Fcr specific blocking mab

Single‐cell suspensions of splenocytes and thymic cells were incubated with an FcR‐specific blocking mAb (eBioscience) and directly stained with fluorescently conjugated mAbs to murine CD3 (PerCP‐Cy5.5), CD4(Alexa Fluor 700), CD8 (FITC),F4/80 (FITC), CD19 (eFluor 405), CD11b (PE), CD11c (APC‐E780). Then, cells were analyzed by flow cytometer (FACS Aria II, BD Biosciences).

The mice were killed by CO₂ inhalation. To acquire single cells, lung was fragmented and digested with collagenase IV, and spleen was fragmented and filtered. Single‐cell suspensions of spleen and lung were incubated with FcR‐specific blocking mAb (eBioscience) and stained with FITC‐anti‐mouse CD3, Pacific blue‐anti‐mouse CD4, Brilliant Violet 605‐anti‐mouse CD8, APC‐anti‐mouse CD19, and Percp‐Cy5.5‐anti‐mouse CD335 (BioLegend, San Diego, CA, USA) and then examined by flow cytometry (LSRFortessa, BD Bioscience). The adhesion cells were excluded by FSC‐H/A and SSC‐H/A, and the lymphocytes were gated by FSC‐A and SSC‐A. The CD3⁺ cells were gated to determine the percentage of CD4⁺ and CD8⁺ cells, the CD3⁻ cells were gated to determine the percentage of CD19⁺ and CD335⁺ cells.