Anti mouse cd45

Type II alveolar cells were isolated according to published procedures.^{11 (link)} In brief, murine lung tissues were perfused with 30 mL of PBS. The whole lung tissue was then injected with 3 mL dispase (5 U/mL, Stem Cell Technology, Vancouver Canada) and placed on ice. Tissues were finely minced, and additional 60 μL dispase (5 U/mL) and 7.5 μL DNase I (2 U/μL, Thermo Fisher Scientific, Fairlawn, NJ) was added prior to incubation at 37 °C for 45 min, with inverting every 5 min. Dissociated cells were pushed through a 100 micron cell strainer, followed by a 40 micron cell strainer (Corning, Corning, NY) to remove connective tissue. To the resulting cells, RBC lysis buffer (e-Biosciences, San Diego, CA) was added and mixed every 5 min at room temperature for a total of 30 min. The suspension was centrifuged, the supernatant was removed, and the cells were resuspended in 50:50 DMEM/F-12 media. The cells were plated on dual antibody coated plates prepared 24 h in advance with anti-mouse CD16/CD32 and anti-mouse CD45.1 (e-Biosciences). After incubation for 2 hours at 37 °C to facilitate negative selection for type II alveolar cells, the media containing the suspended type II alveolar cells was removed and pelleted by centrifugation. Cells were lysed and stored at −80 °C in 750 μL of Trizol Reagent (Thermo Fisher, Fairlawn, NJ).