Optiview dab ihc kit

Four µm sections were cut from FFPE tissue blocks. After deparaffinization, antigen retrieval was performed by CC1 antigen retrieval solution (pH 8.0, Ventana Medical systems, Tuczon, AZ, USA) on the Ventana BenchMark Ultra automated slide stainer. Slides were incubated with the primary PD-L1 antibody (anti-PD-L1, clone 22C3, Merck) at a dilution of 1/100 for 32 min, followed by visualization with the OptiView DAB IHC Kit and OptiView Amplification Kit (Ventana Medical systems). The specimens were then counterstained with haematoxylin and bluing reagent (Ventana Medical systems) and coverslipped.
PD-L1 expression was scored in immune cells in the paracortex and in the sinuses of the lymph node, excluding the germinal centers. Similar to IDO1 scoring, the intensity of PD-L1 staining in the paracortex was evaluated according to a four-tiered grading system (Supplementary Figure 3A): no expression (0), weak expression (1+), moderate expression (2+) or strong expression (3+). PD-L1 expression in the paracortex was dichotomized into an PD-L1-low group (0 and 1+) and an PD-L1-high group (2+ and 3+). In the sinuses, PD-L1 staining was scored as ‘low’ or ‘high’ (Supplementary Figure 3B).