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We use whole mounts of detergent\u2010extracted cells grown on Formvar\u2010coated gold grids as a model system to evaluate various steps in the preparation of biological samples for high resolution scanning electron microscopy (SEM)<\/jats:p><jats:p>Factors that are important in determining the structure and composition of detergent\u2010extracted cells include the nature of the detergent and the composition of the extraction vehicle. Chelation of calcium is extremely important to stabilize and preserve the cytoskeletal filaments. We have also demonstrated both morphologically and by gel electrophoresis that treatment of cells with bifunctional protein crosslinkers before or during extraction with detergent can significantly enhance the preservation of both proteins and supramolecular structures.<\/jats:p><jats:p>The methods used to dry samples are a major determinant of the quality of structural preservation. For cytoskeletons freeze\u2010drying (FD) is superior to critical point\u2010drying (CPD), one reason being that CPD samples have to be dehydrated, thereby causing more shrinkage as compared to FD samples. The high pressures to which samples are exposed during CPD may also cause increased shrinkage, and water contamination during CPD causes severe structural damage. We have obtained the best structural preservation of detergent\u2010extracted and fixed cells by manually plunging them into liquid propane and drying over night in a freeze\u2010drayer.<\/jats:p><jats:p>The factor that most limits achievement of high resolution in SEM is the metal coat, which has to be very thin, uniform, and free of grain in order not to hide structures or to create artifactual ones. We have found that sputter\u2010coating with 1\u20133 nm of tungsten (W) or niobium )Nb( gives extremely fine\u2010grained films as well as satisfactory emission of secondary electrons. These samples can also be examined at high resolution by transmission electron microscopy (TEM) and scanning transmission electron microscopy (STEM). The best preservation and visualization of supramolecular structures have been obtained using cryosputtering, in which the samples are freeze\u2010dried and then sputter\u2010coated within the freeze\u2010dryer while still frozen. \u00a9 1992 Wiley\u2010Liss, Inc.<\/jats:p>","DOI":"10.1002\/jemt.1070220203","type":"journal-article","created":{"date-parts":[[2005,2,23]],"date-time":"2005-02-23T17:43:02Z","timestamp":1109180582000},"page":"130-150","source":"Crossref","is-referenced-by-count":24,"title":["Preservation and visualization of molecular structure in detergent\u2010extracted whole mounts of cultured cells"],"prefix":"10.1002","volume":"22","author":[{"given":"Margaretha","family":"Lindroth","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"suffix":"Jr.","given":"Paul B.","family":"Bell","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bengt\u2010Arne","family":"Fredriksson","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiao\u2010Dong","family":"Liu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2005,2,4]]},"reference":[{"key":"e_1_2_1_2_1","first-page":"159","article-title":"High resolution SEM of cultured cells: Preparatory procedures","author":"Arro E.","year":"1981","journal-title":"Scanning Electron Microsc."},{"key":"e_1_2_1_3_1","doi-asserted-by":"publisher","DOI":"10.1021\/bi00551a041"},{"key":"e_1_2_1_4_1","first-page":"139","article-title":"The application of scanning electron microscopy to the study of the cytoskeleton of cells in culture","author":"Bell P. 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