Abstract: The plasma membrane of mammalian cells is depicted as a two-dimensional hybrid material which is compartmentalized into submicron-sized domains. These membrane domains play a pivotal role in cellular signaling processes due to selective recruitment of specific cell surface receptors. The structural dynamics of the membrane domains and their exact biological functions are, however, still unclear, partially due to the wave nature of light, which limits the optical resolution in the visible light to approximately 400 nm in conventional optical microscopy. Here, we provide a non-fluorescence based approach for monitoring distance changes on subdiffraction limit length scales in a conventional far-field optical microscope. This approach, which is referred to as plasmon coupling microscopy (PCM), utilizes the distance dependent near-field coupling between noble metal nanoparticle (NP) labels to resolve close contacts on the length scale of approximately one NP diameter. We firstly utilize this PCM strategy to resolve interparticle separations during individual encounters of gold NP labeled fibronectin-integrin complexes in living HeLa cells. We then further refine this ratiometric detection methodology by augmenting it with a polarization-sensitive detection, which enables simultaneous monitoring of the distance and conformation changes in NP dimers and clusters. We apply this polarization resolved PCM approach to characterize the structural lateral heterogeneity of cell membranes on sub-micron length scales. Finally, we demonstrate that PCM can provide quantitative information about the structural dynamics of individual epidermal growth factor receptor (ErbB1)-enriched membrane domains in living cells.

Biological membranes play a central role in cell structure, shape and functions. However, investigating the membrane bilayer has proved to be difficult due to its highly dynamic and anisotropic structure, which generates steep gradients at the nanometer scale. Due to the decisive impact of recently developed fluorescence-based techniques, tremendous advances have been made in the last few years in our understanding of membrane characteristics and functions. In this context, the present book illustrates some of these major advances by collecting review articles written by highly respected experts. The book is organized in three parts, the first of which deals with membrane probes and model membranes. The second part describes the use of advanced quantitative and high-resolution techniques to explore the properties of biological membranes, illustrating the key progress made regarding membrane organization, dynamics and interactions. The third part is focused on the investigation of membrane proteins using the same techniques, and notably on the membrane receptors that play a central role in signaling pathways and therapeutic strategies. All chapters provide comprehensive information on membranes and their exploration for beginners in the field and advanced researchers alike.

· Measuring membrane protein distributions using single-molecule localisation microscopy (SMLM) · Measuring membrane protein dynamics and diffusion using fluorescence correlation spectroscopy (FCS) · Mapping membrane lipid backing using environmentally sensitive fluorescence probes · Mapping membrane thickness and rigidity using atomic force microscopy · Mapping membrane proteins and the cytoskeleton using electron microscopy

This book highlights recent advances in and diverse techniques for exploring the plasma membrane’s structure and function. It starts with two chapters reviewing the history of membrane research and listing recent advances regarding membrane structure, such as the semi-mosaic model for red blood cell membranes and the protein layer-lipid-protein island model for nucleated tissue cell membranes. It subsequently focuses on the localization and interactions of membrane components, dynamic processes of membrane transport and transmembrane signal transduction. Classic and cutting-edge techniques (e.g. high-resolution atomic force microscopy and super-resolution fluorescence microscopy) used in biophysics and chemistry are presented in a very comprehensive manner, making them useful and accessible to both researchers in the field and novices studying cell membranes. This book provides readers a deeper understanding of the plasma membrane’s organization at the single molecule level and opens a new way to reveal the relationship between the membrane’s structure and functions, making it essential reading for researchers in various fields.

Cell surface membranes have long been characterized as two-dimensional fluids whose mobile components are randomized by diffusion in the plane of the membrane bilayer. Recent research has indicated that cell surface membranes are highly organized and ordered and that important functional units of membranes appear as arrays of interacting molecules rather than as single, freely diffusing molecules. Mobility and Proximity in Biological Membranes provides an overview of the results obtained from biophysical methods for probing the organization of cell surface membranes. These results are presented in the context of detailed treatments of the theory and the technical demands of each of the methods. The book describes a versatile and easily applied mode for investigating molecular proximities in plasma membranes in a flow cytometer. Its analysis of lipid fluidity and viscosity of membranes and the rotational mobility of proteins offers intimate insight into the physical chemistry of biological membranes. The electrophysiology of lymphocytes is presented with focus on its importance in different diseases. New techniques are described, and new data, new possibilities, and future trends are presented by world experts. This book's chapters can serve both as guides to the existing literature and as starting points for new experiments and approaches associated with problems in membrane function.