemical signal decrease. Under optimized detection conditions, this sensor displayed an effective analytical performance for thrombin, p53 gene, and cholesterol, achieving low detection limits of 61.7 pM, 7.3 nM, and 14.3 μM, respectively. Importantly, this platform enables rapid regeneration in the nucleic acid detection mode and reliable operation in complex samples, outperforming previous multi-target detecting methods in reusability, anti-interference capacity, and ease of construction. By eliminating multi-labeling steps and minimizing cross-talk on a single electrode, our design may inspire more innovative multifunctional sensors for multiplexed biomarker diagnostics.