Application of high‐resolution scanning electron microscopy to biological macromolecules

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Abstract
The development of ultrahigh‐resolution scanning electron microscopes (SEMs) has made the observation of biological macromolecules feasible, but adequate preparation methods have not yet been established. Although it has been possible to observe some molecules after they have been spread on a carbon substrate, this method has not proved suitable for other molecules which exhibit lower contrast, or are more susceptible to damage by the electron beam. In this study we have applied heavy‐metal impregnation methods using phosphotungstic acid, uranyl acetate, or osmium tetroxide mordanted by tannic acid. In addition, contamination due to the electron beam was reduced by improving the vacuum in the specimen chamber, and by the use of a heated specimen stage. Using these measures, haemocyanin, ferritin, apoferritin, thyro‐globulin and immunoglobulin M were successfully imaged. Ultrahigh‐resolution SEM seems likely to become an important means for studying the morphology of biological macromolecules.