An optimized method to process mouse CNS to simultaneously analyze neural cells and leukocytes by flow cytometry

Area of Science:

• Neuroscience
• Immunology
• Cell Biology

Background:

• Flow cytometry is a powerful cell analysis tool.
• CNS tissue preparation is a bottleneck for neuroscientific flow cytometry.
• Previous methods did not simultaneously analyze neural and immune cells.

Purpose of the Study:

• To optimize central nervous system (CNS) tissue preparation for flow cytometry.
• To enable simultaneous quantification of neural and immune cells.

Main Methods:

• Optimized CNS tissue digestion using collagenase D and DNase I.
• Developed a one-step Percoll™ gradient method for cell separation.
• Analyzed neural cells (neurons, oligodendrocytes, microglia) and leukocytes (macrophages, T lymphocytes) via flow cytometry.

Main Results:

• The one-step Percoll™ method yielded higher cell numbers than gradient Percoll™.
• Collagenase D + DNase I digestion maximized cell recovery and preserved marker integrity.
• The combined method proved optimal for CNS cell analysis in healthy and disease models.

Conclusions:

• This optimized method allows concurrent analysis of neural and immune cells in individual mice.
• The technique is applicable to both healthy and pathological CNS conditions.
• This approach will advance the use of flow cytometry in neurosciences.