"Generation of Delta-24-GREAT. ( A ) Schematic representation of Delta-24-GREAT genomic structure. pCMV , cytomegalovirus promoter; BGH pA , bovine growth factor polyadenylation signal; ITR , inverted terminal repeat. ( B and D ) Expression in vitro of the GITR ligand (GITRL) on the surface of Delta-24-GREAT-infected murine glioma cells <t>GL261-5</t> (100 MOIs) (B) and human brain tumor stem cells GSC17 (25 MOIs) ( D ) assessed by FACS, 48 hours after infection. Mock-infected cells stained with IgG isotype as primary antibody, and UV-inactivated Delta-24-GREAT-infected cells were used as control. Non-viable cells were excluded from the analysis using ethidium homodimer. Data are shown as mean ± SD of three independent experiments. α, antibody. (C and E ) Expression of viral proteins, E1A and fiber, and murine GITRL, assessed by western blotting, in Delta-24-GREAT-infected murine glioma GL261-5 cells ( C ) and human glioma U-87MG cells (E) 48 hours after infection at the indicated MOIs. Mock, Delta-24-RGD, and UV-inactivated Delta-24-GREAT were used as controls at the highest MOI. GAPDH expression was used as loading control. "