Goat anti rabbit igg hrp secondary antibody

A total of 10 paired cancer and paracancerous tissues were collected from December 2021 to March 2022 from gastric cancer patients who underwent surgical resection in the Department of Gastrointestinal Surgery at the Third Xiangya Hospital of Central South University, and written informed consent was obtained from all patients prior to the study. The research was approved by Ethics Committee of Xiangya Third Hospital, Central South University (22101). The methods for tissue protein extraction were as follows: tissue clipping, tissue homogenizer homogenization; lysis on ice with RIPA lysis solution; centrifugation; protein quantification using a BCA kit (KGPBCA; KeyGEN BioTECH, Jiangsu, China); appropriate amount of protein up-sampling; electrophoresis; gel cutting; membrane transfer; milk closure; addition of 1:800 dilution of rabbit anti-COMMD10 (123867; ZEN BIO, Chengdu, Sichuan, China); 1:1,000 dilution of rabbit anti-GAPDH monoclonal antibody (GB11002; Servicebio, Wuhan, Hubei, China); incubated overnight at 4 °C; washed the membrane; and added goat anti-rabbit IgG secondary antibody (HRP; Proteintech, Rosemont, IL, USA); incubated at 37 °C for 90 min; membrane washed; developed by ECL luminescence kit (BL520A; Biosharp, Beijing, China); developed and fixed; and the expression level of COMMD10 in cells and tissues was analyzed with GAPDH as the internal reference protein.

As mentioned above, the brain tissues were obtained at 7 days after HI injury and made into paraffin sections. Immunohistochemistry staining was performed as described previously (Wei et al., 2020 (link)). The paraffin-embedded sections were dewaxed and hydrated, and then were boiled in preheated sodium citrate buffer (C1010, Solarbio, Beijing, China) for 20 min and cooled to room temperature for antigen retrieval. The endogenous peroxidase was eliminated by 3% H₂O₂. After washed with PBS, sections were blocked with 10% goat serum (C0265, Beyotime, Shanghai, China) for 1 h at the room temperature. Then, sections were incubated with primary antibodies (listed in Supplementary Table S1) overnight at 4°C, and followed by the incubation of goat anti-rabbit IgG HRP secondary antibody (1:200, Proteintech, SA00001-2) for 1 h at room temperature. Finally, these slides were visualized with 3, 3′-diaminobenzidine DAB solution under the optical microscope (Nikon Corporatin, Tokyo, Japan).