Adult tissue dissection, fixation and resin embedding were performed as described (Wolff, 2000).
A combination of perfusion- and cryo-fixation with resin embedding was used to achieve an optimized balance between high labeling efficiency as obtained by the Tokuyasu cryo immuno-gold labeling technique and ultrastructural visualization as obtained by standard Epon embedding (van Lookeren et al., 1991; Verkade et al., 1995).
Hyphal tissue organization was observed from resin fixation and sectioning of basidiomata.
Examples are provided from samples of cultured cells prepared by high-pressure freezing and freeze-substitution as well as by chemical fixation before epoxy resin embedding.
Among them, the ultrathin-sectioning with chemical fixation, embedding in resin, and chemical staining provides good contrast for electron microscopy imaging.
We thank Wann-Neng Jane, Academia Sinica, for resin fixation and sectioning of the mushroom samples, Jia-Ling Wei, National Museum of Natural Science, Taiwan, for helping collect G. epiphylla, and Huei-Mei Hsieh, Academia Sinica, for assistance in obtaining ITS sequences.
Conventional electron microscopy, involving chemical fixation and embedding in resin, is still an excellent method for visualization of the membrane and cytoskeletal elements of cell-cell contacts.
The processing steps from post fixation to polymerization of resin blocks were carried out in a microwave oven, Pelco Bio Wave 34770 (Pelco International, Clovis, CA, USA) using similar procedures but with a slight modification as recommended by the manufacturer.
Positive staining depends on chemical reactivity with the components of the object and involves fixation, postfixation, embedding in resins, ultrathin sectioning, and multiple staining incubations.
Volume fraction of GNS EMO in composites is given for an optimal quantity of resin epoxy in fixation and formation.
