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Analytical and Bioanalytical Chemistry

, Volume 387, Issue 1, pp 41–44 | Cite as

Single-molecule tracking in eukaryotic cell nuclei

  • Jan Peter Siebrasse
  • David Grünwald
  • Ulrich KubitscheckEmail author
Trends

Introduction

Visualization of single fluorescent molecules (single-molecule tracking, SMT) within cells provides a real-time molecular view of physiological processes in vivo [ 1, 2]. Prerequisites for optical intracellular single-molecule detection are the reduction of background fluorescence, optimization of light transmission in the microscope, utilization of laser light sources for excitation, and high-sensitivity and high-speed CCD camera systems for signal detection in combination with digital image processing (Fig.  1). Also, it is essential to use fluorescence labels of the highest photostability [ 3, 4], especially if autofluorescent proteins are employed [ 5, 6]. Microscopic imaging of single fluorescent molecules produces diffraction-limited light spots in the image plane. The corresponding intensity pattern can be well approximated by a two-dimensional Gaussian with a full width at half maximum of approximately 250 nm, when light with a wavelength of 500 nm and an objective...

Keywords

Splice Factor Fluorescence Correlation Spectroscopy Supramolecular Complex Single Fluorescent Molecule Small Nuclear Ribonucleoprotein Particle 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgements

U.K. gratefully acknowledges financial support by the Volkswagen Foundation (Grant I/78852) and the German Research Foundation (Grant Ku 1507/1-2).

Supplementary material

Movie 1

Single U1 snRNPs diffusing and binding within a living HeLa cell nucleus. Left hand side original but contrast-enhanced data. Right hand side movie data was filtered using a spot-enhancing filter (3-pixel diameter, see ref. [8]) to improve U1 snRNP visibility. The movie was recorded at 200 Hz, the total object field corresponded to an area of 12.2×12.2 μm2 (AVI 3.78 mb)

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Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Jan Peter Siebrasse
    • 1
  • David Grünwald
    • 1
  • Ulrich Kubitscheck
    • 1
    Email author
  1. 1.Department of Biophysical Chemistry, Institute for Physical and Theoretical ChemistryUniversity of BonnBonnGermany

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