Cell Culture Protocols for RPTEC and HNSCC

Renal proximal tubule epithelial cells (RPTEC) and HPV− HNSCC cell lines, FaDu and A253, were obtained from ATCC. Cell lines were authenticated using short tandem repeat DNA profiling. All experiments were conducted with Mycoplasma-free cells that had undergone less than five passages. RPTECs were cultured in renal epithelial cell basal medium (#PCS-400-030) supplemented with renal epithelial cell growth kit (#PCS-400-040) and 2% FBS (ATCC). Cancer cell lines were cultured in RPMI1640 medium supplemented with 10% FBS. All cells were grown at 37°C in 5% carbon dioxide. Harvesting of cells was performed by washing with PBS then incubating cells in a 0.25% trypsin, 2.21 mmol/L Ethylenediaminetetraacetic acid (EDTA), and 1X sodium bicarbonate solution at 37°C. Cell counts were performed using a Beckman Coulter Vi-CELL XR Cell Viability Analyzer (Beckman-Coulter).