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A comprehensive protocol for simplified mouse DRG fixation, processing and F4/80 immunohistochemistry: Overcoming
Nicolette Tay1, Ammar Alshammari2, Simranpreet Kaur3
1School of Pharmacy and Pharmaceutical Sciences, The University of Queensland, Woolloongabba, QLD, Australia.
Journal of Neuroscience Methods
|March 25, 2025
Summary
We optimized a method for fixing and processing mouse dorsal root ganglia (DRGs) for immunohistochemistry (IHC). This technique preserves tissue integrity and cell morphology, enabling better analysis of neuropathic pain mechanisms.
Area of Science:
- Neuroscience
- Immunology
- Histopathology
Background:
- Dorsal root ganglia (DRGs) are crucial in neuropathic pain.
- Immunohistochemistry (IHC) visualizes immune cells in DRGs.
- DRG isolation for IHC is challenging due to size and location.
Purpose of the Study:
- Optimize fixation, harvesting, processing, and IHC staining of mouse DRGs.
- Utilize F4/80, a monocyte-macrophage marker, for analysis.
- Ensure tissue integrity and compatibility with histopathology.
Main Methods:
- Whole mouse spinal column fixation in 10% neutral-buffered formalin.
- DRG isolation after 24h fixation.
- 9h processing and proteinase K antigen retrieval.
Main Results:
- Optimized immersion-fixation preserved cellular morphology and antigenicity.
- Achieved high-quality histological outcomes.
- Method is non-invasive and reduces time to fixation.
Conclusions:
- The protocol provides high-quality histological outcomes.
- It is comparable to perfusion-based techniques.
- A grading system aids in selecting optimal experimental conditions.
