The application of scanning electron microscopy to the study of the cytoskeleton of cells in culture.
This paper reviews the methods available for studying the structural organization of the cytoskeleton of cells in culture. These are transmission electron microscopy of whole cells, immunofluorescence, and detergents extraction procedures. Published methods for detergent extraction are critically reviewed with special emphasis on those factors that are most relevant for the preparation of cytoskeletons for observation by scanning electron microscopy. These include the type of detergent, the use of chemical crosslinking reagents or specially formulated buffers to stabilize the cytoskeleton during extraction, the osmotic and salt composition of the extraction medium, the use of cool sputter coaters, and the application of minimal amounts of metal during coating. Finally, examples are given of the types of information about the three dimensional organization of the cytoskeleton that can be obtained by SEM.
D000818:Animals; D002021:Buffers; D002458:Cell Fractionation; D002478:Cells, Cultured; D003584:Cytological Techniques; D003599:Cytoskeleton / Q000648:ultrastructure*; D003890:Desiccation; D003902:Detergents; D005404:Fixatives; D006863:Hydrogen-Ion Concentration; D008854:Microscopy, Electron; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000648:ultrastructure*; D013048:Specimen Handling / Q000379:methods; D013194:Staining and Labeling; D013696:Temperature
D002021:Buffers; D003902:Detergents; D005404:Fixatives