Figure 2.

Detection and quantification of NETs in the lungs of infected mice. Detection of NETs induced in lung tissues of mice infected with lethal doses of influenza virus followed by S. pneumoniae challenge. (A) For identification of NETs, lung sections were stained with histone H2B (green), MPO (red), and DAPI (blue). (B) NETs formation was analyzed microscopically based on triple immunostaining, and by their morphologic characteristics, i.e., appearing as individual NETS, small, medium, or large clusters of NETs. All scale bars represent 50 μm. (C) A representative cluster with area demarcated for calculation by ImageJ software. (D) Quantification of the total NETs score was obtained from at least 20 fields of each lung section. Values represent the means ± SE of 6 animals per group. ^*Indicates P < 0.05; N.S. = Not significant difference.