Fig. 5
Production of infectious virus by NPC43 upon EBV lytic reactivation. a Expression kinetics of Rta, Zta and EA-D proteins in NPC43 cells upon TPA treatment. Expression of β-actin was included as the loading control. b The schematic diagram showing the workflow to determine whether infectious EBV particles were present in the supernatant of EBV+ve cells (including NPC43 and HONE1-EBV) upon lytic induction. Briefly the EBV+ve cells were induced to undergo lytic EBV reactivation. The cell-free supernatant which contains the infectious EBV was collected and was used to infect EBV–ve Akata cells or human primary B cells. The presence of EBV virions was confirmed by the respective experiments listed. c Identification of EBV+ve Akata cells by FISH analysis. The supernatant of NPC43 upon EBV lytic induction was collected for co-culture with EBV–ve Akata cells. At 96 h after co-culture, part of the infected Akata cells were subjected to FISH analysis. The presence of EBV genome was indicated by punctate red dots in Akata cell nucleus. Scale bar, 10 μm. d Determination of EBV gene expression by real-time PCR analysis in EBV-infected Akata cells. The supernatant collected from NPC43 or HONE1-EBV cells were subjected to co-culture with EBV–ve Akata cells for 96 h. EBV gene expression was evaluated afterwards. Akata cells infected by EBV from NPC43 cells after TPA treatment had a comparable level of EBV RNA expression with that from HONE1-EBV cells. Data are shown as mean ± SD from three independent experiments. e Proliferating foci of human primary B cells infected by NPC43-EBV after 28-day culture. Arrow suggests an NPC43-EBV-transformed B lymphoblastoid cell line. Scale bar, 100 μm
