Isolation and Culture of Renal Proximal Tubule Cells

Primary renal proximal tubule epithelial cells were obtained from ATCC (American Type Culture Collection) and were grown in renal epithelial cell basal medium (RECBM) supplemented with one Renal epithelial cell growth kit as suggested by the manufacturer (ATCC). For JCPyV infection, we used a lab-adapted strain referred to as Mad-1/SVEΔ, which was described previously (90 (link), 91 (link)). For BKPyV infection, we used the Dunlop strain purchased from ATCC. JCPyV and BKPyV were grown in SVGA cells (human glial cells transformed with SV40 large T antigen) and Vero cells, respectively, using 1,700-cm² roller bottles. Cells were cultured for 14 days, with the cell culture medium replaced at 7 days. Viral lysates were harvested by scraping cells in the presence of cell culture medium, and this lysate was frozen and thawed 3 times. When needed, JCPyV and BKPyV lysates were purified as previously described (92 (link)).