Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining

Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining

Ex vivo ELISPOT and multimer staining are well-established tests for the assessment of antigen-specific T cells. Many laboratories are now using a period of in vitro stimulation (IVS) to enhance detection. Here, we report the findings of a multi-centre panel organised by the Association for Cancer I...

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Main Authors: Chudley, Lindsey, McCann, Katy J., Coleman, Adam, Cazaly, Angelica M., Bidmon, Nicole, Britten, Cedrik M., van der Burg, Sjoerd H., Gouttefangeas, Cecile, Jandus, Camilla, Laske, Karoline, Maurer, Dominik, Romero, Pedro, Schröder, Helene, Stynenbosch, Linda F. M., Walter, Steffen, Welters, Marij J. P., Ottensmeier, Christian H.
Format: Online
Language:English
Published: Springer Berlin Heidelberg 2014
Online Access:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4209099/
id pubmed-4209099
recordtype oai_dc
spelling pubmed-42090992014-10-28 Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining Chudley, Lindsey McCann, Katy J. Coleman, Adam Cazaly, Angelica M. Bidmon, Nicole Britten, Cedrik M. van der Burg, Sjoerd H. Gouttefangeas, Cecile Jandus, Camilla Laske, Karoline Maurer, Dominik Romero, Pedro Schröder, Helene Stynenbosch, Linda F. M. Walter, Steffen Welters, Marij J. P. Ottensmeier, Christian H. Original Article Ex vivo ELISPOT and multimer staining are well-established tests for the assessment of antigen-specific T cells. Many laboratories are now using a period of in vitro stimulation (IVS) to enhance detection. Here, we report the findings of a multi-centre panel organised by the Association for Cancer Immunotherapy Immunoguiding Program to investigate the impact of IVS protocols on the detection of antigen-specific T cells of varying ex vivo frequency. Five centres performed ELISPOT and multimer staining on centrally prepared PBMCs from 3 donors, both ex vivo and following IVS. A harmonised IVS protocol was designed based on the best-performing protocol(s), which was then evaluated in a second phase on 2 donors by 6 centres. All centres were able to reliably detect antigen-specific T cells of high/intermediate frequency both ex vivo (Phase I) and post-IVS (Phase I and II). The highest frequencies of antigen-specific T cells ex vivo were mirrored in the frequencies following IVS and in the detection rates. However, antigen-specific T cells of a low/undetectable frequency ex vivo were not reproducibly detected post-IVS. Harmonisation of the IVS protocol reduced the inter-laboratory variation observed for ELISPOT and multimer analyses by approximately 20 %. We further demonstrate that results from ELISPOT and multimer staining correlated after (P < 0.0001 and R2 = 0.5113), but not before IVS. In summary, IVS was shown to be a reproducible method that benefitted from method harmonisation. Springer Berlin Heidelberg 2014-08-19 2014 /pmc/articles/PMC4209099/ /pubmed/25134947 http://dx.doi.org/10.1007/s00262-014-1593-0 Text en © The Author(s) 2014 Open AccessThis article is distributed under the terms of the Creative Commons Attribution License which permits any use, distribution, and reproduction in any medium, provided the original author(s) and the source are credited.
repository_type Open Access Journal
institution_category Foreign Institution
institution US National Center for Biotechnology Information
building NCBI PubMed
collection Online Access
language English
format Online
author Chudley, Lindsey
McCann, Katy J.
Coleman, Adam
Cazaly, Angelica M.
Bidmon, Nicole
Britten, Cedrik M.
van der Burg, Sjoerd H.
Gouttefangeas, Cecile
Jandus, Camilla
Laske, Karoline
Maurer, Dominik
Romero, Pedro
Schröder, Helene
Stynenbosch, Linda F. M.
Walter, Steffen
Welters, Marij J. P.
Ottensmeier, Christian H.
spellingShingle Chudley, Lindsey
McCann, Katy J.
Coleman, Adam
Cazaly, Angelica M.
Bidmon, Nicole
Britten, Cedrik M.
van der Burg, Sjoerd H.
Gouttefangeas, Cecile
Jandus, Camilla
Laske, Karoline
Maurer, Dominik
Romero, Pedro
Schröder, Helene
Stynenbosch, Linda F. M.
Walter, Steffen
Welters, Marij J. P.
Ottensmeier, Christian H.
Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining
author_facet Chudley, Lindsey
McCann, Katy J.
Coleman, Adam
Cazaly, Angelica M.
Bidmon, Nicole
Britten, Cedrik M.
van der Burg, Sjoerd H.
Gouttefangeas, Cecile
Jandus, Camilla
Laske, Karoline
Maurer, Dominik
Romero, Pedro
Schröder, Helene
Stynenbosch, Linda F. M.
Walter, Steffen
Welters, Marij J. P.
Ottensmeier, Christian H.
author_sort Chudley, Lindsey
title Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining
title_short Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining
title_full Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining
title_fullStr Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining
title_full_unstemmed Harmonisation of short-term in vitro culture for the expansion of antigen-specific CD8+ T cells with detection by ELISPOT and HLA-multimer staining
title_sort harmonisation of short-term in vitro culture for the expansion of antigen-specific cd8+ t cells with detection by elispot and hla-multimer staining
description Ex vivo ELISPOT and multimer staining are well-established tests for the assessment of antigen-specific T cells. Many laboratories are now using a period of in vitro stimulation (IVS) to enhance detection. Here, we report the findings of a multi-centre panel organised by the Association for Cancer Immunotherapy Immunoguiding Program to investigate the impact of IVS protocols on the detection of antigen-specific T cells of varying ex vivo frequency. Five centres performed ELISPOT and multimer staining on centrally prepared PBMCs from 3 donors, both ex vivo and following IVS. A harmonised IVS protocol was designed based on the best-performing protocol(s), which was then evaluated in a second phase on 2 donors by 6 centres. All centres were able to reliably detect antigen-specific T cells of high/intermediate frequency both ex vivo (Phase I) and post-IVS (Phase I and II). The highest frequencies of antigen-specific T cells ex vivo were mirrored in the frequencies following IVS and in the detection rates. However, antigen-specific T cells of a low/undetectable frequency ex vivo were not reproducibly detected post-IVS. Harmonisation of the IVS protocol reduced the inter-laboratory variation observed for ELISPOT and multimer analyses by approximately 20 %. We further demonstrate that results from ELISPOT and multimer staining correlated after (P < 0.0001 and R2 = 0.5113), but not before IVS. In summary, IVS was shown to be a reproducible method that benefitted from method harmonisation.
publisher Springer Berlin Heidelberg
publishDate 2014
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4209099/
_version_ 1613148701299048448

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