The transmission electron microscope allows us to see particles, which can be as close as 2 nanometres, as separate structures. The resolution of this microscope is about 2 nanometres. It can be used with high or low magnification. Under low magnification, they can produce a sharp definition of the specimen. The higher the magnification on the specimen, the higher the amount of visible detail.
The scanning electron microscope gives a three-dimensional effect, which shows surface detail. It scans electron beams to and fro across the surface of a complete specimen. It doesn’t have a resolving power as high as the transmission electron microscope. Despite this, it can take larger specimens than the transmission electron microscope.
The specimen for both electron microscopes must be very thin. This is because electrons must be able to pass through parts of the specimen. Molecules in air interrupt the flow of electrons. For this reason, the specimen is placed into a vacuum chamber, which is inside an electron microscope, before it is split along a line of weakness. Very small specimens such as viruses and large molecules do not have to be sectioned.
Photographs of specimens viewed with an electron microscope are called electronmicrographs. Electron-dense areas scatter the electrons and produce dark areas on the electronmicrograph. The atoms in organic molecules have a low atomic number and so do not scatter electrons. Light areas on an electronmicrograph are produced when electrons have been able to pass through the specimen.
Beams of electrons have very short wavelengths and therefore they cannot pass through glass. A high-intensity electron beam can destroy parts of the specimen, producing light-coloured areas on the screen.
When using a transmission electron microscope, you first have to fix the tissue specimen to preserve it. It is preserved using substances which prevent enzyme action. The tissue is then soaked in alcohol to dehydrate it. Once dehydrated, it is embedded in a resin, which becomes hard. The embedded tissue is cut using a microtome. Thin sections are mounted on copper grids which provide support. The sectioned tissue is stained using heavy metal stains. This is because specimens must be stained in order to improve contrast.
When using a scanning electron microscope, the process is different. First the tissue is frozen rapidly in liquid nitrogen at about –210°c. This technique is known as freeze fracture, which allows surface detail to be seen with a transmission electron microscope. The frozen tissue is then fractured into very thin sections with a sharp blade, made either of metal or glass. The ice is evaporated from the surfaced and the exposed surface is sputter coated with carbon or gold. Then carbon or a gold replica is floated off the tissue and finally it is mounted on a disc. The treatment might introduce artefacts.
The electron microscope is a useful tool to look at the ultrastructure of the cells, including the fine structure of the cell organelles. This is because of its ability to magnify specimens up to 500, 000 times.
