EBV Virus Production and Quantification

B95-8 cells (EBV producer cell line) [2 (link)] were cultured in RPMI-1640 (GIBCO, USA), supplemented with 10% FBS (GIBCO, USA), 1% antibiotic antimycotic solution (Santa Cruz, UK), 50 μg/ml gentamycin (Hyclone, USA) and 1× glutamine (GIBCO, USA), as previously described [58 (link)]. Cells were cultured until they reached a density of approximately 5×10⁶ cells/ml. Culture supernatants were centrifuged to remove cells/cell debris and then filtered using a 0.45 μm filter. Fresh filtered virus preparations were used for inoculation of rabbits. EBV copy number in the inoculum was estimated using quantitative real-time PCR (qPCR) (see ahead). The infectivity of the virus was determined by in vitro immortalization of human PBMCs [40 ]. This was approved by the Al Ain Medical District Human Research Ethics Committee (Approval number AAMD HREC 14/13).