To assess osteoblast differentiation, alkaline phosphatase (ALP) activity was determined as described previously52 (link). In brief, differentiated osteoblasts were washed with phosphate-buffered saline (PBS) and incubated with a solution containing 6.5 mM Na2CO3, 18.5 mM NaHCO3, 2 mM MgCl2, and a phosphatase substrate (Sigma, S0942). ALP activity was measured using a spectrophotometer. ALP staining was performed using Fast Blue (Sigma, FBS25) and Naphthol AS-MX (Sigma, 855) after fixation in 10% neutral formalin buffer.63 (link)
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Osteoblast Differentiation Analysis
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Other organizations : Worcester Polytechnic Institute, Baba Ghulam Shah Badshah University, University of Massachusetts Chan Medical School
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Variable analysis
independent variables
- Differentiation of osteoblasts
- Alkaline phosphatase (ALP) activity
- ALP staining
- Phosphate-buffered saline (PBS) wash
- Solution containing 6.5 mM Na2CO3, 18.5 mM NaHCO3, and 2 mM MgCl2
- Phosphatase substrate (Sigma, S0942)
- Fast Blue (Sigma, FBS25) and Naphthol AS-MX (Sigma, 855) staining
- 10% neutral formalin buffer fixation
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