After blood sample collection, tissues of lung and liver were obtained, fixed with 10% formalin for 24 h, and embedded in paraffin. The 5-μm sections were cut and placed on glass slides. To perform hematoxylin and eosin (H&E) staining, these samples were deparaffinized with xylene and rehydrated in graded ethanol. Subsequently, the sections were stained with H&E and observed under a light microscope (Carl Zeiss, Jena, Germany). Five random fields were selected from one slide for scoring, and morphological changes were scored according to a previous study [18 (link)].
For immunohistochemistry analysis, after rehydration in graded ethanol, sections were performed antigen retrieval with the microwave. Then, the slides were incubated with primary antibodies, including anti-CD86 (Cell Signaling Technology, Danvers, MA, USA; Cat. No. 19589, 1:300), anti-CD206 (Cell Signaling Technology, Cat. No. 24595, 1:500), and anti-p-NFκB p65 (p-p65; Santa Cruz Biotechnology, Santa Cruz, CA, USA; Cat. No. sc-166748, 1:200), followed by incubation with secondary antibodies and diaminobenzidine. Lastly, the nuclei were counterstained with hematoxylin. Images were obtained under a light microscope, and the expression level was calculated by the H score system [19 (link)].
For immunohistochemistry analysis, after rehydration in graded ethanol, sections were performed antigen retrieval with the microwave. Then, the slides were incubated with primary antibodies, including anti-CD86 (Cell Signaling Technology, Danvers, MA, USA; Cat. No. 19589, 1:300), anti-CD206 (Cell Signaling Technology, Cat. No. 24595, 1:500), and anti-p-NFκB p65 (p-p65; Santa Cruz Biotechnology, Santa Cruz, CA, USA; Cat. No. sc-166748, 1:200), followed by incubation with secondary antibodies and diaminobenzidine. Lastly, the nuclei were counterstained with hematoxylin. Images were obtained under a light microscope, and the expression level was calculated by the H score system [19 (link)].
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