Integrated ultramicrotome for serial block face scanning electron microscopy (SBF-SEM)

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Recent technical developments in biomedical electron microscopy (EM) aim at visualizing structures (e.g. cells or tissue samples) in their topographic context at high resolution in 3D (volume EM). The analysis of large sample volumes allows to reveal biological "connectomes" in their spatial complexity.We apply for an ultramicrotome, integrated into the column of a scanning electron microscope, for automated 3D analyses by serial block face scanning electron microscopy (SBF-SEM). The system will complement a scanning electron microscope with field emission cathode (FEG-SEM) operated since 2019 by the Core Facility Electron Microscopy (CFEM) of Charité - Universitätsmedizin Berlin. The ultramicrotome is mounted on a table and can be integrated via the front of the FEG-SEM column. By exchange of the table, the FEG-SEM can either be operated in conventional mode or in SBF-SEM mode.An essential approach for generating 3D image datasets in SEM is based on the principle of mechanical removal of thin slices from the sample surface, alternating to imaging the sample surface after each removal. In SBF-SEM, this removal from the sample surface is realized by the principle of ultramicrotomy (i.e. with a diamond knife). After proper alignment, the acquisition of raw data runs automatically. The maximum spatial resolution is about 10-15 (x/y) and 25-50 nm (z), respectively. The maximum sample volume is in the order of about 1 mm (z) over an area of about 0.5 x 0.5 mm (x/y).The microscope, upgraded for SBF-SEM investigations, will be made available to all interested users according to the user policy of the CFEM of Charité - Universitätsmedizin Berlin. Furthermore, it will be embedded into the research infrastructure of the Berlin University Alliance (BUA).