2.7.5. Western Blot was Used to Detect the Expression of Proteins Related to the P13K/AKT/GSK-3β Signaling Pathway

After the right femur of each group of rats was crushed, 1 g of bone tissue was taken, and tissue lysate containing RIPA was added, and protease inhibitor (PMSF) was added at the same time for protein quantification after boiling water for about 15 minutes. Take 50 μg of each sample for sodium dodecyl sulfate (SDS) gel electrophoresis, transfer the membrane to polyvinylidene fluoride (PVDF) membrane by the semidry method, and seal with skim milk powder for 1 hour. The primary antibody was incubated overnight at 4°C, and the secondary antibody was incubated for 1 hour at room temperature. Then, add ECL and expose, use the image processing system to scan, and calculate the absorbance value of each band, and the ratio of it to the absorbance value of the internal reference strip represents the expression of P13K, AKT, P-AKt, and GSK-3β protein in each group of femoral bone tissues.