We know that the technique used in TEM sample preparation involves multiple steps, one of the most important of them is fixation.
Fixation can be of two types:
Cryofixation, that suggests that the specimen are subjected to freezing temperature to perserve the cell's living material before we observe them.
Chemical fixation involves the use of certain chemicals, such as formalin, glutaraldehyde, etc. for almost same purpose.
The question is when it is mandatory to use one of them only? Such as, cryofixation or chemical one? At some instances, have the researchers ever need to use both of the techniques at the same time?