Format

Send to

Choose Destination
Biophys J. 2009 Mar 4;96(5):1999-2008. doi: 10.1016/j.bpj.2008.12.3888.

Accurate determination of membrane dynamics with line-scan FCS.

Author information

1
Technical University of Dresden, Biotechnologisches Zentrum, Dresden, Germany.

Abstract

Here we present an efficient implementation of line-scan fluorescence correlation spectroscopy (i.e., one-dimensional spatio-temporal image correlation spectroscopy) using a commercial laser scanning microscope, which allows the accurate measurement of diffusion coefficients and concentrations in biological lipid membranes within seconds. Line-scan fluorescence correlation spectroscopy is a calibration-free technique. Therefore, it is insensitive to optical artifacts, saturation, or incorrect positioning of the laser focus. In addition, it is virtually unaffected by photobleaching. Correction schemes for residual inhomogeneities and depletion of fluorophores due to photobleaching extend the applicability of line-scan fluorescence correlation spectroscopy to more demanding systems. This technique enabled us to measure accurate diffusion coefficients and partition coefficients of fluorescent lipids in phase-separating supported bilayers of three commonly used raft-mimicking compositions. Furthermore, we probed the temperature dependence of the diffusion coefficient in several model membranes, and in human embryonic kidney cell membranes not affected by temperature-induced optical aberrations.

PMID:
19254560
PMCID:
PMC2717290
DOI:
10.1016/j.bpj.2008.12.3888
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center