What is CryoEM?
Cryo-electron microscopy (also known as electron cryomicroscopy or cryoEM) is the method our lab uses to “take photographs” of viruses and other macromolecular complexes. The following is an abbreviated, layman’s terms explaination of how cryoEM works.
First, we must prepare the specimen for studying with the electron microscope. For the purposes of this example, let’s pretend we’re trying to image a virus. We grow it to a concentrated number (or high titer), isolate it, and purify it. Then, when we have a good sample, we place a drop containing thousands of virions onto a thin film which is then quickly frozen to the temperature of liquid nitrogen in order to protect and preserve the specimen during observation.
When the sample is ready, we can start shooting electrons at it. Cryo-EM uses a very low dose of electrons (about 1-10 electrons per square angstrom) so that the biological sample is not damaged during the study. The electrons pass through empty areas and are bounced or refracted from dense areas.
Please note that although the lenses in the image to the left look an awful lot like a magnifying glass, electron microscopes actually use magnetic coils to “magnify” and focus the electrons.
When imaging is complete, we end up a two-dimensional image similar to the one you see in the diagram. To render these flat images into a three-dimensional model, we must use computerized 3-D data merging. You can read about that here.
What is CryoET?
Cryo-electron tomography is an imaging method similar to cryo-electron microscopy. However, the primary difference between the two lies in that cryoEM relies on hundreds of thousands of different particles to composite into a single 3-D model, and cryoET uses a single particle, but hundreds of images.
The advantage of cryoET over cryoEM is that structures that are flexible or otherwise specific to a single particle can be rendered. The disadvantage is that because one must put the same particle under radiation for the entire duration of imaging, the number of images that can be taken is limited.