In this application protocol, we present a protocol to generate highly purified and viable neurons from adult mouse brain tissue. Brain tissue from mice older than P7 is dissociated into a single-cell suspension using the Adult Brain Dissociation Kit. The extracellular matrix is enzymatically digested using the kit components, while the gentleMACS™ Dissociator with Heaters is used for the mechanical dissociation steps during the on-instrument enzyme incubation. After the dissociation, myelin and cell debris are removed using the Debris Removal Solution and is followed by an subsequent removal of erythrocytes using the Red Blood Cell Removal Solution. The Neuron Isolation Kit, mouse is used to isolate neurons from the single-cell suspension.
|Enzyme mix 1||Enzyme mix 2|
|Debris Removal Solution||D-PBS||Overlay (D-PBS)|
|900 µL||3100 µL||4 mL|
|1800 µL||6200 µL||4 mL|
▲ Note: In case of very small amount of tissue (< 100 mg), cell debris removal can be performed in a 5 mL reagent tube using 450 µL of Debris Removal Solution, 1550 µL of D-PBS for resuspension of the cell pellet, and 2000 µL D-PBS for overlay.
|Column||Max. number of labeled cells||Max. number of total cells||Selector|
VarioMACS, SuperMACS II
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