Practical magnetic bead-based capillary electrophoresis with laser-induced fluorescence for detecting endogenous miRNA in plasma

Abstract

MicroRNAs (miRNAs) are small, non-coding RNAs crucial for gene regulation and implicated in various human diseases. Their potential as clinical prognostic and diagnostic biomarkers in biological fluids necessitates reliable detection methods. In this study, a combination of streptavidin-coupled magnetic beads and capillary electrophoresis with laser-induced fluorescence (CE-LIF) was used to extract and analyze plasma miRNAs. Specifically, miRNAs hybridized with a biotinylated fluorescent DNA probe were isolated from plasma using magnetic beads. These hybridized miRNAs were then directly injected into the CE-LIF system for analysis, eliminating the need for additional processing steps. Both the hybridization and bead-to-probe binding were executed concurrently, regulated by temperature and time. Through the optimization of magnetic bead extraction and CE-LIF conditions, we developed a highly sensitive assay for miR-21 quantification in plasma. The assay displayed remarkable linearity (R2 = 0.9975) within a 0.1–5 pM range and exhibited favorable precision (0.22–1.26 %) and accuracy (98.31–111.19 %). Importantly, we successfully detected endogenous miR-21 in plasma samples from both a lung cancer patient and healthy adults, revealing a 1.7-fold overexpression of miR-21 in lung cancer plasma relative to normal samples. Our findings suggest that this developed system offers a simple and sensitive approach for detecting endogenous miRNAs in plasma, showing its potential utility in disease diagnostics. To our knowledge, this is the first study to utilize CE-LIF for plasma miRNA detection. © 2023 The Authors