A general method to improve fluorophores for live-cell and single-molecule microscopy

By J. Grimm, Brian P. English, Jiji Chen, Joel P Slaughter, Zhengjian Zhang, A. Revyakin, Ronak Patel, J. Macklin, Davide Normanno, R. Singer and 2 more. Nature Methods, 2015

1,300 citations recorded by the source corpus.

Specific labeling of biomolecules with bright fluorophores is the keystone of fluorescence microscopy. Genetically encoded self-labeling tag proteins can be coupled to synthetic dyes inside living cells, resulting in brighter reporters than fluorescent proteins. Intracellular labeling using these techniques requires cell-permeable fluorescent ligands, however, limiting utility to a small number of classic fluorophores. Here we describe a simple structural modification that improves the brightness and photostability of dyes while preserving spectral properties and cell permeability. Inspired by molecular modeling, we replaced the N,N-dimethylamino substituents in tetramethylrhodamine with four-membered azetidine rings. This addition of two carbon atoms doubles the quantum efficiency and improves the photon yield of the dye in applications ranging from in vitro single-molecule measurements to super-resolution imaging. The novel substitution is generalizable, yielding a palette of chemical dyes with improved quantum efficiencies that spans the UV and visible range.

Subfields: Biophysics, Molecular Biology, Organic Chemistry

View Paper: https://www.semanticscholar.org/paper/a7a69b6fc714fbbae99644538cae8f3b92cabdea

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In Biophysics

1. R. Singer

In Molecular Biology

1. A. Senecal
2. Jeetayu Biswas
3. Maria Vera
4. H. Park
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6. L. Lavis
7. R. Tjian
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In Organic Chemistry

1. G. Lessene

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