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[01058] Microscopy - Electron microscopy; [10504] Biophysics - Methods and techniques; [10506] Biophysics - Molecular properties and macromolecules; [10511] Biophysics - Bioengineering; [30500] Morphology and cytology of bacteria; [32000] Microbiological apparatus, methods and media
Abstract:
Electron microscope control, that allows the automatic recording of tilt series for the 3D reconstruction of invididual objects, has been realized. The experimental set-up includes a 200 kV TEM equipped with a 1K .times. 1K CCD camera, both controlled externally by a fast dedicated image-processing computer. For the goniometer control an accurate electronic readout of the tilt angle and a board driving the goniometer motor have been installed. For low-dose imaging, three to five different specimen areas are used: one (or two) for the determination of object displacements during tilting, one (or two) for autofocusing, and another one for recording the tilt series to be used for the 3D reconstruction. Tilt series can be recorded with a rather low total dose, the lower limit being set by the requirement that subsequent projection images have to be aligned by means of cross-correlation functions. The method has been tested with graphitized carbon particles on carbon film and with negative stained proteasomes from the archaebacterium Thermoplasma acidophilum. Some future developments towards fully automatic electron tomography are discussed.
