Single-cell sequencing of FACS-sorted mouse astrocytes, investigating the effect of absence of microglia in a mouse model of ß-amyloidopathy
Single-cell sequencing of FACS-sorted mouse astrocytes, investigating the effect of absence of microglia in a mouse model of ß-amyloidopathy
To sort astrocytes, mouse neocortices were rapidly harvested then dissociated with Adult Brain Dissociation Kit (Miltenyi Biotec) on a gentleMACS Octo Dissociator using program 37C_ABDK_01. Dissociated samples were then treated with debris and red blood cell removal steps to obtain cell suspensions. For FACS, cells were resuspended in a final volume of 100 uL 0.1% PB buffer. Then cells were incubated with fluorescent conjugated antibodies at 4° C for 30 minutes. The following antibodies were used: ACSA2 APC (1:200, Miltenyi Biotec, 130-116-245) and O4 PE (1:100, Miltenyi Biotec, 30-117-357). ACSA2 APC (+) and O4-PE (-) live single cells were selected as the astrocyte population.   Library preparation was conducted with the Chromium Single Cell 3′ v3.1 Reagent Kit (10x Genomics), following the manufacturer's instructions. Library preparation was conducted with the Chromium Single Cell 3′ v3.1 Reagent Kit (10x Genomics), following the manufacturer's instructions.