The conventional sample preparation for electron microscopy makes the artifacts in the process of dehydration and substitution, such as the changes of ultrastructure of cells and outflow of soluble components in the cells. Cryofixation has been proposed as the method to overcome the inconvenient and unexpected problems, by freezing the cells very rapidly. Because water freezes to be ice crystals with increase of the volume, and those formed in the cells destroy the ultrastructure. To preserve

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... e ultrastructure of native cells at the resolution of electron microscopic level, it is required to make amorphous ice at the process of cryofixation. As the methods for it, rapid freezing to cool samples rapidly in normal pressure and high pressure freezing to freeze under 2100 bar have been proposed. In this review, the information of the latest high pressure freezing systems and their application has been introduced. The depth of good freezing in amorphous ice made by rapid freezing is 5 to 20 μm, on the other hand, that of high pressure freezing is about 200 μm. So the high pressure freezing can be applied for the electron microscopic analyses of cells and tissues at closed-to-native state. Workflows after cryofixation are, for example, freeze substitution and cryo-TEM observation of vitreous ice sections, or CEMOVIS. Furthermore, the surface of cryo-sectioned frozen samples has been observed by cryo-SEM.