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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...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2012
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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. |
format | Online Article Text |
id | pubmed-3257230 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
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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