Lipid nanoparticles (LNPs) are widely used for the delivery of therapeutic microRNAs (miRNAs), especially in coformulated systems where multiple targets are combined to enhance therapeutic efficacy. However, the simultaneous quantification of miRNAs within LNP formulations remains a major analytical challenge for pharmaceutical quality control. This work presents a dual-working electrochemical biosensor for multiplexed, selective, and amplification-free quantification of two miRNAs (miR-4676 and miR-6503) encapsulated in LNPs. The biosensor was fabricated on a dual-working screen-printed graphite electrode via gold electrodeposition, followed by site-specific immobilization of methylene blue (MB)-labeled ss-DNA probes. After surfactant-mediated LNP lysis, square wave voltammetry (SWV) enabled hybridization-based signal-off detection of the released targets. The sensor showed excellent analytical performance, with limits of detection of 0.96 nM for miR-4676 and 0.98 nM for miR-6503. High selectivity was confirmed through cross-reactivity experiments. In real formulations with different miRNA ratios (1:1 and 2:1), the sensor achieved quantification accuracies of 88% and 84%, respectively, compared to a standard fluorimetric assay. Unlike conventional methods, the platform enabled individual miRNA quantification in complex mixtures. This study introduces an electrochemical platform enabling multiplexed miRNA quantification within LNPs, providing a rapid, low-cost, and portable solution for quality control of miRNA-based formulations.
Cimmino et al. (2026) studied this question.