Cerebellum Astrocyte Isolation via MACS

The cerebellum was macrodissected for astrocyte isolation using magnetic-activated cell sorting (MACS) [16 (link)]. Briefly, the cerebellum was mechanically dissociated using a scalpel, followed by enzymatic dissociation using the Neural Tissue Dissociation kit (P) (Miltenyi Biotec, Cologne, Germany), according to the manufacturer’s instructions. Myelin and cell debris were removed using the Myelin removal kit (Miltenyi Biotec), followed by microglia removal using Cd11b microbeads (Miltenyi Biotec). Finally, astrocytes were isolated using anti-astrocyte cell surface antigen (ACSA)-2 beads (Miltenyi Biotec). All the separation steps were performed using the AutoMACS Pro Separator equipment (Miltenyi Biotec).