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Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries

The identification of novel T cell antigens is central to basic and translational research in autoimmunity, tumor immunology, transplant immunology, and vaccine design for infectious disease. However, current methods for T cell antigen discovery are low throughput, and fail to explore a wide range o...

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Autores principales: Hondowicz, Brian D., Schwedhelm, Katharine V., Kas, Arnold, Tasch, Michael A., Rawlings, Crystal, Ramchurren, Nirasha, McIntosh, Martin, D'Amico, Leonard A., Sanda, Srinath, Standifer, Nathan E., Shendure, Jay, Stone, Brad
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3257230/
https://www.ncbi.nlm.nih.gov/pubmed/22253836
http://dx.doi.org/10.1371/journal.pone.0029949
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author Hondowicz, Brian D.
Schwedhelm, Katharine V.
Kas, Arnold
Tasch, Michael A.
Rawlings, Crystal
Ramchurren, Nirasha
McIntosh, Martin
D'Amico, Leonard A.
Sanda, Srinath
Standifer, Nathan E.
Shendure, Jay
Stone, Brad
author_facet Hondowicz, Brian D.
Schwedhelm, Katharine V.
Kas, Arnold
Tasch, Michael A.
Rawlings, Crystal
Ramchurren, Nirasha
McIntosh, Martin
D'Amico, Leonard A.
Sanda, Srinath
Standifer, Nathan E.
Shendure, Jay
Stone, Brad
author_sort Hondowicz, Brian D.
collection PubMed
description The identification of novel T cell antigens is central to basic and translational research in autoimmunity, tumor immunology, transplant immunology, and vaccine design for infectious disease. However, current methods for T cell antigen discovery are low throughput, and fail to explore a wide range of potential antigen-receptor interactions. To overcome these limitations, we developed a method in which programmable microarrays are used to cost-effectively synthesize complex libraries of thousands of minigenes that collectively encode the content of hundreds of candidate protein targets. Minigene-derived mRNA are transfected into autologous antigen presenting cells and used to challenge complex populations of purified peripheral blood CD8+ T cells in multiplex, parallel ELISPOT assays. In this proof-of-concept study, we apply synthetic minigene screening to identify two novel pancreatic islet autoantigens targeted in a patient with Type I Diabetes. To our knowledge, this is the first successful screen of a highly complex, synthetic minigene library for identification of a T cell antigen. In principle, responses against the full protein complement of any tissue or pathogen can be assayed by this approach, suggesting that further optimization of synthetic libraries holds promise for high throughput antigen discovery.
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spelling pubmed-32572302012-01-17 Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries Hondowicz, Brian D. Schwedhelm, Katharine V. Kas, Arnold Tasch, Michael A. Rawlings, Crystal Ramchurren, Nirasha McIntosh, Martin D'Amico, Leonard A. Sanda, Srinath Standifer, Nathan E. Shendure, Jay Stone, Brad PLoS One Research Article The identification of novel T cell antigens is central to basic and translational research in autoimmunity, tumor immunology, transplant immunology, and vaccine design for infectious disease. However, current methods for T cell antigen discovery are low throughput, and fail to explore a wide range of potential antigen-receptor interactions. To overcome these limitations, we developed a method in which programmable microarrays are used to cost-effectively synthesize complex libraries of thousands of minigenes that collectively encode the content of hundreds of candidate protein targets. Minigene-derived mRNA are transfected into autologous antigen presenting cells and used to challenge complex populations of purified peripheral blood CD8+ T cells in multiplex, parallel ELISPOT assays. In this proof-of-concept study, we apply synthetic minigene screening to identify two novel pancreatic islet autoantigens targeted in a patient with Type I Diabetes. To our knowledge, this is the first successful screen of a highly complex, synthetic minigene library for identification of a T cell antigen. In principle, responses against the full protein complement of any tissue or pathogen can be assayed by this approach, suggesting that further optimization of synthetic libraries holds promise for high throughput antigen discovery. Public Library of Science 2012-01-12 /pmc/articles/PMC3257230/ /pubmed/22253836 http://dx.doi.org/10.1371/journal.pone.0029949 Text en Hondowicz et al. http://creativecommons.org/licenses/by/4.0/ 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 properly credited.
spellingShingle Research Article
Hondowicz, Brian D.
Schwedhelm, Katharine V.
Kas, Arnold
Tasch, Michael A.
Rawlings, Crystal
Ramchurren, Nirasha
McIntosh, Martin
D'Amico, Leonard A.
Sanda, Srinath
Standifer, Nathan E.
Shendure, Jay
Stone, Brad
Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries
title Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries
title_full Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries
title_fullStr Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries
title_full_unstemmed Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries
title_short Discovery of T Cell Antigens by High-Throughput Screening of Synthetic Minigene Libraries
title_sort discovery of t cell antigens by high-throughput screening of synthetic minigene libraries
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3257230/
https://www.ncbi.nlm.nih.gov/pubmed/22253836
http://dx.doi.org/10.1371/journal.pone.0029949
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