Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model

We used CITE-seq (10x Gemoics-based) to profile and compare the transcriptomes and cell surface expression of a set of immune markers of naïve brain myeloid cells from young-adult (12 weeks old) wild type C57Bl/6 mice, aged (72 weeks old) wild type C57Bl/6 mice and young-adult and aged C57Bl/6 mice with antibiotics induced gut microbiota depletion. In one sequencing round, we sequenced a total of 12 different mouse brains (3 young control, 3 aged control, 3 young depleted gut microbiota (ABX) and 3 aged depleted gut microbiota (ABX)). We created an antibody pool consisting of 31 different antibodies (CCR2/CD192, C117/c-kit, CD11b, CD11c, CD172a/SIRPα, CD25, CD3, CD38, CD4, CD44, CD45, CD45R/B220, CD86, CD8a, CD90.1, Cx3cr1, F4/80, I-A/I-E, Ly6C, Ly6G, NK1.1, PD-1, PD-L1, CD169/Siglec-1, Siglec-H, TMEM119, XCR1, CD24, CD103, CD64, CD83) and stained each brain individually with this antibody pool. Then, we stained each brain with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium. Young Control and Young ABX brains were tagged with HTOs 1-6 (HTO 1-3 for young control and HTO 4-6 for young ABX). Aged control and Aged ABX brains were tagged with HTOs 1-6 (HTO 1-3 for aged control and HTO 4-6 for aged ABX). Young and aged were loaded in separate wells on the 10X Chromium. The samples were tagged with unique nextera index primers to allow for preparation of one sequencing library consisting of all 12 samples and were separated in silico by it's unique hashing antibody and nextera index barcode combinations.