Ultramicrotomy is widely regarded as a thin section preparation method for transmission electron microscopy (TEM) investigations. It is shown that ultramicrotomy can also provide a simple path for microstructure analysis and assessment of mechanical properties for a sectioned block-face. Furthermore, electron backscatter diffraction (EBSD) analysis can be applied directly on ultramicrotomed surfaces without any additional polishing or etching. EBSD analysis relates the inherent cutting artefacts to the crystallographic orientations of the grains, hence delivering a rough assessment of their deformation resistance. TEM investigations revealed that crystallographic-related cutting artefacts, which exhibit a wave-like pattern, are the result of the dislocation pile-ups close to the knife-specimen interface. This technique is considered suitable to be coupled with EBSD for three-dimensional microstructure reconstructions when used for serial sectioning of large volumes.@en

Cryo-ultramicrotomy can be used to obtain ultrathin cryo-sections from cryo-fixed or aldehyde-fixed cryo-protected vitreous biologic samples. For immuno-gold EM, cryo-sections are retrieved from the cryo-chamber on a droplet of a pick-up solution (paste-like and almost frozen) to which the sections attach. The sections are then placed on an EM specimen grid at room temperature. This procedure compromises the ultrastructure, resulting in folds, holes, and loss of the original material. In this paper we show the critical influence of humidity, stretching, and relief of compression during thawing of the sections. We show a new lift-up hinge device for semi-automated retrieval of cryo-sections that results in significantly improved section quality. This approach was also applied successfully to vitreous sections from high pressure frozen samples. An important advance is that these vitreous cryo-sections can now successfully be post-fixed and immunolabelled after thawing: this allows cryo-EM comparison with adjacent ribbons of sections still in the frozen hydrated state. These findings call for technical innovations aiming at automated cryo-ultramicrotomy in a fully controlled environment for improved localization of proteins within their 'close to native' cellular context and correlative electron cryo-tomography of consecutive ribbons of sections of one frozen hydrated sample@en