IC50 and Viability Assay Determination by CCK8 Assay

Ishikawa and HEC-1A cells at logarithmic growth stage were selected. The target cells were divided into DMSO group and drug group according to different treatment measures. The concentration of OSU-03012 (Selleck Chemicals, Houston, Texas, USA) was 0μL, 2.5μL, 5μL, 7.5μL, 10μL and 15μL, respectively. It was dissolved in dimethyl sulfoxide (DMSO, Solarbio, Beijing, China), and dilutions were made in McCoy’5A or DMEM to the desired in vitro concentrations. The maximum concentration of DMSO does not exceed 0.1% and will not influence cell growth. The single cell suspension was placed into 96-well plates and incubated in the incubator for 12 hours, 24 hours, 36 hours and 48 hours respectively. In the dark state, 10μL CCK-8 solution (dojindo, Shanghai, China) was added to each well, and then the plates were placed in the incubator at 37°C for 1 hour. The absorbance of each cell at 450nm was measured with a microplate reader (Thermo Fisher Scientific Inc.). The IC50 value and proliferation rate of cells were calculated by SPSS software.