**2.3 Staining of cell suspensions for flow cytometry analysis**

Monoclonal antibodies used for phenotypic analysis of DC and T cells were anti-CD3-FITC (BD Pharmingen 553062), anti-CD4-PerCP (BD Pharmingen 553052), anti-CD8a-APC (BD Pharmingen 553035), PD-1-PE (BD Pharmingen 551892), anti-CD11c-APC (BD Pharmingen 550261), anti-CD40-PE (BD Pharmingen 553791), anti-MHCII-FITC (BD Pharmingen 553623), anti-Ly6c-A700 (e-biosciences 56-5981-32), and Streptavidin-conjugated with PerCP fluorochrome (SAV-PerCP, BD Pharmingen 554064), biotinylated anti-CD103 (R&D Systems BAF1990). Cell suspensions were prepared by disgregating the organs using a 70m cell strainer (BD Falcon 352350) and the piston of a 3mL Syringe (BD 309585). Spleen, Lungs, BAL and Mediastinal lymph nodes cell suspensions were washed, incubated 10 min at 4°C with Power Block reagent (Biogenex, HK085-5K) to block Fc receptors, washed, and stained with fluorochrome-coupled mAbs for 15 min at 4°C. Cells were centrifuged and resuspended in FACS buffer. 106 and 105 live MHC-II high or CD3+ cell cells were acquired respectively. Data was acquired in a Dako Cyan Flow Cytometer and analyzed with FlowJo Software 7.2 (Tree Star, Inc., San Carlos, CA).
