Defining standard enzymatic dissociation methods for individual brains and spinal cords in EAE

Area of Science:

• Neuroimmunology
• Cellular Biology
• Experimental Pathology

Background:

• Experimental autoimmune encephalomyelitis (EAE) is a mouse model for multiple sclerosis (MS).
• Isolating mononuclear cells from the central nervous system (CNS) is crucial for understanding EAE pathogenesis.
• Standardizing CNS tissue dissociation methods is essential for reliable experimental outcomes.

Purpose of the Study:

• To evaluate and compare the capacity, effectiveness, efficiency, and reliability of different tissue dissociation methods.
• To identify the optimal method for isolating mononuclear cells from the CNS of EAE mice.
• To qualify an assay for downstream analysis in EAE research.

Main Methods:

• Tested Percoll PLUS, a commercial Neural Tissue Dissociation Kit, accutase, and papain.
• Applied criteria: ≥80% cell viability, ≥5 × 10^5 cells/sample, test-retest reliability, and correlation with EAE disease course.
• Utilized C57BL/6 mice with active EAE.

Main Results:

• Enzymatic dissociations significantly increased mononuclear cell yield.
• All enzymatic methods were more efficient and effective than Percoll PLUS.
• Only the Neural Tissue Dissociation Kit showed a significant correlation between cell numbers and EAE disease severity.

Conclusions:

• The Neural Tissue Dissociation Kit is recommended as the standard method for CNS tissue dissociation in EAE.
• Optimized CNS dissociation methods can aid in identifying cellular events relevant to MS pathogenesis.
• This standardization supports reproducible research in neuroinflammatory diseases.