Highly Accurate Multiplexed Nanoplasmonic Detection of MicroRNAs using Splinted Ligation

MicroRNAs (miRNAs) are short, non-coding single-stranded RNAs that regulate gene expression by binding to mRNAs. Because miRNAs are abnormally expressed in cancer cells, they have been considered an important cancer biomarker. However, the short length and high homology of miRNAs make them difficult to detect accurately. In this study, we developed a highly accurate miRNA detection method using a multi-channel localized surface plasmon resonance (LSPR) sensing platform. MiRNAs were labelled with a DNA probe using splinted ligation and immobilized on a gold nano-truncated cone (GNTC) array chip fabricated by nanoimprinting. After enzyme-assisted precipitation was performed, the LSPR spectral shift was measured as a function of miRNA concentration. Three miRNAs (miR-34c, miR-99a, and miR-125b) were sensitively detected with the limit of detection ranging from 1.8 to 26.1 fM. Single nucleotide variation (SNV) was clearly discriminated with high contrast. In addition, several miRNAs could be simultaneously detected in the multi-channels of a GNTC chip with high quantitative accuracy. Furthermore, we demonstrated the clinical applicability of our method by accurately detecting miRNAs in total RNA samples extracted from cancer cells.