Profiling Immune Responses in Influenza-Infected Mice

Lung tissues of challenge-infected mice were collected at 4 dpi, and single-cell suspensions were used to analyze CD4+ T cell, CD8+ T cell, germinal center-like (GC) cell, and B cell populations [7 (link),24 (link)]. Harvested cells were stimulated with H5N1 and H3N2 inactivated virus antigens (0.5 ug/mL) for 5 h at 37 °C before staining with anti-CD3 (PeCy 5), CD4 (FITC), CD8 (PE), B220 (FITC), GL7(PE), CD19 (PeCy5), and IgD (PE) fluorophore-conjugated antibodies (BD Biosciences, San Diego, CA, USA) as described previously [31 (link)]. Cell acquisition and analyses were performed using the Accuri C6 flow cytometer and the C6 analysis software, respectively (BD Biosciences, San Diego, CA, USA).