Mts cell viability kit

MSCs were cultured in α-MEM with 10% FBS containing the BMP-2 released from the CAH/B2 scaffolds. When osteoblast differentiation was to be induced, 50 µg/mL of ascorbic acid and 5 mM β-glycerophosphate were added, and the culture media was changed every 3 days. To investigate the cytotoxicity of the CAH/B2 scaffolds to MSCs, the MTS assay was performed using the MTS cell viability kit (Promega) after 24, 48, 72 and 96 hr. Then the cells at different time points were processed in the MTS assay according to the user manual. OD values were measured at 490 nm using a microplate reader. CAH scaffold and plate group were set as controls.

The pan caspase inhibitor z-VAD-fmk (Biovision, USA) was used to inhibit caspases that are responsible for apoptosis induction. Briefly, 1 × 10⁴ cells/well/100 μL culture media were seeded in 96-well plate and incubated at 37 °C for 24 h. The cells were then treated with MP-HX and MP-EA extracts for 48 h in the absence and presence of z-VAD-fmk (4 μM and 8 μM). The concentration of the extracts used were at ~IC₅₀ values for HCT116, HCC1937, HepG2 and MDA-MB-231 cells, where MP-HX concentrations were 70 μg/mL, 90 μg/mL, 75 μg/mL, and 45 μg/mL, respectively, while MP-EA concentrations were 75 μg/mL, 90 μg/mL, 130 μg/mL and 90 μg/mL, respectively. After 48 h, the cell viability was determined using Promega’s MTS cell viability kit, following the manufacturer’s instruction.

A549 cells were seeded into 96-well plates at 5000 cells/well. After cell adhesion, the PC-specific siRNA or control RNA were then transfected into A549 cells using Lipofectamine 2000 reagent (Invitrogen). Forty-eight hours after transfection, MTS assays were performed using the MTS cell viability kit (Promega) according to the manufacturer’s instructions. Cell viability was then calculated.