Cd49b pe clone dx5

Four days after RSV A2 challenge, neonatal and adult mice were sacrificed and their lungs harvested. Lungs were chopped and incubated with lung enzyme cocktail including 10% RPMI (Sigma, St. Louis, MO), collagenase D (Roche, Mannheim, Germany), and DNase1 (Roche) for 30 min at 37°C with stirring. After incubation, lung homogenates were transferred to a 70-μm strainer on a 50-mL conical tube and cells were analyzed by flow cytometry as described above. Anti-CD1d-APC (clone 1B1; eBioscience, San Diego, CA), CD3ε-PerCP-Cy5.5 (clone 145-2C11; BD Pharmingen), and CD49b-PE (clone DX5; BD Pharmingen) were used for lung cell staining.

Flow cytometry analysis was performed by using a BD FACSVerse (BD Biosciences). Human TILs were stained with the following monoclonal antibodies: CD45-APC-H7, CD3ε-PerCP, CD8α-V500, CD56-PE-Cy7, CD19-FITC, IFN-γ-FITC, IL-17A-V450 (all from BD Pharmingen, Milan, Italy), IL-21-PE, T-bet-PE-Cy7 (clone 4B10), RORγt-APC (clone AFKJS-9), Foxp3-FITC (clone PCH101), IL-22-APC, IL-6 PerCP (both from eBioscience), CD68-APC (Biolegend, San Diego, CA). Mouse LPMCs and TILs were stained with the following monoclonal antibodies: CD45-APC-Cy7, CD4-PerCP, CD8α-FITC, CD49b-PE (clone DX5), FoxP3-APC, IL-17A-PE, IFN-γ-PE-Cy7 (both from BD Pharmingen), perforin-APC, granzyme B-PE-Cy7 (all from eBioscience). In parallel, cells were stained with the respective control isotype antibodies.