Probing Ionic Strength Effects on miR-141 Hybridization Kinetics by Surface Plasmon Resonance

Abstract MicroRNAs (miRNAs) are short noncoding RNAs involved in numerous cellular processes, and their dysregulation is frequently associated with human diseases. Consequently, the development of reliable strategies for miRNA detection has become increasingly important for diagnostic purposes. Many miRNA detection methods employ a complementary strand as the capture probe immobilized on a substrate and a target free in solution. Because of electrostatic repulsion between the negatively charged RNA strands, a relatively high ionic strength is required to promote hybridization. However, this can limit the sensitivity in many biosensing methods. In this work, we applied surface plasmon resonance (SPR) to investigate the hybridization kinetics between miR-141 and its complementary strand (anti-miR-141) immobilized on the chip, over a 50–400 mM ionic strength range, covering relevant physiological and biosensing conditions. For a comprehensive characterization of the interaction between the partners, both single-cycle kinetics and multicycle kinetics modes were used. At the lowest ionic strength (50 mM), a marked increase in hybridization occurred from monovalent to divalent salts, suggesting a strategy for improving the detection. In all cases, the interaction was accurately described by a 1:1 Langmuir binding model over the entire ionic strength range. On the basis of the results, an SPR biosensing strategy was developed, achieving the best limit of detection (LoD) of 0.2 nM at an ionic strength of 300 mM. These results provide a quantitative description of the influence of ionic strength on miRNA hybridization and offer a rational framework for optimizing nucleic acid hybridization and designing miRNA biosensing platforms.