The Transmission Electron Microscope
- Preparation of Specimen

In a TEM, the specimen you want to look at must be of such a low density that it allows electrons to travel through the tissue. There are different ways to prepare your material for that purpose. You can cut very thin slices of your specimen from a piece of tissue either by fixing it in plastic or working with it as frozen material. Another way to prepare your specimen is to isolate it and study a solution of for example viruses or molecules in the TEM.

You can also stain the specimen in different ways and use markers to locate specific things in the tissue. It can for example, be stained with heavy metals like uranium and lead, which scatters electrons well and improves the contrast in the microscope.

Below are two examples described in more detail.

1. Sections of Embedded Material
Biological material contains large quantities of water. Since the TEM works in vacuum, the water must be removed. To avoid disruption as a result of the loss of water, you preserve the tissue with different fixatives. These cross-link molecules with each other and trap them together as stable structures. The tissue is then dehydrated in alcohol or acetone.

After that, your specimen can be embedded in plastic that polymerize into a solid hard plastic block. The block is cut into thin sections by a diamond knife in an instrument called ultramicrotome. Each section is only 50-100 nm thick.

The thin sections of your sample is placed on a copper grid and stained with heavy metals. The slice of tissue can now be studied under the electron beam.

2. Negative Staining of Isolated Material
The isolated material (can be a solution with bacteria or a solution with isolated molecules) is spread on a support grid coated with plastic. A solution of heavy metal salt is added. The metal salt solution does not bind to the material but forms a "shadow" around it on the grid. The specimen will appear as a negative picture when viewing it in the TEM.