From fix to fit into the autoptic human brains

Area of Science:

• Neuroscience
• Histology
• Biochemistry

Background:

• Formalin-fixed, paraffin-embedded (FFPE) human brain tissues are crucial for research but long-term storage degrades tissue quality.
• Standard fixation methods often compromise immunostaining and nucleic acid extraction, hindering detailed analysis.
• A gold standard protocol for FFPE tissue fixation and analysis is still lacking.

Purpose of the Study:

• To evaluate different fixation systems for FFPE human brain tissues.
• To develop an improved fixation method for enhanced immunostaining and morphology preservation.
• To optimize protocols for analyzing neuronal markers, neurodegeneration, and stem cell antigens in postmortem brain tissue.

Main Methods:

• Comparison of four different fixation systems on FFPE human brain tissues.
• Histological and immunohistochemical staining analyses were performed.
• A modified combined fixation method using formalin and formic acid was developed and tested.

Main Results:

• The modified fixation method demonstrated improved preservation of cellular morphology.
• Successful postmortem immunostaining for NeuN (neuronal marker) was achieved, overcoming common issues of faint or lost staining.
• The protocol enabled Fluoro Jade C staining (neurodegeneration marker) and immunofluorescent staining for stem cell antigens.

Conclusions:

• A modified combined fixation method using formalin and formic acid offers an efficient, safe, and fast approach for immunolabelling long-stored FFPE human brain tissues.
• This method significantly improves the quality of immunostaining for key markers, aiding in histopathology and topographic studies.
• The optimized fixation procedure is valuable for analyzing various cellular and molecular features in postmortem brain samples.