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Specimen Preparation Techniques

Visualizing a biological specimen with an electron microscope is not a trivial task, mostly because of the intrinsic nature of the electron and matter interactions that are responsible for the image formation. In order to be imaged using TEM the specimen has to be very thin and for using SEM the specimen needs a conductive surface. In both cases the specimen has to be placed inside high vacuum. Thus, biological specimens cannot be imaged in their native state and need to be heavily processed. Accordingly, sample preparation is a crucial step, which represents an essential activity and service in our facility.

For immunodetection

Immuno-preparations aim to preserve, expose and detect epitopes, in some cases at a cost to ultrastructure. The samples are crosslinked weekly or cryo-immobilised and processed more gently; solvent dehydration is omitted or performed at sub-zero temperatures and resin is cured with UV light instead of heat. The epitopes are detected using 1° antibodies and 2° probes tagged with gold particles.

For SEM topography

Similarly to the ultrastructure preparations described above this technique ensures the preservation of fine ultrastructural details and stabilisation of the specimen. However, the specimen is not in any way embedded. Since charging is much more of an issue than in TEM it is crucial to render the specimen conductive to avoid image distortions.

Plunge-freezing

At our facility the plunge-freezing technique is used primarily for cryo-screening of particulate samples.