Mouse fcγ receptor block reagent

Mouse brains were dissociated into single cell suspension using the Adult Brain Dissociation Kit (Miltenyi Biotec, Cat# 130-107-677) with cell debris removal following manufacturer’s instructions. Single cells were resuspended in cold MACS buffer supplemented with mouse Fcγ receptor block reagent (553141; BD Biosciences) and incubated on ice for 5 min, followed by staining with fluorescently tagged antibodies in the brilliant stain buffer (563794; BD Biosciences) on ice for 20min. The cells were washed twice with MACS buffer, and used for FACS sorting. Antibodies: APC-Cy7 rat anti-CD11b (clone M1/70, Cat# 557657; BD Biosciences), ACSA2-PE (Miltenyi Biotec, Cat# 130-123-284), O4-APC (Miltenyi Biotec, Cat# 130-118-978), BV421 Rat Anti-Mouse CD24 (clone M1/69, Cat# 562563; BD Biosciences).

2 FE4-female, 4 TAFE4-TAM-female, and 4 TFE4-TAM-female mice, around 285 days, were randomly selected and perfused with perfusion buffer (DPBS + 1 X Glutamax). Mouse brains were dissociated into single cell suspension using the Adult Brain Dissociation Kit (Miltenyi Biotec, Cat# 130-107-677) with cell debris removal following manufacturer’s instructions. Single cells were resuspended in cold MACS buffer supplemented with mouse Fcγ receptor block reagent (553141; BD Biosciences) and incubated on ice for 5 min, followed by staining with APC-Cy7 rat anti-CD11b (clone M1/70, Cat# 557657; BD Biosciences) on ice for 20min. The cells were washed twice with MACS buffer, and microglia were isolated by fluorescence-activated cell sorting.