Freeze-Fracture Techniques Applied to Biological Membranes

Abstract

The development and application of the freeze-fracture technique to membrane research (Moor and Mühlethaler, 1963; Mühlethaler et al., 1965) led to a better understanding of the complex structural arrangement of lipids and proteins. It is now used worldwide for topographical studies, being the only available method for producing a direct high-resolution image of large areas of membrane surfaces and internal fracture faces. At first, however, it was thought that freezing would cause phase transformation and structural rearrangement of the original membrane structure. This has been disproved with the help of indirect methods such as X-ray diffraction. As shown by Gulik-Krzywicki and Costello (1977), a perturbation of the molecular organization of hydrocarbon chains does not occur provided freeze quenching is carried out rapidly enough to avoid the formation of large ice crystals between lipid lamellae. Based on these results, much work has been carried out to improve the main preparative steps such as freezing, fracturing, and replication. In addition to these procedures, new methods for labeling membrane components, special devices for rapid quenching, handling procedures for split membranes, and elaborate image-processing procedures became available. Some of these achievements will be discussed in the following chapters.