Anti opg antibody

Femur specimens were set in 10% neutral formaldehyde and embedded in 4 μm thick paraffin. After dewaxing, the sections were prepared with sodium citrate buffer, washed in 1X phosphate‐buffered saline (PBS), and incubated overnight at 4°C with anti‐COX2 antibody (Abcam) or anti‐OPG antibody (Abcam) diluted in PBS containing 3% BSA. Anti‐mouse HRP‐conjugated secondary antibody (DAKO) was added, and the specimens were incubated at 37°C for 30 minutes. The scoring criteria are as follows: percentage of positive cells (0%‐5%, 0; 6%‐25%, 1; 26%‐50%, 2; 51%‐75%, 3; 76%‐100%, 4), positive staining intensity (negative, 0; yellow, 1; brownish yellow, 2; brown, 3), and the final total score is equal to the percentage of positive cells multiplied by the positive staining intensity.

Total Drynariae Rhizoma flavonoids (DRTF) were purchased from Beijing Qihuang Pharmaceutical Manufacturing Co., Ltd. (National Medicine Permit No. Z20030007, number of production: 04080081, the content of DRTF ≥ 80%). Alpha-Minimum Essential Medium (α-MEM) was purchased from Life Technologies (USA). Fetal bovine serum (FBS) was obtained from ExCell Biology, Inc. (China). Ascorbic acid (AA), β–glycerophosphate (β-GP) and Alizarin Red S were purchased from Sigma-Aldrich (USA). Rat turnover markers ELISA kits were purchased from Biocalvin (Suzhou, China). RIPA lysis solution and a BCA protein assay kit were purchased from HEART biological technology Co., Ltd. (China). The RNAprep Pure Tissue Kit was purchased from TianGen Biotech Co., Ltd. (China). The PrimeScript™ RT reagent kit (Perfect Real Time) and SYBR Premix Ex Taq™ reagent were purchased from TAKARA Biotechnology (China). Anti-OPG antibody was purchased abcam (USA). Anti-ERα and anti-ERβ antibodies were purchased from R&D Systems (USA). Anti-GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) antibody was obtained from Proteintech (China). Anti-RANKL and Horseradish peroxidase (HRP) conjugated antibodies were purchased from Ruiying bio (China).