Related Experiment Video
Updated: Feb 22, 2026
MicroRNA Amplification and Recognition through Locked-nucleic-acid In situ Hybridization as a Novel Detection and Quantification Method
Published on: October 7, 2025
Single-step, homogeneous and sensitive detection for microRNAs with dual-recognition steps based on luminescence
Dong Zhu1, Zhao Yi Miao2, Yue Hu2
1School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing, Jiangsu 210023, PR China; Jiangsu Key Laboratory for Functional Substance of Chinese Medicine State Key Laboratory Cultivation Base for TCM Quality and Efficacy, Nanjing 210023, PR China.
Abstract:
A single-step, homogeneous and sensitive LRET assay is presented for the detection of miRNAs. The amplification-free assay provides a unique combination of high specificity with dual-recognition approach of different hybridization and ligation steps and preventing background auto-fluorescence in biological samples using upconversion nanoparticles (UCNPs) as signal-producing nanoprobes. The assay probe is composed of signal-producing unit (a pair of homogeneous upconversion luminescence resonance energy transfer (UC-LRET)-based oligonucleotides) and recognition unit (two adaptor oligonucleotides). In the presence of target miRNAs, the probe and target miRNAs leads to the formation of stable double-strands and semi-stable adaptor-miRNAs complexes with an adaptor nick. Ligation of the nick using ligase cause the formation of stable double-strands, resulting in UCNPs-to-dye UC-LRET for detection of the miRNAs with near-infrared radiation (980nm). Sensitive detection of miRNA-21 at concentrations of 200pM to 1.4nM and detection limits of 0.095nM with good precision of 3.9% (RSD) for seven repeated measurements of 500pM miRNAs demonstrate the feasibility of both high throughput and point-of-care clinical diagnostics. The homogeneous UC-LRET assay without any washing can be extended to the application in other important types of nucleic acid analysis.
