In vitro stimulation of lung classical monocytes

B6 mice and CD93 deficient mice were intranasal injected with 100ug HDM on Day 0 and sacrificed on Day 3. Lung classical monocytes were sorted by BD FACSAria, the gating strategy for classical monocytes is single cell Live CD45+ Ly6Glo Siglecf- CD11b+ Ly6Chi. And then monocytes were cultured in completed medium as negative control, or completed medium with 1ug per ml IL-17D as stimulated group. After 15h culture, TRIzol reagent were added and the RNA samples were collected for library construction and sequencing.