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Preparation of ultrathin amorphous ice films for cryo‐electron microscopy
Author(s) -
Jakubowski U.,
Mende M.
Publication year - 1991
Publication title -
journal of microscopy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.569
H-Index - 111
eISSN - 1365-2818
pISSN - 0022-2720
DOI - 10.1111/j.1365-2818.1991.tb03087.x
Subject(s) - cryofixation , materials science , monolayer , composite material , thin film , adsorption , electron microscope , optics , nanotechnology , chemistry , physics , organic chemistry
SUMMARY Electron microscopy of biological macromolecules embedded in vitrified ice films suffers from serious problems caused by excessive inelastic scattering, beam‐induced movements of the specimen, deformation of the molecules by adsorption at the water‐air interface and insufficient mechanical stability of the films. We have built an environmental chamber to control temperature and humidity independently in order to produce ultrathin water films (< 20 nm) spanning holes with diameters of 20 μm to 1 mm. The surface tension of the water films was reduced by adding lipid monolayers, thus prolonging the usable time for thinning of the film and avoiding adsorption artefacts in the embedded material. After cryofixation in ethane a carbon film was evaporated on each side of the specimen to stabilize the ultrathin ice—lipid film. Mechanical stability and charging effects could successfully be reduced by this preparation method. Collapsing water films could be cryofixed and the shape of the hole was analysed. By the eccentricity of the elliptical holes an estimation could be made of the burst velocity of the rim of the hole and of the cooling rate of the cryofixation process.