Single‐Molecule Studies of Biomolecules

Abstract

Imaging biological processes at the single‐molecule scale enables the direct observation of the temporal and spatial dynamics of cellular processes. This relatively new branch of biophysical investigation, which was founded on a platform of physics and chemistry research, is pushing forward the frontiers of medical science by providing unprecedented new illuminations into the molecular basis of life. Although it is far from comprehensive, this article provides the advanced reader with a review of single‐molecule methods, as well as a discussion of the aims, considerations, achievements, and future directions of this field in the context of ongoing investigations of structure–function relationships inherent to biomolecules.

Xanthene, cyanine, oxazine and BODIPY: the four pillars of the fluorophore empire for super-resolution bioimaging

“…For the initial implementation of SMLM, cyanine dyes were used overwhelmingly by the researchers as this fluorescent dye system possessed good water solubility and easy functionalization aptitude (for attaching to the biomolecules) besides showing outstanding photo-switching properties. 23,[157][158][159][160][161][162] It will not be an exaggeration to say that among the small moleculebased fluorophores for SMLM studies, cyanine dyes are the most prevalent class of fluorescent probes other than rhodamine dyes. Commonly, the thiol (BME and MEA) imaging buffers, containing enzymatic oxygen scavengers are used in imaging practices to expedite the fluorescence blinking/photo-switching behaviour in cyanine dyes, so that the fluorophores could endure repetitive transitions between fluorescent and nonfluorescent forms until they become fully photobleached.…”