Cell proliferation assay
MTT
assay was used to evaluate the inhibition of cell proliferation. Cells
were collected, adjusted to a cell concentration of 4.1 × 104 cells /
ml, and 100 microliters were seeded in a 96-well plate. After 24 hours
of incubation, NCTD with final concentrations of 0, 0.160, 0.312, 0.625,
1.25, 2.5, 5, 10, 20, 40, and 80 μM were added to each well. The group
without NCTD treatment was the control group. Then continue to incubate
for 72 hours, then add MTT solution (15.0 μl / well), and continue
incubation for 4 hours in a 5% CO2 incubator. Remove the culture plate
and add 150 μl of dimethyl sulfoxide (DMSO) to dissolve the purple
precipitate. A 96-well plate was analyzed on an automatic microplate
spectrophotometer (Thermo Molecular De-vices Co., United States) at a
wavelength of 560 nm. Calculate each cell (A560 control cells-A560
treated cells) / A560 control cells x 100% cytostatic rate. The IC50
value is calculated by the logit equation. Experiments were performed in
triplicate.Clone formation assay