Research ResourceBiochemistry

Rapid multiplex analysis of lipid raft components with single-cell resolution

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Sci. Signal.  22 Sep 2015:
Vol. 8, Issue 395, pp. rs11
DOI: 10.1126/scisignal.aac5584

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Lipid raft analysis by flow cytometry

Lipid rafts are dynamic regions of membranes that are involved in cell signaling but are challenging to study because of their small size and dynamic nature. Schatzlmaier et al. found that lipid raft components became associated with nuclei during lysis of cells as the cells passed through a detergent-containing layer in a gradient. In experiments with lymphocytes, the authors demonstrated that this association enabled the quantitative analysis by flow cytometry of the composition of lipid rafts and of the dynamic association of proteins with these membrane microdomains at single-cell resolution.

Abstract

Lipid rafts, a distinct class of highly dynamic cell membrane microdomains, are integral to cell homeostasis, differentiation, and signaling. However, their quantitative examination is challenging when working with rare cells, developmentally heterogeneous cell populations, or molecules that only associate weakly with lipid rafts. We present a fast biochemical method, which is based on lipid raft components associating with the nucleus upon partial lysis during centrifugation through nonionic detergent. Requiring little starting material or effort, our protocol enabled the multidimensional flow cytometric quantitation of raft-resident proteins with single-cell resolution, thereby assessing the membrane components from a few cells in complex cell populations, as well as their dynamics resulting from cell signaling, differentiation, or genetic mutation.

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