Selection of CMV-specific CD8+ and CD4+ T cells by mini-EBV-transformed B cell lines

Eur J Immunol. 2005 Jul;35(7):2110-21. doi: 10.1002/eji.200425936.


Efficient protocols to generate cytomegalovirus (CMV)-specific T cells are required for adoptive immunotherapy. Recombinant Epstein-Barr virus (EBV) vectors called mini-EBV can be used to establish permanent B cell lines in a single step, which present the CMV antigen pp65 in a constitutive manner. These B cell lines, coined pp65 mini-LCL, were successfully used to reactivate and expand CMV-specific cytotoxic T cells. Here we evaluate this pp65 mini-EBV system in closer detail, focusing on (1) the quantification of T cells with specific effector function and (2) the identification of CMV-specific CD4(+) helper T cells. The co-expansion of various functional CMV epitope specificities was demonstrated by IFN-gamma enzyme-linked immunospot assay (ELISPOT) assays and HLA-peptide tetramer staining. Single-cell cloning resulted in both CD4(+) and CD8(+) T cell clones, the majority of which was CMV specific. Thus, mini-LCL present the pp65 antigen on HLA class I and II, mobilizing both arms of the T cell response. Using a peptide library covering the pp65 sequence for further analysis of T cell clones, we identified new pp65 CD8(+) and CD4(+) T cell epitopes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Antigen Presentation / immunology
  • B-Lymphocytes / immunology*
  • CD4-Positive T-Lymphocytes / immunology*
  • CD4-Positive T-Lymphocytes / virology
  • CD8-Positive T-Lymphocytes / immunology*
  • CD8-Positive T-Lymphocytes / virology
  • Cell Differentiation / genetics
  • Cell Differentiation / immunology
  • Cell Line, Transformed
  • Clone Cells
  • Cytomegalovirus / genetics
  • Cytomegalovirus / immunology*
  • Epitopes, T-Lymphocyte / immunology*
  • HLA Antigens / immunology
  • Herpesvirus 4, Human / physiology*
  • Humans
  • Lymphocyte Activation / immunology*
  • Phosphoproteins / genetics
  • Phosphoproteins / immunology
  • Phosphoproteins / metabolism
  • Viral Matrix Proteins / genetics
  • Viral Matrix Proteins / immunology
  • Viral Matrix Proteins / metabolism


  • Epitopes, T-Lymphocyte
  • HLA Antigens
  • Phosphoproteins
  • Viral Matrix Proteins
  • cytomegalovirus matrix protein 65kDa