Quantitative Imaging of Membrane Lipid Order in Cells and Organisms

Overview

Journal Nat Protoc

Specialties Biology
Pathology
Science

Date 2011 Dec 14

PMID 22157973

Citations 178

Authors

Dylan M Owen

Carles Rentero

Astrid Magenau

Ahmed Abu-Siniyeh

Katharina Gaus

Affiliations

Soon will be listed here.

Abstract

It is now recognized that lipids and proteins in cellular membranes are not homogenously distributed. A high degree of membrane order is the biophysical hallmark of cholesterol-enriched lipid rafts, which may induce the lateral sorting of proteins within the membrane. Here we describe a quantitative fluorescence microscopy technique for imaging localized lipid environments and measuring membrane lipid order in live and fixed cells, as well as in intact tissues. The method is based on the spectral ratiometric imaging of the polarity-sensitive membrane dyes Laurdan and di-4-ANEPPDHQ. Laurdan typically requires multiphoton excitation, making it suitable for the imaging of tissues such as whole, living zebrafish embryos, whereas di-4-ANEPPDHQ imaging can be achieved with standard confocal microscopes. This approach, which takes around 4 h, directly examines the organization of cellular membranes and is distinct from alternative approaches that infer membrane order by measuring probe partitioning or dynamics.

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