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Alkaline phosphatase (ALP) activity and quantification

BA Basem M. Abdallah
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ALP activity was performed after 6 days of osteoblastic induction. Cell viability was determined using the Cell Titer-Blue cell viability assay according to the manufacturer’s instructions (Promega, USA) and the viability measured at 560Ex/590Em nm using a FLUO star Omega plate reader (BMG Laboratories). ALP activity was determined following incubation with 1 mg/ml of P-nitro phenyl phosphate in 50 mMNAHCO3 and 1 mM MgCl2 buffer (pH 9.6) at 37 °C for 20 min. The activity was stopped by addition of 3 M NaOH. The reaction absorbance was measured at 405 nm using a FLUO star Omega plate reader and ALP activity corrected for cell viability.

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