Brucella melitensis infectious foci in liver.

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Brucella melitensis infectious foci in liver. Brucella melitensis infectious foci in liver. At 3 weeks post-infection, pathogenic B. melitensis-infected cells developed multiple granulomas (A,D; H&E staining). Immunohistochemical analysis of bacterial foci was performed using biotinylated primary antibodies (1.0 μg/ml, #TC-7011, Tetracore Inc.), endogenous biotin blocking, and streptavidin-conjugated to horseradish peroxidase (St∼HRP) using 3,3′-diaminobenzidine (DAB) as the final chromogen (B,E; brown precipitate). Staining of normal liver of healthy non-infected mice produced no signal (not shown). Staining of liver tissue at 3 weeks post-infection using the same protocol but substituting biotinylated anti-Brucella antibodies with normal serum produced no signal (F). Immunohistochemical analysis of macrophage lineage cells using antibodies to IBA-1 (dilution 1:500, #019-19741, Wako Chemicals USA, Inc.) followed by HRP rabbit polymer conjugate (#87-9263, Invitrogen Life Technologies) revealed positive staining of cells at the granuloma periphery (C). Residential macrophages – Kupffer cells were also positive for IBA-1 staining (C). Although Brucella was found in the granuloma core, these core cells were basically negative for IBA-1 staining (compare B versus C). Sections were counterstained with hematoxylin (B,C,E,F). Scale bars: 20 μm (A,B,C,E), 50 μm (F) and 200 μm (D). Diogo M. Magnani et al. Dis. Model. Mech. 2013;6:811-818 © 2013. Published by The Company of Biologists Ltd