Target-Mediated EV-SNA Clustering as an Amplifier for In Situ miRNA Imaging in Living Cells

within living cells. The miRDAS consists of two types of extracellular vesicle spherical nucleic acid (EV-SNA), named EVsnater and EVsnafier, which are generated by hydrophobic coassembly of natural EVs with cholesterol-modified oligonucleotides and exhibit rapid cellular uptake dynamics. Upon specifically binding to its target miRNA, the EVsnater activates the fluorescence and then self-assembles with EVsnafier to form giant fluorescent clusters, resulting in a brighter and significantly amplified fluorescence signal. Additionally, the miRDAS exhibits excellent flexibility and orthogonality, thus enabling simultaneous detection of multiple miRNA targets. As a proof of concept, a miR-124 miRDAS was designed and employed for successfully monitoring miR-124 expression levels during P19 neuronal differentiation.