Fluorescence correlation spectroscopy for the study of membrane dynamics and protein/lipid interactions.
Fluorescence correlation spectroscopy (FCS) is a powerful technique to study dynamic biomolecular processes. It allows the estimation of concentrations, diffusion coefficients, molecular interactions, and other processes causing fluctuations in the fluorescence intensity, thus yielding information about aggregation processes, enzymatic reactions, or partition coefficients. During the last years, FCS has been successfully applied to model and cellular membranes, proving to be a promising tool for the study of membrane dynamics and protein/lipid interactions. Here we describe the theoretical basis of FCS and some practical implications for its application in membrane studies. We discuss sources of potential artifacts, such as membrane undulations, positioning of the detection volume, and photobleaching. Special attention is paid to aspects related to instrumentation and sample preparation as well as data acquisition and analysis. Finally, we comment on some strategies recently developed for the specific improvement of FCS measurements on membranes.
García-Sáez
Ana J
AJ
Department of Biotec, Biophysics group, Biotechnologisches Zentrum, Tatzberg 47-51, TU Dresden, 1307 Dresden, Germany.
Schwille
Petra
P
D005456:Fluorescent Dyes; D008563:Membrane Lipids / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods; D013050:Spectrometry, Fluorescence / Q000379:methods*
D005456:Fluorescent Dyes; D008563:Membrane Lipids; D008565:Membrane Proteins