Syllabus
1. Why electrons?
2. How does a transmission electron microscope work?
3. How does a scanning electron microscope work?
4. Methods of biological specimen preparation at room temperature.
5. How to cut section with the thickness below 100 nm?
6. How to vitrify water in biological samples?
7. Methods of biological sample preparation for SEM.
8. Cellular ultrastructure and interpretation of electron-microscopic images.
9. Immunolabeling methods – the tool how to localize and identify molecules in cell structure.
10. Volume electron microscopy – from 2D to 3D imaging.
11. How to visualise macromolecules in TEM?
12. How to record electron microscopic images and how to process them.
Annotation
The course is focused on explaining how electron microscopes work and how can be used for the observation of biological objects. The course can be divided into three parts: 1/ physical principles of electron microscopy and description of instrumentation: Properties of accelerated electrons, basic components of electron microscopes – sources of primary electrons, lenses and their aberrations, the interaction of primary electrons with specimen atoms, formation of images in TEM and SEM, construction of TEM and SEM. 2/ specimen preparation of biological objects for TEM and SEM: Chemical methods (fixation, dehydration, embedding), cutting of ultrathin sections, cryo-methods (high-pressure freezing, plunge freezing, cryo-cutting, freeze substitution), special methods for SEM (critical point drying, freeze fracturing, freeze-drying, methods of specimen coating by conductive layer), cellular ultrastructure and interpretation of electron-microscopic images. 3/ advanced methods of current biological electron microscopy: high-resolution electron microscopy in biology (SPA), immunolocalization of molecules of interest in cell ultrastructure, volume electron microscopy (electron tomography, dual-beam SEM, serial block-face SEM, micro-array tomography), data recording and processing.
The course consists of a theoretical part (lectures) and several practical tasks.
Recommended literature:
MJ Dykstra, LE Reuss: Biological Electron Microscopy – Theory, Techniques, and Troubleshooting. 2nd Edition. Kluwer Academic Plenum Publisher. ISBN 0-306-47749-1. 2003 https://myscope.training/: an open online platform for those who ho want to learn about microscopy. The platform provides insights into the fundamental science behind different microscopes, explores what can and cannot be measured by different systems, and provides a realistic operating experience on high-end microscopes.