Sample preparation and sectioning

Only embedded specimens can be cut sufficiently thinly for EELS and ESI from mycorrhizal material. Samples must therefore be fixed chemically or by freezing and then embedded conventionally or by freeze-substitution (Lehmann et al., 1990). From personal experience, only pre-fixation in glutaraldehyde-formaldehyde (Karnovsky, 1965), postfixation in osmium tetroxide and embedding using Spurr's procedure (1969) can be recommended for ectomycorrhiza (Kottke and Oberwink-ler, 1988). En bloc staining with uranyl acetate must be avoided otherwise high background noise will occur in EELS and ESI. Good cryofixation of mycorrhiza is extremely difficult to achieve and results in tremendous structural disturbance. It is therefore not advisable to use freeze-substitution followed by high resolution analysis in the TEM902.

The thin sections required for EELS and ESI can best be cut using a diamond knife. Because of their fragility these sections cannot be mounted on copper grids after dry sectioning. Sections have to be removed from the water trough and mounted on copper grids (600-700 mesh) immediately after sectioning to limit leaching of ions. They are analysed without any additional staining.

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