Journal
EUROPEAN BIOPHYSICS JOURNAL WITH BIOPHYSICS LETTERS
Volume 38, Issue 6, Pages 813-828Publisher
SPRINGER
DOI: 10.1007/s00249-009-0499-9
Keywords
Fluorescence correlation spectroscopy; Single-particle tracking; Promyelocytic leukemia nuclear body; Green fluorescent protein; Chromatin-remodeling complex; Protein dynamics in living cells
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Funding
- Volkswagenstiftung
- Deutsche Forschungsgemeinschaft [GRK 886/1, Ri 1283/5- 3]
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The spatial and temporal fluctuation microscope (STFM) presented here extends the concept of a fluorescence confocal laser scanning microscope to illumination and detection along a line. The parallel multichannel acquisition of the fluorescence signal was accomplished by using a single line of an electron-multiplying charge-coupled device camera at 14 mu s time resolution for detection of the fluorescence signal. The STFM system provided fast confocal imaging (30 images per second) and allowed for the spatially resolved detection of particle concentration fluctuations in fluorescence correlation spectroscopy experiments. For the application of the STFM, an approximated theoretical description of the beam geometry, the point-spread function, and the fluorescence auto- and cross-correlation functions were derived. The STFM was applied to studies of the dynamics of promyelocytic leukemia nuclear bodies, green fluorescent protein, and chromatin-remodeling complexes in living cells. The results demonstrate the unique capabilities of the STFM for characterizing the position-dependent translocations and interactions of proteins in the cell.
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