Single-molecule fluorescence spectroscopy and super-resolution microscopy are important elements of the ongoing technical revolution to reveal biochemical and cellular processes in unprecedented clarity and precision. Demands placed on the photophysical properties of the fluorophores are stringent and drive the choice of appropriate probes. Such fluorophores are not simple light bulbs of a certain color and brightness but instead have their own "personalities" regarding spectroscopic parameters, redox properties, size, water solubility, photostability, and several other factors. Here, we review the photophysics of fluorescent probes, both organic fluorophores and fluorescent proteins, used in applications such as particle tracking, single-molecule FRET, stoichiometry determination, and super-resolution imaging. Of particular interest is the thiol-induced blinking of Cy5, a curse for single-molecule biophysical studies that was later overcome using Trolox through a reducing/oxidizing system but a boon for super-resolution imaging owing to the controllable photoswitching. Understanding photophysics is critical in the design and interpretation of single-molecule experiments.

Taekjip Ha

Philip Tinnefeld

Annual Review of Physical Chemistry

自 1950 年出版的《物理化学年度评论》涵盖了物理化学领域的重大发展，包括生物物理化学、化学动力学、胶体、电化学、地球化学和宇宙化学、大气和气候化学、激光化学和超快过程，液态、磁共振、物理有机化学、聚合物和大分子等。

The Annual Review of Physical Chemistry, published since 1950, covers significant developments in the field of physical chemistry, including biophysical chemistry, chemical kinetics, colloid chemistry, electrochemistry, geochemistry and cosmochemistry, atmospheric and climate chemistry, laser chemistry and ultrafast processes, liquid state, magnetic resonance, physical organic chemistry, polymers, and macromolecules.

《Annual Review of Physical Chemistry》自1950年出版，涵盖了物理化学领域的重大进展，包括生物物理化学、化学动力学、胶体化学、电化学、地球化学和宇宙化学、大气与气候化学、激光化学与超快过程、液态物质、磁共振、物理有机化学、聚合物及大分子等多个方面。

Annu. Rev. Phys. Chem.

Photophysics of Fluorescent Probes for Single-Molecule Biophysics and Super-Resolution Imaging

The thiol-induced blinking of Cy5, a curse for single-molecule biophysical studies that was later overcome using Trolox through a reducing/oxidizing system but a boon for super-resolution imaging owing to the controllable photoswitching, is reviewed.

Category	Quartile
化学	1
化学, 物理化学	1

Photophysics of Fluorescent Probes for Single-Molecule Biophysics and Super-Resolution Imaging

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Journal Info

Category	Quartile
CHEMISTRY, PHYSICAL	1