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Fluorescence Correlation Spectroscopy Diffusion Laws to Probe the Submicron Cell Membrane Organization

Laure Wawrezinieck ^* , Hervé Rigneault ^*, Didier Marguet  and Pierre-François Lenne ^*

^* Institut Fresnel, MOSAIC Group, CNRS UMR 6133-Université Paul Cézanne Aix-Marseille III, Domaine Universitaire de Saint Jérôme, F-13397 Marseille Cedex 20, France; and Centre d'Immunologie de Marseille-Luminy, MOSAIC group, CNRS UMR 6102-INSERM UMR 631-Université de la Méditerranée, Parc Scientifique de Luminy, Case 906, F-13288 Marseille Cedex 9, France

Correspondence: Address reprint requests to Pierre-François Lenne, Institut Fresnel, MOSAIC Group, CNRS UMR 6133-Université Paul Cézanne Aix-Marseille III, Domaine Universitaire de Saint Jérôme, F-13397 Marseille Cedex 20, France. Tel: 33-491-28-8049; Fax: 33-491-28-8067; E-mail: lenne{at}fresnel.fr.

To probe the complexity of the cell membrane organization and dynamics, it is important to obtain simple physical observables from experiments on live cells. Here we show that fluorescence correlation spectroscopy (FCS) measurements at different spatial scales enable distinguishing between different submicron confinement models. By plotting the diffusion time versus the transverse area of the confocal volume, we introduce the so-called FCS diffusion law, which is the key concept throughout this article. First, we report experimental FCS diffusion laws for two membrane constituents, which are respectively a putative raft marker and a cytoskeleton-hindered transmembrane protein. We find that these two constituents exhibit very distinct behaviors. To understand these results, we propose different models, which account for the diffusion of molecules either in a membrane comprising isolated microdomains or in a meshwork. By simulating FCS experiments for these two types of organization, we obtain FCS diffusion laws in agreement with our experimental observations. We also demonstrate that simple observables derived from these FCS diffusion laws are strongly related to confinement parameters such as the partition of molecules in microdomains and the average confinement time of molecules in a microdomain or a single mesh of a meshwork.

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