Hydrogel Microneedle-Integrated Electrochemical Sensor With Negative-Potential Readout Toward Wash-Free and...

Electrochemical quantification of microRNAs (miRNAs) in tumor interstitial fluid (TIF) may provide complementary molecular information for tumor microenvironment assessment, but analytical accuracy is often limited by extraction-induced biomarker loss/degradation and background interference from endogenous electroactive metabolites. Herein, a hydrogel microneedle-integrated electrochemical biosensor is developed as a complementary ex vivo molecular sensing approach for wash-free and anti-interference monitoring of miRNAs via negative-potential readout. The sensing platform employs a four-way junction (FWJ) nucleic acid nanoreservoir to encapsulate the electroactive indicator doxorubicin (DOX), which is released upon target miRNA-triggered strand displacement. The characteristic oxidation potential of DOX (-0.65 V) enables detection in a negative potential window, thereby reducing co-oxidation interference by endogenous electroactive species. The porous hydrogel microneedle architecture facilitates rapid TIF sampling and directional diffusion of released DOX toward the electrode interface. The biosensor achieved a detection limit of 1.25 pM for miRNA-21 with an interassay variability of <1.9%. Preliminary analysis of clinical TIF samples from breast cancer and healthy donors further supported the feasibility of direct miRNA detection, yielding an AUC of 0.92. This work offers a novel strategy by integrating ex vivo in situ TIF sampling with negative-potential readout to achieve wash-free and anti-interference detection of biomarkers in complex physiological matrices.