formaldehydifixaationa

Formaldehyde fixation is a common chemical method used to preserve biological tissues for microscopic examination. By creating covalent cross-links within and between proteins and nucleic acids, formaldehyde stabilizes cellular architecture and prevents enzymatic degradation, enabling accurate morphological assessment.

Chemical mechanism: formaldehyde reacts primarily with amino groups in proteins to form methylene bridges that link molecules and create a cross-linked network. In practice, fixed tissues are prepared in buffered formaldehyde solutions, commonly 10% neutral buffered formalin, which contains about 3.7-4% formaldehyde. Paraformaldehyde solutions can also be used, often in phosphate buffer.

Procedure: tissues are fixed after collection, typically by immersion or by perfusion in larger specimens. Fixation is followed by dehydration, clearing, and embedding in paraffin for sectioning, or by cryopreservation for frozen sections. Fixation times vary by tissue type and fixative concentration.

Applications and limitations: widely used in histology, pathology, and immunohistochemistry due to good overall preservation and compatibility with standard stains. Drawbacks include cross-linking-induced masking of epitopes and nucleic acids, potential autofluorescence, and tissue shrinkage or hardening. Antigen retrieval and optimized protocols are often required for downstream assays.

Safety and alternatives: formaldehyde is a hazardous substance, a suspected carcinogen, and should be handled in well-ventilated areas with appropriate PPE and waste disposal. Alternatives include glutaraldehyde, ethanol or methanol-based fixatives, Bouin’s or Bouin-like solutions for special applications, or fixation methods such as ethanol fixation or cryofixation when appropriate.