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There are many ways of preparing specimens for electron microscopy. The EMP staff are available to help investigators determine the best way of getting the information they need from their specimens.

In life science specimen preparation methods perform the task of stabilizing the initially hydrated molecule so that in can be placed and observed in the vacuum of the electron microscope. Biological specimens are typically prepared using negative staining or cryo-fixation. The facilities for transmission electron microscope biological specimens include cryo-fixation (vitrification - cryo-EM) and negative staining preparation techniques.

In materials science brittle materials specimens can often be crushed, dispersed in alcohol and a drop of liquid placed onto a grid supporting an amorphous carbon film. Nanoparticles are deposited on electron microscopy grids by drop casting or dip coating methods.

EMP TEMs take standard size specimens (3mm in diameter). The maximum thickness of the specimen depends on the density of the material, the accelerating voltage of the microscope and the resolution of the analysis to be carried out. The specimen have to be thin samples (less than 100nm thick) of the correct diameter (3 mm).

Negative Staining

The contrast produced by the molecules itself is normally insufficient for direct observation in the electron microscope and a contrasting method has to be used. Negative Staining with heavy metal salts such as uranyl acetate produce high contrast and protects the molecule from collapsing. Instead of the molecule (with its interior density variations) only a cast of exterior surface of the molecule is imaged, and only its shape can be reconstructed.

[Principe of negative staining showing side view of specimen surrounded by stain] [Principe of negative staining preparation laboratory]

Cryo-fixation (vitrification)

Cryo-fixation is a method to “sustain” the molecule in a medium that closely approximates the aqueous environment. The samples are embedded in vitreous ice and we can obtain images of fully hydrated macromolecules (“Frozen-hydrated”). The vitrification process enables the structure of macromolecules and the cellular architecture to be studied in a frozen hydrated near native state.

For preparing the cryo-samples a Vitrobot is available.

[picture of cryo-sample preparation laboratory] [Introducing the cryo-holder in the Electron Microscope]