Double-Stranded Ligation Assay for the Rapid Multiplex Quantification of MicroRNAs.

Abstract

MicroRNAs are auspicious candidates for a new generation of biomarkers. The detection of microRNA panels in body fluids promises early diagnosis of many diseases, including cancer or acute coronary syndrome. For a fast, sensitive, and specific detection of microRNA panels on the bedside, medical point-of-care systems that measure those biomarkers are required. As microchips are promising technical tools for a robust signal measurement at biochemical interfaces, we developed an assay for the electrochemical multiplex quantification of microRNAs on a CMOS chip with interdigitated gold electrode sensor positions. The method is based on the formation of a tripartite hybridization complex and subsequent both-sided ligation of the target nucleic acid to a reporter and capture strand. With a time to results of 30 min, the reported assay achieves a limit of detection below 1 pM, at a specificity down to single mismatch discrimination. It also offers very good signal dynamics between 1 pM and 1 nM, thus, allowing reliable quantification of the detected microRNAs and easy implementation into automated devices due to a simple workflow.