Jump to main content
Jump to site search

Issue 2, 2010
Previous Article Next Article

Higher resolution in localization microscopy by slower switching of a photochromic protein

Author affiliations

Abstract

Photoswitchable fluorophores play an essential role in super-resolution fluorescence microscopy, including techniques such as photoactivated localization microscopy (PALM). A determining factor in the precision of the images generated by PALM measurements is the photon numbers that can be detected from the fluorophores. Dronpa is a reversibly photoswitchable fluorescent protein that has been successfully used in PALM experiments. The number of photons per switching cycle that can be acquired for Dronpa depends on its off-switching rate, limiting the number of photons that can be recorded. In this study we report our discovery that the tetrameric ancestor of Dronpa, 22G, shows slower switching, and develop a mutant that displays switching kinetics between those of Dronpa and 22G. We show that the kinetics of the photoswitching are strongly related to self-association of the protein, supporting our view of dynamic flexibility as determining in the photoswitching. Similarly we find that higher-resolution PALM images can be acquired with slower-switching proteins due to their higher number of emitted photons per switching cycle.

Graphical abstract: Higher resolution in localization microscopy by slower switching of a photochromic protein

Back to tab navigation

Publication details

The article was received on 06 Oct 2009, accepted on 11 Nov 2009 and first published on 18 Jan 2010


Article type: Paper
DOI: 10.1039/B9PP00124G
Citation: Photochem. Photobiol. Sci., 2010,9, 239-248
  •   Request permissions

    Higher resolution in localization microscopy by slower switching of a photochromic protein

    H. Mizuno, P. Dedecker, R. Ando, T. Fukano, J. Hofkens and A. Miyawaki, Photochem. Photobiol. Sci., 2010, 9, 239
    DOI: 10.1039/B9PP00124G

Search articles by author

Spotlight

Advertisements