MTT Assay for Zoledronic Acid Sensitivity

The MTT method was applied to test the sensitivity of PDLSCs to ZOL as previously reported with some adjustments^{27 (link)}. Briefly, cells (2 × 10³) were seeded into 96-well plates (Eppendorf, Milan, Italy) with 100 µL culture medium per well. Each drug concentration was tested on 16 wells. After 3 and 6 days of treatment, 10 µL MTT solution (M2128, Sigma, Milan, Italy) (2.5 mg/mL) was added to each well. Plates were incubated at 37°C for 3 h and then the supernatant was removed. Formazan crystals were solubilized by adding 0.08N HCl (H1758, Sigma, Milan, Italy) in isopropanol (I9516, Sigma, Milan, Italy) for 30 min at 37°C. The absorbance was measured at 595 nm using a microplate reader (iMark, Bio-Rad Laboratories, Milan, Italy). Values obtained in the absence of cells were considered as control.