Overview
Fixation preserves tissue morphology and antigens by cross-linking or precipitating biomolecules. The choice of fixative and method strongly influences sectioning and staining quality.
Common Fixatives
- Formaldehyde / paraformaldehyde (PFA): the most common; good morphology and broad antigen compatibility. 4% PFA is standard for brain tissue.
- Glutaraldehyde: stronger cross-linking, excellent ultrastructure, but reduces antibody accessibility.
- Alcohol-based: fast, good for some histochemical stains, but weaker morphology.
Immersion vs Perfusion Fixation
Perfusion fixation delivers fixative through the vascular system for rapid, uniform preservation and is preferred for brain studies. Immersion fixation is simpler but can be uneven and slower.
Fixation Time
Over-fixation can mask antigens; under-fixation can cause poor morphology. Optimise and keep fixation time consistent across samples, then move to sectioning or storage.
Best Practices
Use fresh fixative, keep pH buffered and temperature controlled, and document fixation conditions — they are a frequent source of between-lab variability.