Cryo scanning electron microscopy (cryo-SEM)

About this technique

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Samples for conventional scanning electron microscopy (SEM) must be prepared for the high-vacuum environment of the microscope by being fixed, dehydrated and dried. In some cases, this processing technique cannot be used because it would either destroy the sample or cause unacceptable artifacts.

An excellent alternative in such situations is cryo scanning electron microscopy (cryo-SEM). This technique allows wet or liquid samples to be snap frozen, prepared and held at very low temperature while being examined in the microscope. Sample preparation time is relatively short, so can be readily repeated if necessary. An added advantage is that the frozen sample can be fractured during preparation to reveal internal structures. For example, a leaf can be fractured to reveal the internal detail, or a suspension of nanoparticles can be examined to assess the dispersion. Sublimation of some of the ice at the surface of the fracture will help to enhance detail. After cooling below sublimation temperature, the sample is coated with a conductive metal to prevent electrical charging during examination. Freeze-fracture can also be extremely valuable in working with lipid bilayers and membranes as fracturing can split the bilayers. Such preparations can also be visualised by transmission electron microscopy (TEM) for high-resolution imaging of these structures.

Samples for cryo-SEM are generally not prepared at all beforehand; the best results are obtained from samples that are as close to their natural state as possible. The technique has proved its effectiveness for samples such as plant and animal tissues, biofilms, food, pharmaceuticals and nanoparticles in suspensions, among numerous others.