Impact of assay format on miRNA sensing: Electrochemical microfluidic biosensor for miRNA-197 detection

MicroRNAs (miRNAs) are important biomarkers for the early detection of various diseases, especially cancer. Therefore, there is a continuing interest in different biosensing strategies that allow for the point-of-care measurement of miRNAs. Almost all miRNA sensors utilize cross-hybridization of the target miRNA with a capture probe for the recognition, which can be designed in either a sandwich or a competitive format. In this work, we present a low-cost microfluidic biosensor platform for the electrochemical measurement of miRNA-197 (a tumor biomarker candidate) in undiluted human serum samples, operating with very low sample volumes (580 nl) and a sample-to-result time of one hour. For this purpose, different on-chip miRNA bioassays based on sandwich and competitive formats are developed and compared in terms of their sensitivity, dynamic range, selectivity, precision, and simplicity. The obtained results show that, despite having a narrower dynamic range when compared to the competitive format, the sandwich assay has superior performance regarding its sensitivity and selectivity. The lowest limit of detection which can be achieved with the sandwich assay is 1.28 nM (0.74 fmole), while 4.05 nM (2.35 fmole) with the competitive format. Moreover, the sandwich assay proves to have a better distinction against single-base mismatch oligonucleotide sequences compared to the competitive one. Due to its versatility and easy handling, overcoming the issue with the sensitivity, the implemented electrochemical microfluidic biosensor could pave the way for rapid and low-cost on-site miRNA diagnostics.