Linear fidelity in quantification of anti-viral CD8+ T cells
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Flesch, Inge E. A.; Hollett, Natasha A.; Wong, Yik Chun; Tscharke, David C.
Description
Enumeration of anti-viral CD8+ T cells to make comparisons between mice, viruses and vaccines is a frequently used approach, but controversy persists as to the most appropriate methods. Use of peptide-MHC tetramers (or variants) and intracellular staining for cytokines, in particular IFNγ after a short ex vivo stimulation are now common, as are a variety of cytotoxicity assays, but few direct comparisons have been made. It has been argued that use of tetramers leads to the counting of...[Show more]
dc.contributor.author | Flesch, Inge E. A. | |
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dc.contributor.author | Hollett, Natasha A. | |
dc.contributor.author | Wong, Yik Chun | |
dc.contributor.author | Tscharke, David C. | |
dc.date.accessioned | 2012-08-14T00:22:36Z | |
dc.date.available | 2012-08-14T00:22:36Z | |
dc.identifier.issn | 1932-6203 | |
dc.identifier.uri | http://hdl.handle.net/1885/9176 | |
dc.description.abstract | Enumeration of anti-viral CD8+ T cells to make comparisons between mice, viruses and vaccines is a frequently used approach, but controversy persists as to the most appropriate methods. Use of peptide-MHC tetramers (or variants) and intracellular staining for cytokines, in particular IFNγ after a short ex vivo stimulation are now common, as are a variety of cytotoxicity assays, but few direct comparisons have been made. It has been argued that use of tetramers leads to the counting of non-functional T cells and that measurement of single cytokines will fail to identify cells with alternative functions. Further, the linear range of these methods has not been tested and this is required to give confidence that relative quantifications can be compared across samples. Here we show for two acute virus infections and CD8+ T cells activated in vitro that DimerX (a tetramer variant) and intracellular staining for IFNγ, alone or in combination with CD107 to detect degranulation, gave comparable results at the peak of the response. Importantly, these methods were highly linear over nearly two orders of magnitude. In contrast, in vitro and in vivo assays for cytotoxicity were not linear, suffering from high background killing, plateaus in maximal killing and substantial underestimation of differences in magnitude of responses. | |
dc.description.sponsorship | NHMRC (National Health and Medical Research Council of Australia) | |
dc.format | 9 pages | |
dc.publisher | Public Library of Science | |
dc.rights | "2012 Flesch et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited." - from article | |
dc.source | PLoS ONE 7.6 (2012): e39533 | |
dc.subject | virus | |
dc.subject | CD8 T cell | |
dc.subject | vaccinia virus | |
dc.subject | herpes simplex virus | |
dc.title | Linear fidelity in quantification of anti-viral CD8+ T cells | |
dc.type | Journal article | |
local.description.notes | This work was funded by Project grants (APP1005846; APP1023141) and a CDA fellowship (CDA 418108) from the Australian National Health and Medical Research Council (NHMRC). | |
local.identifier.citationvolume | 7 | |
dcterms.dateAccepted | 2012-05-23 | |
dc.date.issued | 2012-06-20 | |
local.identifier.absfor | 110700 - IMMUNOLOGY | |
local.identifier.ariespublication | f5625xPUB2275 | |
local.publisher.url | http://www.plos.org/ | |
local.type.status | Published Version | |
local.contributor.affiliation | Flesch, Inge E. A., ANU, Research School of Biology, Division of Biomedical Science and Biochemistry | |
local.contributor.affiliation | Hollett, Natasha A., ANU, Research School of Biology, Division of Biomedical Science and Biochemistry | |
local.contributor.affiliation | Wong, Yik Chun, ANU, Research School of Biology, Division of Biomedical Science and Biochemistry | |
local.contributor.affiliation | Tscharke, David C., ANU, Research School of Biology, Division of Biomedical Science and Biochemistry | |
dc.relation | http://purl.org/au-research/grants/nhmrc/1005846 | |
dc.relation | http://purl.org/au-research/grants/nhmrc/1023141 | |
dc.relation | http://purl.org/au-research/grants/nhmrc/418108 | |
local.bibliographicCitation.issue | 6 | |
local.identifier.doi | 10.1371/journal.pone.0039533 | |
dc.date.updated | 2015-12-10T11:56:42Z | |
local.identifier.scopusID | 2-s2.0-84862682075 | |
local.identifier.thomsonID | 000305693200087 | |
Collections | ANU Research Publications |
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