Identification of EBNA2-, EBNA-LP- and BHRF1-specific CD8+ T cell responses.

<p>(A) EBNA2-specific responses. Top left panel: In vitro expanded, CD8-enriched polyclonal T cells from Donor 17 were screened against overlapping 20mer peptides spanning the complete unique sequence of EBNA2 (sub-divided into 18 pools). T cell recognition was assessed by IFNγ production, measured in an ELISA. Results are expressed as the mean IFNγ concentration +/- SD for duplicate wells. Top right panel: Individual component peptides from pool 6 were screened for their ability to mediate IFNγ production by the CD8-enriched T cell population. Middle panel: To identify the HLA restriction, LCLs sharing one or more class I alleles with Donor 17 (class I type in bold) were pre-loaded with peptides 6.4/6.5 (1μg/ml) and co-cultured overnight with a specific T cell clone. Results are expressed as the mean IFNγ concentration +/- SD for triplicate wells. Table: Peptides 6.4, 6.5 and the predicted minimal epitope were screened for their ability to induce IFNγ production by the CD8-enriched polyclonal T cell population. (B) EBNA-LP-specific responses. Left panel: In vitro expanded CD8-enriched polyclonal T cells from Donor 20 were screened against overlapping 15mer peptides spanning the complete unique sequence of EBNA-LP (sub-divided into 4 pools); T cell recognition was assessed by IFNγ production. Right panel: Individual component peptides from pool 3 were screened for their ability to induce IFNγ production by the CD8-enriched T cell population. Sequences and T cell recognition data for peptide 3.6 and the minimal epitope are shown in the table. (C) BHRF1-specific responses. Top left panel: In vitro expanded CD8-enriched polyclonal T cells from Donor 8 were screened against overlapping 15mer peptides spanning the complete unique sequence of BHRF1 (sub-divided into 6 pools). Top right panel: Individual component peptides from pool 1 were screened for their ability to mediate IFNγ production by the total polyclonal T cell population. Middle panel: To identify the HLA restriction, LCLs sharing one or more class I alleles with Donor 8 (class I type in bold) were pre-loaded with peptides 1.3/1.4 (1μg/ml) and co-cultured overnight with a specific T cell clone. Results are expressed as the mean IFNγ concentration +/- SD for triplicate wells. Table: Peptides 1.3/1.4 and the predicted minimal epitope were screened for their ability to mediate IFNγ production by the CD8-enriched T cell population.