CRYOFIXATION
Supercool water
When water freezes, it expands because of ice crystals. That is why ice-cubes float up in your drink. So when we normally freeze cells, the water inside also expands and breaks the cell structure. We have two methods that can freeze cells so fast, water does not have time to form ice crystals and thus is actually still "liquid". These methods called "plunge freezing" and "high-pressure freezing" can be used to investigate cells and tissues as they are frozen in time, which is the closest we can get to anpicture of nanoscale processes in living cells.
Sample preparation
After freezing your sample, we can do two things to investigate it by electron microscopy. We look at the sample at temperatures below 150 C as it it is frozen, this gives the closest to natural state image of your sample, but is limited by how thick the sample is and also no contrast reagents can be used.
We can also replace the frozen water for plastic in a process called freeze-substitution. This is the best way to investigate thicker smaples and allows the use of contrast reagents.