Rabbit specific ihc polymer detection kit

Hematoxylin and eosin (H&E) staining was performed as described previously (11 (link)). Briefly, skin tissue was fixed in 10% buffered formalin and embedded in paraffin, then cut into 5-μm sections using a microtome (HM 325, ThermoFisher, Grand Island, NY, USA). At least 3 independent skin tissue sections from each group were H&E stained and examined for histological changes using a Keyence Fluorescence Microscope Model BZ-X710 (Osaka, Japan).
Immunohistochemistry (IHC) was conducted with a rabbit-specific IHC polymer detection kit (Abcam, Cambridge, MA, USA) per the manufacturer’s protocol. The macrophage marker F4/80 was detected using antibody (Cat No. ab100790, Abcam) with 3,3’-diaminobenzidine substrate (Abcam), and slides were examined under the microscope.

Whole tumor Sects. (4-μm-thick) of formalin-fixed paraffin-embedded tissue were deparaffinized in xylene and rehydrated in ethanol and distilled water. Antigen retrieval was performed by microwaving sections in Universal HIER antigen retrieval reagent (Abcam) for 20 min. Endogenous peroxidase activity was blocked by treatment with Peroxidase Block (DAKO, Santa Clara, CA, USA) for 10 min, and non-specific binding was blocked by treatment with Protein Block serum-free Ready-to-use (DAKO) for 10 min at room temperature. The sections were then incubated overnight at 4 °C with anti-human PD-L1 monoclonal antibody (clone 28–8, 1:500; Abcam). The sections were then incubated with Rabbit-specific IHC polymer detection kit and visualized using DAB substrate (both Abcam). Finally, sections were counterstained with hematoxylin. For negative control staining, the anti-PD-L1 primary antibody was replaced with a rabbit IgG monoclonal antibody (Abcam).