Akira Kitamura, Masataka Kinjo
"Determination of diffusion coefficients in live cells using fluorescence recovery after photobleaching with wide-field fluorescence microscopy"
Biophysics and Physicobiology, Vol. 15, pp. 1-7 (2018)
Fluorescence recovery after photobleaching (FRAP), that is widely used for analysis of molecular dynamics, provides quantitative determination of static and dynamic properties of molecules of interest. In order to investigate diffusion states of rapidly-diffusing fluorescent molecules such as green fluorescent protein (GFP) monomers and tandemly oligomerized GFPs in living cells, the authors proposed a straightforward procedure for measuring diffusion coefficients using FRAP with an epi-fluorescence microscope. In that procedure, the apparent spreading of the effective radius of the photobleached area works as a correction factor. The measurement procedure and the data quality are described in details comparing the results by the conventional fluorescence correlation spectroscopy (FCS). They conclude that the method can be used to measure accurate diffusion coefficients of almost any fluorescently-labeled fusion proteins in living cells without using FCS. This technique has been widely applied in the world to measure the diffusion coefficients of many kinds of molecules, and the article has been frequently cited as one of the original papers in this fields.
Thus, it is an excellent article, contributing to analysis of molecular dynamics as the general method applied in a variety of fields, not only molecular cell biology but also developmental biology and chemical biology.
June, 2022
Biophysics and Physicobiology Award Selection Committee