PepTivator® EBV EBNA-1 ( #130-093-613 #130-093-614 )

PepTivator® EBV EBNA-1

Description

The PepTivator® EBV EBNA-1 is a peptide pool that consists mainly of 15-mer peptides with 11 amino acids overlap, covering the complete sequence of the Epstein-Barr virus EBNA-1 protein.
PepTivator peptide pools have been developed for the efficient in vitro stimulation of antigen–specific CD4⁺ and CD8⁺ T cells, as peptides of 15-aa length with 11 aa overlap represent an optimized solution for stimulating both CD4⁺and CD8⁺ T cells in various applications.¹
Quantitative, phenotypical, or functional analysis of EBV EBNA-1–specific T cell immunity can provide important information on the natural course of immune responses in healthy or immunocompromised individuals.

Details

Background information
Epstein-Barr virus (EBV) is a human γ-herpes virus with B cell growth–transforming ability and lymphomagenic potential. More than 90% of human adults are infected with EBV. Usually the primary infection is asymptomatic, but some individuals develop an infectious mononucleosis, a self-limiting lymphoproliferative disease. The Epstein-Barr nuclear antigen 1 (EBNA-1) protein is involved in the replication of viral episomes and therefore crucial for the persistence of the infection. Its expression is maintained in all latency cycles of infection and in all EBV-associated malignancies. EBNA-1-specific CD4⁺ and CD8⁺ T cells have been found in EBV-infected individuals.^2–5

Downstream applications
The in vitro stimulation of EBV EBNA-1–specific CD4⁺ and CD8⁺ T cells with PepTivator EBV EBNA-1 causes the secretion of effector cytokines and the upregulation of activation markers, which then allow the detection and isolation of EBV EBNA-1–specific T cells ⁶:

Detection and analysis of EBV EBNA-1–specific CD4⁺ and CD8⁺ effector/memory T cells in PBMCs by MACS® Cytokine Secretion Assays, intracellular cytokine staining, or other technologies.
Isolation of viable EBV EBNA-1–specific CD4⁺ T cells with the CD154 MicroBead Kit, or of CD4⁺ and CD8⁺ T cells using the CD137 MicroBead Kit or MACS Cytokine Secretion Assay – Cell Enrichment and Detection Kits. Subsequently, cells can be expanded for generation of T cell lines.
Generation of EBV EBNA-1–specific CD4⁺ and CD8⁺ effector/memory T cells from naive T cell populations for research on immunotherapy and vaccination
Pulsing of antigen-presenting cells, for example, for research on dendritic cell vaccination.

Figure 1

10⁷ PBMCs of an EBV⁺ donor were restimulated for 4 hours with 20 μL/mL of reconstituted PepTivator EBV EBNA-1. EBNA-1-specific cells were stained and magnetically enriched according to their secretion of IFN-γ using the IFN-γ Secretion Assay - Cell Enrichment and Detection Kit. T cells were counterstained for CD4 and CD8 expression. Cell debris and dead cells were excluded from the analysis based on scatter signals and PI fluorescence. IFN-γ secretion of viable lymphocytes is shown.

CD4⁺ T cells

Before enrichment

After enrichment

CD8⁺ T cells

Before enrichment

After enrichment

Figure 2

10⁶ PBMCs of an EBV⁺ donor were restimulated for 6 hours with 20 μL/mL of reconstituted PepTivator EBV EBNA-1 or without antigen. After 2 hours 1µg/mL brefeldin A was added. Cells were fixed, permeabilized and EBNA-1-specific cells were intracellularly stained with Anti-IFN-γ-PE. T cells were counterstained for CD4 expression. IFN-γ production of lymphocytes is shown.

Stimulated cells

Unstimulated control