Fig. 2.

Gating strategy to identify macrophage subsets in human corneas. Cells were harvested from enzymatically digested human corneas. CD45+ was used to identify leukocytes after exclusion of debris, doublets, and dead cells. Macrophages were identified by means of CD11b, CD14, and CD68 expression. A sequential gating strategy was used to identify macrophage subsets based on their surface markers: HLA-DR+, CD282+, and CD284+ cells were identified as M1 macrophages, whereas CD163+ and CD206+ cells were identified as M2 macrophages. CD86+ was only rarely detected. FMO controls (in red) of the corresponding gate (cells in light blue) were included to determine the baseline for positive events in the particular channels. FMO, fluorescence minus one; M1, pro-inflammatory macrophages; M2, regulatory macrophages.