A. Western blotting of total protein extracts from primary cultured human normal urothelial cells (HNUC), SV40T-immortalized urothelial cells (UROtsa), cell lines derived from low (RT4) and moderate (RT112) grade human UC, and cell lines derived from high-grade human UC (J82, T24, T24T (a metastatic derivative of T24), and UMUC3). Note that, with the exception of UMUC3, cell lines derived from both low- and high-grade human UC overexpress PKM2, but not PKM1, compared with primary cultured and immortalized human urothelial cells. B. Immunofluorescence staining of T24 cells with DAPI counterstaining showing both cytoplasmic and nuclear localization of PKM2. Magnification: 400x.