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Cell Biology and Cell Biophysics Programme, European Molecular Biology Laboratory, 69117 Heidelberg, Germany
Correspondence: Address reprint requests to Matthias Weiss, Cell Biology and Cell Biophysics Programme, European Molecular Biology Laboratory, Meyerhofstr. 1, 69117 Heidelberg, Germany. Tel.: +49-6221-387-408; Fax: +49-6221-387-512; E-mail: mweiss{at}embl-heidelberg.de.
We investigate the challenges and limitations that are encountered when studying membrane protein dynamics in vivo by means of fluorescence correlation spectroscopy (FCS). Based on theoretical arguments and computer simulations, we show that, in general, the fluctuating fluorescence has a fractal dimension D0 
1.5, which is determined by the anomality 
of the diffusional motion of the labeled particles, i.e., by the growth of their mean square displacement as (
x)2 
t. The fractality enforces an initial power-law behavior of the autocorrelation function and related quantities for small times. Using this information, we show by FCS that Golgi resident membrane proteins move subdiffusively in the endoplasmic reticulum and the Golgi apparatus in vivo. Based on Monte Carlo simulations for FCS on curved surfaces, we can rule out that the observed anomalous diffusion is a result of the complex topology of the membrane. The apparent mobility of particles as determined by FCS, however, is shown to depend crucially on the shape of the membrane and its motion in time. Due to this fact, the hydrodynamic radius of the tracked particles can be easily overestimated by an order of magnitude.
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