title: GSM6720067: RH22TSB; Mus musculus; RNA-Seq
library_construction_protocol: We performed a papain enzymatic digestion followed by serial trituration to prepare a single cell suspension from the hippocampus. We enriched for microglia by applying a 40% Percoll centrifugation to our cells and collecting the cell pellet, and further purified by fluorescence activated cell sorting (FACS). We sorted CD11b+/CD45low double positive cells, representing microglia, rather than CD11b+/CD45high cell surface markers, which represent macrophages. scRNA-seq was performed by the Advanced Genomics Core at the University of Michigan using the 10X Genomics Chromium system (10X Genomics): An automated counter (Countess II, Thermo Fisher) was used to quantify cells/µl. The single cell suspension was then diluted to a final concentration ranging from 700 to 1000 cells/µl. Using the Chromium Controller, 3' libraries of single cells were created leveraging 3' V3.1 chemistry using NextGEM Chip G reagents, according to the manufacturer's protocol (all from 10X Genomics). The quality of the final library was evaluated by Tapestation 4200 (Agilent) and Kapa qPCR (Roche) was used to quantify libraries. Pooled libraries were sequenced using 150 bp paired-end format (Illumina NovaSeq 6000). De-multiplexed Fastq files were generated (Bcl2fastq2 Conversion Software, Illumina) and reads were aligned and counted (CellRanger Pipeline, 10X Genomics).