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In vitro osteoclast differentiation assay

BS Bing Shu
YZ Yongjian Zhao
SZ Shitian Zhao
HP Haobo Pan
RX Rong Xie
DY Dan Yi
KL Ke Lu
JY Junjie Yang
CX Chunchun Xue
JH Jian Huang
JW Jing Wang
DZ Dongfeng Zhao
GX Guozhi Xiao
YW Yongjun Wang
DC Di Chen
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BMM cells were isolated from long bone of 1-month-old WT mice and plated into 24-well culture plates at a density of 4 × 105 cells per well and cultured in αMEM with 10% FBS. Cells were treated with 20 ng·mL−1 macrophage colony-stimulating factor (M-CSF) for 3 days, and then switched to the medium with 10 ng·mL−1 M-CSF and 50 ng·mL−1 RANKL for 7 days. To test Axin1Osx CM, BMM cells were treated with CM collected from cultured calvarial osteoblasts isolated from Axin1Osx KO mice and Cre-negative controls for 7 days. The culture medium was changed every 3 days. TRAP staining was performed using a TRAP assay kit (Sigma, St. Louis, MO, USA).

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