Cd11c positive selection magnetic bead kit

CD11c⁺ cells were enriched from collagenase/DNAse-digested lungs on days 0 or 3 p.i. using a CD11c positive selection magnetic bead kit (StemCell Technologies). To sort lung DC subsets on day 5 p.i., DCs within a lung digest were enriched by gradient separation with OptiPrep (Axis-Shield) according to the manufacturer’s protocol. DC subsets were identified and sorted after staining with mAbs to DC surface markers listed above as in Fig. S2b. In KO mice, Irf4-deficient DCs were sorted as GFP⁺, since not all Ly6C⁺ iDCs were GFP⁺. Tet⁺CD8⁺ MPECs (GFP-) in lung and CD8⁺ T cells in the mLN were sorted on day 10 p.i. after enrichment on a 40% Percoll gradient and staining with mAbs specific for CD3-PECy7 (145-2C11), CD8a-APC or APCfire750 (53-6.7), IL7Rα-BIO (A7R34), KLRG1-BV421 (2F1/KLRG1), Streptavidin BV605 and H-2D^b- NP366-374-PE tetramer as in Fig. 3. T cell and DC subsets were sorted directly into ARCTURUS PicoPure RNA extraction buffer (ThermoFisher Scientific) using a BD FACS Aria II instrument.