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Fluorescent proteins for single-molecule fluorescence applications

Britta Seefeldt1, Robert Kasper, Thorsten Seidel

  • 1Applied Laser Physics and Laser Spectroscopy, University of Bielefeld, Physics Department, Universitätsstrasse 25, 33615 Bielefeld, Germany.

Journal of Biophotonics
|April 4, 2009
PubMed

Insights

mCherry fluorescent protein is ideal for single-molecule studies due to its photostability and minimal intensity fluctuations, making it a valuable tool for advanced microscopy techniques.

Area of Science:

  • Biophysics
  • Molecular Biology
  • Microscopy

Background:

  • Fluorescent proteins are crucial tools in biological research.
  • Existing fluorescent proteins have limitations for advanced single-molecule applications.

Purpose of the Study:

  • To evaluate mCherry, a DsRed derivative, as a fluorophore for single-molecule fluorescence experiments.
  • To compare mCherry's photophysical properties with other common fluorescent proteins.

Main Methods:

  • Single-molecule fluorescence spectroscopy.
  • Ensemble fluorescence measurements.
  • Photostability and fluorescence intensity fluctuation analysis.

Main Results:

  • mCherry demonstrated high photostability and rare fluorescence-intensity fluctuations.
  • Despite lower quantum yield, mCherry's superior photophysics make it suitable for single-molecule applications.
  • mCherry has a fluorescence lifetime of 1.46 ns.

Conclusions:

  • mCherry is an excellent alternative for single-molecule fluorescence experiments.
  • Its spectral characteristics and short lifetime enable accurate tracking, localization, FRET, FLIM, and multicolor applications.

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