Screening of HCMV- or EBV-specific CD8 T cell lines for cross-recognition of allogeneic MHC molecules.

<p>CD8 T cell lines, sorted with recombinant pMHC multimeric complexes specific to HCMV (pp65_495–503/A*0201) or EBV lytic epitopes (BMLF1_259–267/A*0201 or BZLF1_54–64/B*3501), were tested: (A) for cytotoxicity toward a panel of 30 HLA-typed LCL, in a 4-h chromium release assay (Effector-Target ratio 15∶1). HLA-specific killing of LCL was observed for 3 of the 11 pp65/A2-specific T cell lines tested: T cell line from D01 killed A*3101⁺ LCL, T cell line from D03 killed A*3201⁺ LCL, and T cell line from D08 killed A*3001⁺ LCL. The BZLF1/B*3501-sorted T cell line from D15 killed LCL sharing Cw*0602 expression. Only T cell lines that showed alloresponse are displayed. LCL triggering an alloresponse are shown as well as some of the tested LCL which do not elicit a cytotoxic response. Cognate peptide-HLA complexes (pp65_495–503/A*0201 or BZLF1_54–64/B*3501) were used as positive control. Data are presented as the mean percentage lysis and are representative of 3 different experiments. (<b>B</b>) for TNF-α production toward COS-7 cells transfected with plasmids encoding class I HLA alleles. T cells were added 2 days after the transfection, and the TNF-α content of the supernatant, expressed in pg/mL, was estimated 6 h later by testing the toxicity of the supernatants for TNF-α sensitive WEHI-164 clone 13 cells. TNF-α production was observed for the pp65/A*0201-sorted T cell line from D03, that recognized selectively COS cells expressing HLA-A*3201, and for the BZLF1/B*3501 T cell line from D15 that recognized selectively COS cells expressing HLA-Cw*0602. Plasmids encoding class I HLA alleles that elicits a response are shown as well as some of the plasmid encoding class I HLA alleles that do not induce TNF-α secretion. T cell lines that did not produce TNF-α are not represented. Cognate peptide-HLA complexes (pp65_495–503/A*0201 or BZLF1_54–64/B*3501) were used as positive control. One out three independent experiments is shown.