Cryotechniques

Abstract

The first question that arises when considering cryotechniques in general is, what advantages are there to cryofixation? As we discussed in Chapter 1, fixation ideally instantaneously stops biological activity, immobilizes cellular components, and enables them to withstand any further processing procedures. In principle, biological materials can be fixed with either chemical or physical (cold, heat) techniques. Good chemical fixation requires the fast diffusion of chemical agents through membranes and cytoplasmic components. Cryofixation, on the other hand, requires the rapid diffusion of heat out of the specimen. The reason cryofixation can be superior to chemical fixation is that the rate of chemical diffusion into the specimen is much slower than the rate of heat diffusion out of the specimen. However, most cryofixation techniques adequately freeze samples to no more than 15 µm, in contrast to chemical fixation, which consistently fixes samples to a depth of 0.5 mm. Cryofixation is often inferior for structural studies because of the limited sample size available, but is vastly superior to chemical fixation for many microanalytical or immunocytochemical procedures.