Generation of Lymphoblastoid Cell Lines (LCLs)

Epstein-Barr virus (EBV) is a lymphotropic γ herpes virus. Infection of human B cells with EBV in vitro results in their immortalization and the resulting cell lines are named lymphoblastoid cell lines (LCLs) (1 ). In these cells, EBV establishes mainly a latent infection, characterized by the expression of a limited number of viral proteins. LCLs express 6 EBV nuclear proteins (EBNA1 to 6), 3 membrane proteins (LMP1, LMP2A, and LMP2B) and two small untranslated nuclear RNA molecules (EBER1 and EBER2) (reviewed in refs. 2 , 3 ). LCLs have the phenotype of highly activated B cells as assessed by expression of activation markers (CD23, CD39), high levels of expression of adhesion molecules (LFA1, LFA3, ICAM1) and MHC class I and II alleles (reviewed in ref. 4 ). Owing to these characteristics, these cells are highly immunogenic (reviewed in ref. 5 ) and provide a useful tool for reactivation of EBV-specific cytotoxic T cells (CTLs) in vitro. EBV-transformed LCLs can be obtained by explantation of blood or lymphoid tissues from EBV seropositive individuals without need for exogenous infection (6 ). In addition, LCLs from EBV seropositive and seronegative donors can be obtained by in vitro infection of peripheral blood mononuclear cells (PBMCs) with EBV. The most commonly used strain for laboratory work is derived from the marmoset cell line B95.8 (see Chapter 1 and ref. 7 ). Production of supernatant from this virus producer cell line is described in Subheading 3.4.