Abstract
Heavy metal shadowing is one of the oldest and most useful methods of enhancing the contrast of biological molecules for visualization by electron microscopy. A critical step in this method is the dehydration of the specimen prior to metal shadowing without causing distortion and collapse of the structure and consequent loss of the three-dimensional information. Two methods commonly employed today are glycerol spraying/vaccuum evaporation and freeze-drying. While glycerol spraying is the easier method of the two, it does not significantly prevent collapse artifact during air-drying and thus is limited to single extended molecules. Freeze-drying minimizes the collapse artifact of flattening and distortion due to surface tension. Freeze-drying preparations suffer their own problems of thermal vibrations and possible contamination by a eutectic of remaining salts and proteins.
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