Cell-surface proteins display non-random distribution patterns ranging in size from a few nanometers to microns. Here, we describe some widely used fluorescence-based biophysical methods that can be used to study the different hierarchical levels of the lateral organization and interactions of membrane proteins. Classical biochemical and immunological methods (chemical cross-linking, co-immu-noprecipitation, detergent resistance analysis, etc.) provide valuable information on the interaction of membrane proteins, but have several drawbacks. First, with these methods proteins cannot be studied in their native environment. Second, the use of conventional extraction and isolation procedures inherent to these techniques may also disrupt protein-protein interactions, or may induce the formation of artificial protein aggregates. These methods provide information only on the bulk properties of interactions, without resolving cell-to-cell or subcellular variations. On the other hand, fluorescence-based observations of appropriately labeled molecules allow the detection of interactions with a subcellular resolution. Fluorescence detection is sensitive (down to the level of single molecules under select conditions), relatively non-destructive, and specific due to the high affinity of interaction between commonly used markers (specific antibodies, toxins, etc.) and their targets. Genes tailored to code for proteins carrying a fluorescent tag (GFP and its spectral variants) are routinely used to report on the subcellular distribution of proteins.
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This ebook provides an introductory explanation of the workings of the human body, with an effort to draw connections between the body systems and explain their interdependencies. A framework for the book is homeostasis and how the body maintains balance within each system. This is intended as a first introduction to physiology for a college-level course.