Horseradish peroxidase hrp conjugated goat anti rabbit igg

The prepared paraffin wax blocks were sliced into sections, as described previously. BSA (Servicebio, Wuhan, China) was added dropwise to the sections, and then, the sections were incubated for 30 minutes. The following primary antibodies were added to the sections: rabbit anti‐rat Smoothened antibody (1:200; Affinity), rabbit anti‐rat Gli‐1 antibody (1:100; Novus, CO), rabbit anti‐rat collagen‐1 antibody (1:200; Servicebio), and rabbit anti‐rat collagen‐3 antibody (1:200; Proteintech, Wuhan, China). The sections were placed flat inside a wet box and incubated overnight at 4 °C. The next day, the tissues were covered with secondary antibody corresponding to the primary antibody at a specific dilution and incubated at room temperature for 50 minutes. Horseradish peroxidase (HRP)–conjugated goat anti‐rabbit IgG was added as a secondary antibody to the samples (1:200; Servicebio).
Finally, the samples were sealed with a fluorescence quencher. The slices were placed under a scanner (Pannoramic DESK) to capture images, and the fluorescence intensity of each image was analyzed using ImageJ software.