Cookies on this website
We use cookies to ensure that we give you the best experience on our website. If you click 'Continue' we'll assume that you are happy to receive all cookies and you won't see this message again. Click 'Find out more' for information on how to change your cookie settings.

We describe an optical method capable of tracking a single fluorescent molecule with a flexible choice of high spatial accuracy (approximately 10-20 nm standard deviation or approximately 20-40 nm full-width-at-half-maximum) and temporal resolution (< 1 ms). The fluorescence signal during individual passages of fluorescent molecules through a spot of excitation light allows the sequential localization and thus spatio-temporal tracking of the molecule if its fluorescence is collected on at least three separate point detectors arranged in close proximity. We show two-dimensional trajectories of individual, small organic dye labeled lipids diffusing in the plasma membrane of living cells and directly observe transient events of trapping on < 20 nm spatial scales. The trapping is cholesterol-assisted and much more pronounced for a sphingo- than for a phosphoglycero-lipid, with average trapping times of approximately 15 ms and < 4 ms, respectively. The results support previous STED nanoscopy measurements and suggest that, at least for nontreated cells, the transient interaction of a single lipid is confined to macromolecular dimensions. Our experimental approach demonstrates that fast molecular movements can be tracked with minimal invasion, which can reveal new important details of cellular nano-organization.

Original publication

DOI

10.1073/pnas.0912894107

Type

Journal article

Journal

Proc Natl Acad Sci U S A

Publication Date

13/04/2010

Volume

107

Pages

6829 - 6834

Keywords

Animals, Cell Membrane, Diffusion, Kidney, Membrane Lipids, Membrane Microdomains, Microscopy, Confocal, Potoroidae, Probability, Reproducibility of Results, Signal Processing, Computer-Assisted, Time Factors