Document Detail


Dual-color fluorescence cross-correlation spectroscopy with continuous laser excitation in a confocal setup.
MedLine Citation:
PMID:  23276535     Owner:  NLM     Status:  In-Data-Review    
Abstract/OtherAbstract:
Fluorescence correlation spectroscopy evaluates local signal fluctuations arising from stochastic movements of fluorescent particles in solution. The measured fluctuating signal is correlated in time and analyzed with appropriate model functions containing the parameters that describe the underlying molecular behavior. The dual-color extension, fluorescence cross-correlation spectroscopy (FCCS) allows for a comparison between spectrally well-separated channels to extract codiffusion events that reflect interactions between differently labeled molecules. In addition to solution measurements, FCCS can be applied with subcellular resolution and is therefore a very promising approach for a quantitative biochemical assessment of molecular networks in living cells. To derive thermodynamic and kinetic reaction parameters, the influence of a number of other factors like background noise, illumination intensity profiles, photophysical processes, and cross talk between the channels have to be treated. Here, we provide a roadmap to derive binding reaction data with dual-color FCCS using continuous wave laser excitation, as it is now accessible with many state-of-the-art confocal microscopes.
Authors:
Thomas Weidemann; Petra Schwille
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Methods in enzymology     Volume:  518     ISSN:  1557-7988     ISO Abbreviation:  Meth. Enzymol.     Publication Date:  2013  
Date Detail:
Created Date:  2013-01-01     Completed Date:  -     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  0212271     Medline TA:  Methods Enzymol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  43-70     Citation Subset:  IM    
Copyright Information:
Copyright © 2013 Elsevier Inc. All rights reserved.
Affiliation:
Biophysics/BIOTEC, Technische Universität Dresden, Tatzberg 47-51, Dresden, Germany. Electronic address: weidemann@biochem.mpg.de.
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