Renal Cancer Cell Viability Assay

Renal cancer cell lines (Caki-1, Caki-2, A498) were obtained from the Korean Cell Line Bank (Seoul, Korea) and American Type Culture Collection (Manassas, VA, USA), and cultured in McCoy’s 5A and RPMI1640 supplemented with 10% fetal bovine serum. VHL-null RCC4 cell lines were kind gift from Dr. Jong-Wan Park [38 (link)], and cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal bovine serum. Cells were incubated in a humidified atmosphere at 37 °C under normoxia (20% O₂ and 5% CO₂) or hypoxia (1% O₂ and 5% CO₂). Cell viability assay was performed using crystal violet staining [39 (link)]. Briefly, cells were cultured in 24-well tissue culture dishes with or without drug treatment. After incubation with drugs, washed cells were fixed using 4% paraformaldehyde, and then stained with 0.5% crystal violet solution for 20 min at room temperature. Optical density of crystal violet was analyzed using 1% SDS solution and measured by an absorbance reader (BioTek, Winooski, VT, USA) (OD570).