Dakocytomation labelled streptavidin biotin reagent

Selected sections were deparaffinized, rehydrated, and heated in a microwave oven in 0.01 M citrate buffer (pH 6.0; Química Contemporânea, Diadema, Brazil) for 30 min. Endogenous peroxidase activity was blocked by 3% hydrogen peroxide for 10 min, followed by a wash with phosphate buffered saline. The sections were incubated overnight at 4°C with the antibodies. The antibody was then detected using avidin-biotin peroxidase detection solution (Dakocytomation labelled streptavidin biotin reagent; Dakocytomation, Glostrop, Denmark and System-horseradishperoxidase; Dako, Glostrup, Denmark) and the signal was visualized using diaminobenzidine (Dakocytomation) and Substrate Chromogen-System (Dako). Slides were counterstained with Harris’s hematoxylin, dehydrated, cleared and mounted. Positive controls from the appendix and tonsils were used. The cells were initially observed at a low magnification (×100) to assess the general distribution of the antibody. The samples were subsequently examined at a higher magnification (×400). The evaluation of cell staining was performed in tumor tissue. The tumor cells (exhibiting gross and evident nucleoli, and irregular chromatin) were identified and counted at the higher magnification.

Selected liver sections were deparaffinized, rehydrated, and heated in a microwave oven in 0.01 M citrate buffer (pH 6.0; Química Contemporânea, Diadema, Brazil) for 30 min. Endogenous peroxidase activity was blocked by 3% hydrogen peroxide for 10 min, followed by a wash with PBS. Sections were incubated overnight at 4°C with the following primary antibodies: anti-FAS (1 : 50), anti-OX62 (1 : 10), and anti-TNFR1 (1 : 100). The primary antibody was then detected using avidin-biotin peroxidase detection solution (DakoCytomation labelled streptavidin biotin reagent; DakoCytomation, Glostrop, Denmark; and System-horseradishperoxidase; Dako, Glostrop, Denmark), and the signal was visualized using diaminobenzidine (DakoCytomation) and Substrate Chromogen System (Dako). Slides were counterstained with Harris's hematoxylin, dehydrated, cleared, and mounted. Positive controls from the appendix and tonsils were used. Cells were initially observed at a low magnification (×100) to assess the general distribution of the primary antibody. Samples were subsequently examined at a higher magnification (×400) [16 (link)]. Liver cells (exhibiting gross and evident nucleoli and irregular chromatin) were identified, and stained cells were counted at the higher magnification.