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2D TCR-pMHC-CD8 kinetics determines T-cell responses in a self-antigen-specific TCR system.

ABSTRACT: Two-dimensional (2D) kinetic analysis directly measures molecular interactions at cell-cell junctions, thereby incorporating inherent cellular effects. By comparison, three-dimensional (3D) analysis probes the intrinsic physical chemistry of interacting molecules isolated from the cell. To understand how T-cell tumor reactivity relates to 2D and 3D binding parameters and to directly compare them, we performed kinetic analyses of a panel of human T-cell receptors (TCRs) interacting with a melanoma self-antigen peptide (gp100209 -217 ) bound to peptide-major histocompatibility complex in the absence and presence of co-receptor CD8. We found that while 3D parameters are inadequate to predict T-cell function, 2D parameters (that do not correlate with their 3D counterparts) show a far broader dynamic range and significantly improved correlation with T-cell function. Thus, our data support the general notion that 2D parameters of TCR-peptide-major histocompatibility complex-CD8 interactions determine T-cell responsiveness and suggest a potential 2D-based strategy to screen TCRs for tumor immunotherapy.

SUBMITTER: Liu B

PROVIDER: S-EPMC3941036 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature

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2D TCR-pMHC-CD8 kinetics determines T-cell responses in a self-antigen-specific TCR system.

Liu Baoyu B Zhong Shi S Malecek Karolina K Johnson Laura A LA Rosenberg Steven A SA Zhu Cheng C Krogsgaard Michelle M

European journal of immunology 20131020 1

Two-dimensional (2D) kinetic analysis directly measures molecular interactions at cell-cell junctions, thereby incorporating inherent cellular effects. By comparison, three-dimensional (3D) analysis probes the intrinsic physical chemistry of interacting molecules isolated from the cell. To understand how T-cell tumor reactivity relates to 2D and 3D binding parameters and to directly compare them, we performed kinetic analyses of a panel of human T-cell receptors (TCRs) interacting with a melanom ...[more]

PMID: 24114747

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Project description:This experiment compares the transciptional changes in antigen specific murine CD8 T cells (P14 T cells) after exposure in vivo to dendritic cells (DC) pulsed with low dose cognate peptide (1uM KAVYNFATC), high dose cognate peptide (100uM KAVYNFATC) or no antigen. Splenic dendritic cells were freshly isolated, peptide pulsed, washed and then adoptively transferred s.c. to the right footpad of C57BL/6 hosts. After 18h, freshly isolated P14 CD8 T cells were labelled with CMFDA and adoptively transferred iv. Two hours after T cell transfer, anti-L selectin antibody was given iv. At 12 and 24 hours, recipients were sacrificed and The right popliteal LN was harvested at 12 or 24h post T cell transfer and a single cell suspension was created and stained with PE CD4, B220 and CD19 (dump channel). Cells were then sorted on a FacsARIA for being non-doublets, CMFDA positive and dump channel negative. This experiment compares the transciptional changes in antigen specific murine CD8 T cells (P14 T cells) after exposure in vivo to dendritic cells (DC) pulsed with low dose cognate peptide (1uM KAVYNFATC), high dose cognate peptide (100uM KAVYNFATC) or no antigen. Splenic dendritic cells were freshly isolated, peptide pulsed, washed and then adoptively transferred s.c. to the right footpad of C57BL/6 hosts. After 18h, freshly isolated P14 CD8 T cells were labelled with CMFDA and adoptively transferred iv. Two hours after T cell transfer, anti-L selectin antibody was given iv. At 12 and 24 hours, recipients were sacrificed and The right popliteal LN was harvested at 12 or 24h post T cell transfer and a single cell suspension was created and stained with PE CD4, B220 and CD19 (dump channel). Cells were then sorted on a FacsARIA for being non-doublets, CMFDA positive and dump channel negative. The experiment was conducted for 39 samples out of which 35 passsed transcriptional quality control tests. The phenotypic distribution for the 35 samples includes: (1) high dose (100uM KAVYNFATC ) cognate peptide pulsed samples harvested at 12h post T cell transfer: 6 biological replicates (2) high dose (100uM KAVYNFATC ) cognate peptide pulsed samples harvested at 24h post T cell transfer: 7 biological replicates (3) low dose (1uM KAVYNFATC) cognate peptide pulsed samples harvested at 12h post T cell transfer: 6 biological replicates (4) low dose (1uM KAVYNFATC) cognate peptide pulsed samples harvested at 24h post T cell transfer: 9 biological replicates (5) no antigen pulsed samples harvested at 12h post T cell transfer: 3 biological replicates (6) no antigen pulsed samples harvested at 24h post T cell transfer: 4 biological replicates.

Dataset Information

2D TCR-pMHC-CD8 kinetics determines T-cell responses in a self-antigen-specific TCR system.

Publications

2D TCR-pMHC-CD8 kinetics determines T-cell responses in a self-antigen-specific TCR system.

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