High-resolution SEM of biological macromolecular complexes

For HRSEM imaging of insulating specimens, thin “homogeneous” metal films with a thickness smaller than the SE-escape depth (1-2 nm for metals) and with a neglectable or very low BSE-coefficient (low Z e.g. Cr or Ge), continuous enveloping the surface fine structures, are supposed to reveal a high resolution topographic SE-1 signal (Joy 1984). Hermann and Müller (1991) have reported a resolution of about 3-4 nm on freeze-dried thin biological specimens, coated with Cr at low temperature (−80°C). High resolution SE-images were also shown on ft or W sputter coated, frozen-hydrated yeast cells (Walther et al., 1992 and Hermann and Müller, 1993) and on coated, freeze-dried macromolecular structures (Walther et al. 1992; Ya Chen et al. 1993). To study the obtainable resolution and the extractable surface information in SEM from a variety of coating techniques and metals, thin specimens (<20nm) were used as test specimens in order to reduce the signal contribution of the background (SEII).