Highly Sensitive Hybridization Chain Reaction-Based miRNA Detection Technology Using Diffusivity Analysis of Fluorescent Probe-Modified miRNA Particles

MicroRNAs (miRNAs) are promising biomarkers for the early detection of various diseases, particularly cancer, driving active development of highly sensitive and selective detection technologies. This study aims to establish a novel miRNA detection technique that utilizes image analysis to track the Brownian motion (diffusivity) of fluorescent probe-modified miRNA particles. This method identifies the presence and concentration of miRNAs by exploiting the change in particle size upon hybridization with the target. Furthermore, the use of a probe modified with a photo-crosslinkable artificial nucleic acid (CNV-D) enables the covalent capture of the target miRNA, ensuring high selectivity in biological samples even under stringent washing conditions. By integrating Hybridization Chain Reaction (HCR), the complex size is significantly amplified, dramatically enhancing the detection sensitivity. Consequently, we successfully demonstrated the highly sensitive and specific detection of the cancer biomarker miR-21 in serum, achieving an exceptionally low limit of detection (LOD) of 1 fM. This technology holds great potential to contribute to the early diagnosis of cancer.