Ds u3 camera interface

IgA⁺ cells were identified with the immunohistochemical staining method as described previously [23 (link)]. Briefly, fixed lung tissues were first embedded in paraffin, sectioned at 6 µm thickness, and mounted onto poly-lysine-coated glass slides. The slides were then washed with 0.01 M PBS (pH 7.4). Endogenous peroxidase activity was blocked by incubation with 3% H₂O₂ in methanol for 20 min. After rinsing three times with 0.01 M PBS (pH 7.4), the slides were then incubated sequentially with goat anti-mouse IgA antibody (1:400) and HRP-conjugated rabbit anti-goat IgG (1:250). 3,3′-diaminobenzidine (DAB) detection was performed according to the manufacturer’s instructions. After immunohistochemical staining, the slides were counterstained with hematoxylin, dehydrated with alcohol, cleared with xylene, and sealed with neutral gum. Images were captured with a Nikon DS-U3 camera interface and measured using image analysis software Image-Pro Plus 6.0. Three fields of each slide were randomly selected at ×200 magnification and the integrated optical density (IOD) was analyzed. The average value was calculated as the IOD value of the lung specimen.

IgA⁺ cells were identified with an immunohistochemical staining method described previously [15 (link)]. Briefly, fixed intestinal samples were embedded in paraffin, sectioned at a thickness of 6 µm, and mounted onto glass slides. Endogenous peroxidase activity was blocked by incubation with 3% H₂O₂ in methanol for 20 min. After rinsing three times with PBS, the slides were successively incubated with a goat anti-mouse IgA antibody (1:400) and HRP-conjugated rabbit anti-goat IgG antibody (1:250). 3, 3′-Diaminobenzidine (DAB) (BOSTER, Wuhan, China) detection was performed according to the manufacturer’s instructions. Images were captured with a DS-U3 camera interface (Nikon, Tokyo, Japan) and evaluated using the image analysis software Image-Pro Plus 6.0. Three fields per slide were randomly selected at 400× magnification, and the integrated optical density (IOD) was analyzed.
After hematoxylin–eosin staining, the number of iIELs in five different fields of intestinal villi on each slide was counted to statistically analyze the data.