Avidity-based bright and photostable light-up aptamers for single-molecule mRNA imaging

Tracking the Message: Applying Single Molecule Localization Microscopy to Cellular RNA Imaging

“…Their approach has enabled single‐molecule mRNA tracking in live cells while simultaneously maintaining a high signal‐to‐background ratio. Although this approach cannot be used for SMLM, these avidity‐based probes enable single‐molecule RNA imaging with a smaller tag size than protein‐based methods and have the ptoential for use in multiplex RNA imaging [83] . These proof of concept experiments have opened a new door for RNA labeling and aptamer development, holding strong promise for future SMLM studies of cellular RNA localization.…”

Harmonizing the growing fluorogenic RNA aptamer toolbox for RNA detection and imaging

“…113 This concept of engineering dimerization-based fluorogenic module provides new guides of engineering ultrabright fluorogenic aptamer-ligand system and is promising for the development of heterodimerization modules. Recently, an avidity-based dimeric aptamer named biRhoBAST 78 (Table 1, No. 67) has been reported that it binds towards a fluorogenic bivalent fluorophore TMR2 with K D = 40 pM.…”

RNA Condensate as a Versatile Platform for Improving Fluorogenic RNA Aptamer Properties and Cell Imaging

“…Cell imaging is a powerful technique for monitoring the abundance of molecules, their localization, dynamic changes, and cellular biological functions. , Biomolecular fluorescent reporters are essential components for developing cell imaging strategies . Due to their small size, high specificity and programmability, and low background fluorescence, fluorogenic RNA aptamers that selectively bind nonfluorescent small molecular fluorophores and activate their fluorescence have been successfully used as a kind of particularly attractive reporting unit for imaging diverse intracellular target analytes, , such as signaling molecules, metabolites, , and RNAs. − However, these fluorogenic RNA aptamer-based biosensing systems suffer from more or less pronounced shortcomings associated with easy degradation, limited photostability, and low fluorescence enhancement. , …”

“…9−11 However, these fluorogenic RNA aptamer-based biosensing systems suffer from more or less pronounced shortcomings associated with easy degradation, limited photostability, and low fluorescence enhancement. 12,13 RNA aptamer function and activity rely on structure and folding, which are heavily affected by the molecular microenvironment. 14 Inspired by protocells that confine functional RNA inside vesicles to stabilize the structure, promote folding, and improve activity, artificial molecular crowding systems have been constructed to improve RNA properties.…”