Adult brain workflow - 
Isolation and analysis of neural cells

Isolation and cultivation of:

astrocytes from adult mouse brain 

oligodendrocytes from adult mouse or rat brain 

adult neural stem cells from adult mouse brain 

microglia from adult mouse or rat brain 

endothelial cells from adult mouse brain 

neurons from adult mouse brain 

Highly purified cells after sample clearing

The cell suspension from adult brain tissue contains a significant amount of cell debris (A) and erythrocytes (B), which hampers subsequent cell isolation, cultivation, and analysis. The Adult Brain Dissociation Kit optimizes the tissue dissociation process, and after sample clearing yields highly purified, living cells with much less cell debris (C) and erythrocytes (D).

Isolation of adult astrocytes

A single-cell suspension from a 3 month old mouse brain was prepared using the Adult Brain Dissociation Kit, mouse and rat. Adult astrocytes were isolated from the single-cell suspension with the Anti-ACSA-2 MicroBead Kit, mouse. Cells were fluorescently stained with the Anti-ACSA-2 antibody and analyzed by flow cytometry on the MACSQuant Analyzer.

Isolation of adult neurons

A single-cell suspension from a 3 month old mouse brain was prepared using the Adult Brain Dissociation Kit, mouse and rat. Subsequently, adult neurons were isolated from the single-cell suspension with the Neuron Isolation Kit, mouse. Cells were fluorescently stained with antibodies specific for non-neuronal cell marker and the ACSA-2 antigen and analyzed by flow cytometry using a MACSQuant Analyzer.  
 

Isolation of adult microglia

Isolation of mouse microglia using the CD11b MicroBeads, human and mouse from a 2 month old mouse brain. The cell separation using the MicroBeads resulted in a high purity of 96,5% and a viability of 95,2 %. Cell were stained with CD11b-FITC and CD45-APC for further analysis. 

Isolation of adult pre-oligodendrocytes

Single-cell suspension of adult brain tissue was used for cell separation with anti-O4 MicroBeads, human, mouse, rat. Cells were stained with Anti-O4-PE to verify a pure cell population of pre-oligodendroytes. 

Primary mouse neuron cultivation

Primary adult neurons were cultured in MACS Neuro Medium and MACS Neurobrew-21 for 7 days after isolation from adult brain tissue. Cells were fixed and stained with anti-MAP2 (green) and anti-βIII Tubulin (red) antibodies. 

AstroMACS separation buffer and medium enhances growth of astrocytes

Adult astrocytes were isolated from 8-week old mouse brains using the Adult Brain Dissociation Kit and the Anti-ACSA-2 MicroBeads. When using the AstroMACS Separation Buffer and the AstroMACS Medium, growth of astrocytes is strongly improved (A) in comparison to astrocyte culture without the AstroMACS Separation Buffer (B). Cells were fixed and stained with GLAST Antibody (green) and DAPI (blue).

Single-cell analysis of unseparated neural cells

Single-cell analysis of non-sorted adult neural cells led to identification of 24 different subclusters, representing neuronal and different glial cell types, including astrocytes, microglia, oligodendrocytes, vascular cells, and immune cells.

Single-cell analysis of isolated neurons

Isolation of neurons using the Neuron Isolation Kit resulted in an increased resolution of neuronal clusters, including several subpopulations of immature and mature neurons. Moreover, the amount of non-neuronal cell types was significantly decreased. 

Cultivation and differentiation of adult neural stem cells

Adult neural stem cell sorting and analysis kit, mouse results in a pure population of neural stem cells from the subventricular zone, which is ready for culturing of neurospheres (A, B) and differentiation into neuronal and glial cells (C-F).