Ab16054

Bone tissue sample were ground with liquid nitrogen and then mixed with RIPA lysate solution. After sufficient lysis, the mixture was centrifuged for 10 min, and the supernatant was collected. Measure the protein concentration using the BCA kit (KeyGEN Bio TECH, China). Then, 5 × SDS loading buffer was added to the extracted proteins, which were subsequently heated in a boiling water bath for 5 min. Equal amounts of protein lysates were separated by 10% SDS-polyacrylamide gel electrophoresis (PAGE) and transferred onto polyvinylidene difluoride membranes. The membranes were incubated overnight at 4 °C with anti-VEGF (1:1000, Boster, BA0407), anti-Notch1 (1:1000, Abcam, ab52627) and anti-Noggin antibodies (1:1000, Abcam, ab16054). After washing with TBST buffer, horseradish peroxidase-labeled Goat Anti-rabbit (ZSGB-Bio, ZB2301) and Anti-Mouse (ZSGB-Bio, ZB2305) antibodies, diluted at 1:5000, were added and incubated for 1 h at room temperature. The membrane was then treated with an appropriate amount of ECL luminescent liquid (Beijing Dingguo Changsheng Biotechnology Co., Ltd., China) and imaged using an integrated chemiluminescence instrument (ChemiScope 5300 Pro).
(We have provided an unprocessed original version of the Western blot experimental images in Additional file 1.)