SPECTROSCOPY, IMAGING, OTHER TECHNIQUES |
Total Internal Reflection Fluorescence Correlation Spectroscopy: Effects of Lateral Diffusion and Surface Generated Fluorescence
Jonas Ries 1, Eugene P. Petrov 2 and Petra Schwille 1*
1 TU Dresden
2 Biophysics / BIOTEC
* To whom correspondence should be addressed. E-mail: schwille{at}biotec.tu-dresden.de.
Submitted on November 27, 2007
Revised on January 22, 2008
Accepted on 26 February 2008
 |
Abstract |
|---|
Fluorescence Correlation Spectroscopy with Total Internal Reflection excitation (TIR-FCS) is a promising method with emerging biological applications for measuring binding dynamics of fluorescent molecules to a planar substrate as well as diffusion coefficients and concentrations at the interface. Models for correlation functions proposed so far are rather approximate for most conditions, since they neglect lateral diffusion of fluorophores. Here we propose accurate extensions of previously published models for axial correlation functions taking into account lateral diffusion through detection profiles realized in typical experiments. In addition, we consider the effects of surface-generated emission in objective-based TIR-FCS. The expressions for correlation functions presented here will facilitate quantitative and accurate measurements with TIR-FCS.
Key Words:
Binding, Diffusion, FCS, Membranes, TIRF
