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Chem Biol. 2011 Oct 28;18(10):1241-51. doi: 10.1016/j.chembiol.2011.08.007.

Rational design of photoconvertible and biphotochromic fluorescent proteins for advanced microscopy applications.

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1
Laboratory of Photochemistry and Spectroscopy, Department of Chemistry, Katholieke Universiteit Leuven, 3001 Heverlee, Belgium. virgile.adam@ibs.fr

Abstract

Advanced fluorescence imaging, including subdiffraction microscopy, relies on fluorophores with controllable emission properties. Chief among these fluorophores are the photoactivatable fluorescent proteins capable of reversible on/off photoswitching or irreversible green-to-red photoconversion. IrisFP was recently reported as the first fluorescent protein combining these two types of phototransformations. The introduction of this protein resulted in new applications such as super-resolution pulse-chase imaging. However, the spectroscopic properties of IrisFP are far from being optimal and its tetrameric organization complicates its use as a fusion tag. Here, we demonstrate how four-state optical highlighting can be rationally introduced into photoconvertible fluorescent proteins and develop and characterize a new set of such enhanced optical highlighters derived from mEosFP and Dendra2. We present in particular NijiFP, a promising new fluorescent protein with photoconvertible and biphotochromic properties that make it ideal for advanced fluorescence-based imaging applications.

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PMID:
22035793
DOI:
10.1016/j.chembiol.2011.08.007
[Indexed for MEDLINE]
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