Cell and tissue staining hrp dab kit

The tissues were fixed, dehydrated, embedded in paraffin (Servicebio), and cut into 4-µm specimens. Subsequently, the specimens were deparaffinized using xylene and rehydrated with graded alcohols. After restoration with sodium citrate and blocking using hydrogen peroxide and bovine serum albumin (Servicebio), the specimens were incubated with the primary antibodies, including KI67, proliferating cell nuclear antigen (PCNA), AKT1, mTOR, HIF1α, vascular endothelial growth factor (VEGF), and Glut1 (Proteintech, Wuhan, China) for 12 h at 4°C. Subsequently, the sections were immunohistochemically stained with horseradish peroxidase-conjugated secondary antibodies (Servicebio) for 2 h at room temperature. After incubation with the Cell and Tissue Staining HRP-DAB Kit (Servicebio), and orthophotomicroscope (Olympus, Japan) was used to capture the images.

After fixation using 4% paraformaldehyde for 24 h at room temperature, dehydration and embedding in paraffin (Servicebio), the tissues were cut into 4-µm sections. The specimens were then deparaffinized using xylene and rehydrated through graded alcohols. After restoration with sodium citrate and blocking using H₂O₂ and bovine serum albumin (Servicebio), the specimens were incubated with the primary antibodies (both 1:400), including Ki-67 (cat. no. 27309-1-AP) and proliferating cell nuclear antigen (PCNA; cat. no. 10205-2-AP; ProteinTech Group, Inc.) for 12 h at 4°C. Subsequently, the sections were immunohistochemically stained with horseradish peroxidase (HRP)-conjugated goat anti-rabbit secondary antibodies (cat. no. G1213; Servicebio) for 2 h at room temperature. Following incubation with the Cell and Tissue Staining HRP-DAB kit for 5 min at room temperature (Servicebio), an orthophotomicroscope (Nikon Corporation; magnification, ×400) was used to capture images.

Osteosarcoma and adjacent tissues were fixed, dehydrated, and embedded in paraffin (Servicebio), and then cut into 2‐µm specimens. Specimens were deparaffinized and rehydrated with graded xylene and alcohols, respectively. After restoration by sodium citrate and blocking using H₂O₂ and bovine serum albumin (Servicebio), the specimens were incubated with primary anti‐CHRDL2 antibody (Cat No. ab198786; Abcam) and anti‐BMP‐9 antibody (Cat No: ab30588; Abcam) for 16 h at 4°C. Subsequently, the sections were immunohistochemically stained with horseradish peroxidase‐conjugated goat anti‐rabbit antibodies (Servicebio) for 2 h at room temperature. After incubation with the Cell and Tissue Staining HRP‐DAB Kit (Servicebio), an orthophoto light microscope was used to collect images (magnification ×200 and ×400).