The alkaline phosphatase activity (ALP) was determined after 7 days and 14 days of culture on cell lysates using a commercial kit (Alkaline Phosphatase Activity Colorimetric Assay Kit, BioVision; Milpitas, CA, USA), as presented in a previous paper [67 (link)]. Alkaline phosphatase activity was calculated according to the formula:
where A represents the amount of p-nitrophenol (pNP) expressed by the samples (in μmol), V is the volume of lysate used in the reaction (in mL), and T represents the reaction time (in minutes).
To avoid variations due to different protein concentrations, all samples were reported to the protein concentration which was previously determined using the Bradford method. Thus, the concentrations of the reaction product (p-nitrophenol) were normalized to 1 μg protein.
where A represents the amount of p-nitrophenol (pNP) expressed by the samples (in μmol), V is the volume of lysate used in the reaction (in mL), and T represents the reaction time (in minutes).
To avoid variations due to different protein concentrations, all samples were reported to the protein concentration which was previously determined using the Bradford method. Thus, the concentrations of the reaction product (p-nitrophenol) were normalized to 1 μg protein.
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