Progress in scanning electron microscopy of frozen-hydrated biological specimens. 1993

R Hermann, and M Müller
Laboratory for Electron Microscopy I, ETH Zentrum, Institute for Cell Biology, Zürich, Switzerland.

Modern scanning electron microscopy yields structural information down to 2 to 5 nm from thin, beam transparent biological specimens. This paper examines the possibilities of garnering this level of structural information from bulk, frozen-hydrated samples. Freeze-fractured, frozen-hydrated yeast cells, frequently taken as a yardstick to monitor progress in low-temperature scanning electron microscopy, have been used to optimize both metal shadowing methods and observation parameters (e.g. accelerating voltage, electron beam irradiation of the specimen). Uncoated frozen-hydrated yeast cells do not change electrically at an accelerating voltage of 30 kV. Increasing charging effects are however observed with decreasing accelerating voltages. Very thin metal films are therefore used for specimen coating to localize and enhance the specific secondary electron signal. Planar-magnetron sputtering of a 1 nm metal layer provides high resolution secondary electron images, at 30 kV, of freeze-fractured, frozen-hydrated yeast cells in an in-lens field-emission scanning electron microscope. Structural information comparable to that of transmission electron microscopy of freeze-fractures is attained. Planar-magnetron sputtering of either chromium, tungsten or platinum results in essentially the same information density (smallest visible significant structural detail). Frozen-hydrated samples are very beam sensitive and have to be observed under minimum dose conditions.

UI MeSH Term Description Entries
D008855 Microscopy, Electron, Scanning Microscopy in which the object is examined directly by an electron beam scanning the specimen point-by-point. The image is constructed by detecting the products of specimen interactions that are projected above the plane of the sample, such as backscattered electrons. Although SCANNING TRANSMISSION ELECTRON MICROSCOPY also scans the specimen point by point with the electron beam, the image is constructed by detecting the electrons, or their interaction products that are transmitted through the sample plane, so that is a form of TRANSMISSION ELECTRON MICROSCOPY. Scanning Electron Microscopy,Electron Scanning Microscopy,Electron Microscopies, Scanning,Electron Microscopy, Scanning,Electron Scanning Microscopies,Microscopies, Electron Scanning,Microscopies, Scanning Electron,Microscopy, Electron Scanning,Microscopy, Scanning Electron,Scanning Electron Microscopies,Scanning Microscopies, Electron,Scanning Microscopy, Electron
D005614 Freeze Fracturing Preparation for electron microscopy of minute replicas of exposed surfaces of the cell which have been ruptured in the frozen state. The specimen is frozen, then cleaved under high vacuum at the same temperature. The exposed surface is shadowed with carbon and platinum and coated with carbon to obtain a carbon replica. Fracturing, Freeze,Fracturings, Freeze,Freeze Fracturings
D012441 Saccharomyces cerevisiae A species of the genus SACCHAROMYCES, family Saccharomycetaceae, order Saccharomycetales, known as "baker's" or "brewer's" yeast. The dried form is used as a dietary supplement. Baker's Yeast,Brewer's Yeast,Candida robusta,S. cerevisiae,Saccharomyces capensis,Saccharomyces italicus,Saccharomyces oviformis,Saccharomyces uvarum var. melibiosus,Yeast, Baker's,Yeast, Brewer's,Baker Yeast,S cerevisiae,Baker's Yeasts,Yeast, Baker
D013048 Specimen Handling Procedures for collecting, preserving, and transporting of specimens sufficiently stable to provide accurate and precise results suitable for clinical interpretation. Specimen Collection,Collection, Specimen,Collections, Specimen,Handling, Specimen,Handlings, Specimen,Specimen Collections,Specimen Handlings
D014867 Water A clear, odorless, tasteless liquid that is essential for most animal and plant life and is an excellent solvent for many substances. The chemical formula is hydrogen oxide (H2O). (McGraw-Hill Dictionary of Scientific and Technical Terms, 4th ed) Hydrogen Oxide
D015925 Cryopreservation Preservation of cells, tissues, organs, or embryos by freezing. In histological preparations, cryopreservation or cryofixation is used to maintain the existing form, structure, and chemical composition of all the constituent elements of the specimens. Cryofixation,Cryonic Suspension,Cryonic Suspensions,Suspension, Cryonic

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