Abstract
Scanning electron microscopy (SEM) can be used to image nuclear pore complex (NPC) surface structure of from a number of organisms and model systems. With a field emission SEM , this is a medium resolution technique where details of the organization of various components can be directly imaged. Some components, such as the NPC baskets and cytoplasmic filaments, are difficult to visualize in any other way. Protein components can be identified by immunogold labeling. Any surface that can be exposed can potentially be studied by SEM . Several overlapping protocols for SEM sample preparation and immunogold labeling of NPCs are given here. Various parameters for sample preparation, fixation, immunogold labeling, drying, metal coating, and imaging are detailed which have been optimized for different types of specimens and desired endpoints.
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Acknowledgments
Thanks to Christine Richardson and Helen Grindley for technical support; the Biotechnology and Biological Sciences Research Council, UK, grant number BB/E015735/1 and BB/G011818/1; and the Grant Agency of the Czech Republic, grant number 15-08835Y for funding.
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Goldberg, M.W., Fišerová, J. (2022). Scanning Electron Microscopy (SEM) and Immuno-SEM of Nuclear Pore Complexes from Amphibian Oocytes, Mammalian Cell Cultures, Yeast, and Plants. In: Goldberg, M.W. (eds) The Nuclear Pore Complex. Methods in Molecular Biology, vol 2502. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2337-4_27
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DOI: https://doi.org/10.1007/978-1-0716-2337-4_27
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