Graduate student Incheon National University Incheon, Inch'on-jikhalsi, Republic of Korea
Abstract: Microfluidics-based microwells with picoliter volumes are widely used for single-molecule analysis in the field of bioanalysis. Microwells play a crucial role in digitally detecting nucleic acids and proteins, offering significantly higher sensitivity compared to bulk-scale assays. However, conventional microwell systems for detecting multiple targets often require multiple fluorescence probes, resulting in high per-assay costs and complex experimental setups.
In this study, we have developed a multiplexed digital detection platform by integrating microwell technology with barcoded magnetic microparticles (b-MMPs) to identify various microRNAs. Marked with unique barcodes, the probe-labeled b-MMPs allow for the distinct identification of target microRNAs. This approach enables the detection of a range of microRNAs with high sensitivity without the need for nucleic acid amplification techniques such as rolling circle amplification (RCA) or hybridization chain reaction (HCR).
To realize this concept, we manufactured b-MMPs using the discontinuous dewetting method and captured target miRNAs on their surfaces through a sandwich hybridization process. Subsequently, signal amplification took place within the segregated microwells through the interaction of a labeled enzyme (streptavidin-HRP) with QuantaRed™ substrate solution. Our results demonstrated that miR-21 could be detected with increased sensitivity compared to conventional FAM probe-based methods.
Moving forward, this developed platform holds promise for the broad-spectrum detection of microRNAs, viruses, pathogenic bacteria, and proteins.
