Figure 149.

Gating strategy for Kupffer cell identification in LNPC samples isolated from C57BL/6 mice (9 weeks-old). Single cells were discriminated by plotting FSC-A against FSC-H (gate 1). Live cells were defined as DAPI negative (gate 2), while hepatic leukocytes are CD45⁺(gate 3). Note that in this gate liver sinusoidal endothelial cells (LSEC) can be identified as CD45⁻ CD31⁺. Gate 4 is then used to get rid of lineage⁺cells (CD3⁺, CD19⁺, CD49⁺, Ly6G⁺). Note that the concomitant use of CD11b allows the identification of neutrophils as Lineage⁺CD11b^high, while T and B cells are Lineage⁺CD11b⁻. F4-80⁺macrophages are then defined as CD11b^int F4-80⁺(gate 5). Finally, plotting I-A/I-E against TIM4 (gate 6) allows the discrimination between Kupffer cells (TIM4⁺, I-A/I⁻ Eint) and capsular macrophages (TIM4⁻, I-A/I-E⁺).