2.7 Pathological examination and virus protein antigen
detection
Respiratory tract tissues of mice and lung tissues of humans were
dehydrated, embedded, and serially sectioned with a thickness of 4 μm.
The sections were stained with HE, and the pathological changes were
observed under an optical microscope. An immunohistochemical method for
the detection of viral protein was conducted as previously described (J.
Wang et al., 2016). After antigen repairing, the primary nucleoprotein
(NP) antibody (1:500) (kindly provided by the National Institute of
Diagnostics and Vaccine Development in Infectious Diseases, Xiamen
University) was added in sections. The sections were incubated overnight
at 4°C, followed by the addition of goat anti-mouse IgG-specific biotin
conjugate (Calbiochem) (1:50), development by DAB stain, and
counterstaining slightly by hematoxylin. Human lung tissue sections
infected with influenza virus H5N1 were used as a positive control, and
10% normal mouse serum was used as a MOCK control. The positive results
were judged by the brownish coloring of the nucleus, and the cytoplasm
around the nucleus appeared slightly brownish in color.