Signalstain boost ihc anti rabbit hrp reagent

Formalin-fixed, paraffin-embedded tissue samples were sectioned with a standard microtome at 3- to 5-μm thickness. After deparaffinization and rehydration, heat-induced (98°C) antigen retrieval was performed in a 10 mM sodium citrate buffer (pH 6.0) at a sub-boiling temperature for 10 min. The slides were incubated with hydrogen peroxide 3% (v/v) for 10 min, washed and blocked with 5% FBS in TBST for 1h at room temperature. Next, slides were incubated with primary antibodies anti-ITGA2 (1:500) (Abcam) at 4°C overnight. Primary antibodies were detected using a SignalStain® Boost IHC anti-rabbit HRP Reagent (Cell signaling Technology). The signal was visualized using a diaminobenzidine substrate kit (DAB, Thermo Fisher Scientific) according to the manufacturer’s instructions and nuclei were counterstained with hematoxylin.

Formalin-fixed, paraffin-embedded tissue samples were sectioned with a standard microtome at 3- to 5 µm thickness. After deparaffinization and rehydration, heat-induced (98°C) antigen retrieval was performed in 10 mM sodium citrate buffer (pH 6.0) at a sub-boiling temperature for 10 mins. The slides were incubated with hydrogen peroxide 3% (v/v) for 10 min, washed and blocked with 5% FBS in TBST for 1 hr at room temperature. Next, slides were incubated with primary antibodies anti-COL1A1 (1:100) (Sigma-Aldrich) and anti-ITGA2 (1:500) (Abcam) at 4°C overnight. Primary antibodies were detected using a SignalStain Boost IHC anti-rabbit HRP Reagent (Cell signaling Technology). The signal was visualized using a diaminobenzidine substrate kit (DAB, Thermo Fisher Scientific) according to the manufacturer's instructions and nuclei were counterstained with hematoxylin. Immunostaining was scored by the weighted average score (intensity: 0–3, coloring: 0–100% of ITGA2 expression) by two trained scientists independently and discrepancies were resolved by consensus.