Pv6000 histostain kit

The wound healing samples in vivo or cultured HS tissues ex vivo were embedded in paraffin blocks and cut into 4-μm thick tissue sections. One section was used for routine H&E staining and another was used for Masson’s trichrome staining analysis of collagen fibers. For immunohistochemistry staining, sections were dewaxed and endogenous peroxidase activity was quenched with 3 % hydrogen peroxide for 15 min. The sections were then blocked with normal goat serum for 30 min to eliminate non-specific binding and incubated overnight at 4 °C with primary antibodies against Col1 or Col3 (1:100; Abcam). On the next day, the sections were treated with a PV6000 Histostain™ kit (ZSGB, Beijing, China) and stained with diaminobenzidine (ZSGB, Beijing, China). Finally, the sections were counterstained with hematoxylin. An isotype-matched IgG served as the negative control for each immunostaining procedure. Sections were analyzed and images acquired with FSX100.

The samples were fixed with 4% paraformaldehyde, dehydrated in graded ethanol, embedded in paraffin, and then cut into 5-μm-thick sections. H&E and Masson’s trichrome staining were used to detect the histological change and collagen deposition. For immunohistochemistry staining, the sections were immersed in 3% H₂O₂ after deparaffinization to eliminate the activity of endogenous peroxidase at 37 °C for 15 min and blocked with 5% BSA in PBS for 1 h to exclude the non-specific binding. Then, the slides were incubated with the primary antibodies against α-SMA and IL-17RA overnight at 4 °C. The next day, the slides were incubated with a PV6000 Histostain™ kit (ZSGB, Beijing, China) and stained with diaminobenzidine (ZSGB, Beijing, China). Images were obtained by FSX100 Bio Imaging Navigator (Olympus, Tokyo, Japan).