Single cell RNA seq analysis of microglia from aged wild-type, NLRP3-/-, APP/PS1 and APP/PS1.NLRP3-/- mice.

NLRP3 inflammasome activation is involved in the progression of Alzheimer's disease. To unravel the unique cell populations and clusters regulated by NLRP3 in the brain, we enriched CD11b+ cells from the brains of 18 month old wild-type, NLRP3-/-, APP/PS1 and APP/PS1.NLRP3-/- mice for single cell RNAseq. The brains from wild-type, NLRP3-/-, APP/PS1 and APP/PS1.NLRP3-/- mice were isolated and dissociated using the Neural Tissue Dissociation kit (P) (Miltenyi Biotec), and using MACS isolation the myelin was labelled and removed magnetically before the enrichment of CD11b+ microglia following the manufacturers guidelines (Miltenyi Biotec). Single, live, CD11b+ cells were FACS sorted at 4C (BD FACS Aria III) into 384-well plates containing 384 primers and Mineral oil (Sigma). After sorting, plates were snap-frozen on dry ice and stored at -80CNLRP3炎性小体（NLRP3 inflammasome）的激活与阿尔茨海默病（Alzheimer's disease）的病程进展密切相关。为阐明大脑中受NLRP3调控的独特细胞群与细胞簇，我们从18月龄野生型、NLRP3基因敲除（NLRP3-/-）、APP/PS1及APP/PS1.NLRP3-/-小鼠的脑组织中富集CD11b+细胞，用于单细胞RNA测序（single cell RNAseq）。我们先分离上述四种基因型小鼠的脑组织，采用神经组织解离试剂盒（P型）（Miltenyi Biotec）进行组织解离；随后依照厂商操作指南（Miltenyi Biotec），使用磁激活细胞分选（Magnetic Activated Cell Sorting，MACS）技术对髓磷脂进行磁标记并磁去除，进而富集CD11b+小胶质细胞。将单个活态CD11b+细胞在4℃条件下，通过BD FACS Aria III流式细胞仪分选至含有384种引物与矿物油（Sigma）的384孔板中。分选完成后，将培养板置于干冰上快速冷冻，并于-80℃冰箱保存。