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Targeting membrane proteins for antibody discovery using phage display

A critical factor in the successful isolation of new antibodies by phage display is the presentation of a correctly folded antigen. While this is relatively simple for soluble proteins which can be purified and immobilized onto a plastic surface, membrane proteins offer significant challenges for an...

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Autores principales: Jones, Martina L., Alfaleh, Mohamed A., Kumble, Sumukh, Zhang, Shuo, Osborne, Geoffrey W., Yeh, Michael, Arora, Neetika, Hou, Jeff Jia Cheng, Howard, Christopher B., Chin, David Y., Mahler, Stephen M.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4870581/
https://www.ncbi.nlm.nih.gov/pubmed/27189586
http://dx.doi.org/10.1038/srep26240
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author Jones, Martina L.
Alfaleh, Mohamed A.
Kumble, Sumukh
Zhang, Shuo
Osborne, Geoffrey W.
Yeh, Michael
Arora, Neetika
Hou, Jeff Jia Cheng
Howard, Christopher B.
Chin, David Y.
Mahler, Stephen M.
author_facet Jones, Martina L.
Alfaleh, Mohamed A.
Kumble, Sumukh
Zhang, Shuo
Osborne, Geoffrey W.
Yeh, Michael
Arora, Neetika
Hou, Jeff Jia Cheng
Howard, Christopher B.
Chin, David Y.
Mahler, Stephen M.
author_sort Jones, Martina L.
collection PubMed
description A critical factor in the successful isolation of new antibodies by phage display is the presentation of a correctly folded antigen. While this is relatively simple for soluble proteins which can be purified and immobilized onto a plastic surface, membrane proteins offer significant challenges for antibody discovery. Whole cell panning allows presentation of the membrane protein in its native conformation, but is complicated by a low target antigen density, high background of irrelevant antigens and non-specific binding of phage particles to cell surfaces. The method described here uses transient transfection of alternating host cell lines and stringent washing steps to address each of these limitations. The successful isolation of antibodies from a naive scFv library is described for three membrane bound proteins; human CD83, canine CD117 and bat CD11b.
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spelling pubmed-48705812016-06-01 Targeting membrane proteins for antibody discovery using phage display Jones, Martina L. Alfaleh, Mohamed A. Kumble, Sumukh Zhang, Shuo Osborne, Geoffrey W. Yeh, Michael Arora, Neetika Hou, Jeff Jia Cheng Howard, Christopher B. Chin, David Y. Mahler, Stephen M. Sci Rep Article A critical factor in the successful isolation of new antibodies by phage display is the presentation of a correctly folded antigen. While this is relatively simple for soluble proteins which can be purified and immobilized onto a plastic surface, membrane proteins offer significant challenges for antibody discovery. Whole cell panning allows presentation of the membrane protein in its native conformation, but is complicated by a low target antigen density, high background of irrelevant antigens and non-specific binding of phage particles to cell surfaces. The method described here uses transient transfection of alternating host cell lines and stringent washing steps to address each of these limitations. The successful isolation of antibodies from a naive scFv library is described for three membrane bound proteins; human CD83, canine CD117 and bat CD11b. Nature Publishing Group 2016-05-18 /pmc/articles/PMC4870581/ /pubmed/27189586 http://dx.doi.org/10.1038/srep26240 Text en Copyright © 2016, Macmillan Publishers Limited http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in the credit line; if the material is not included under the Creative Commons license, users will need to obtain permission from the license holder to reproduce the material. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
spellingShingle Article
Jones, Martina L.
Alfaleh, Mohamed A.
Kumble, Sumukh
Zhang, Shuo
Osborne, Geoffrey W.
Yeh, Michael
Arora, Neetika
Hou, Jeff Jia Cheng
Howard, Christopher B.
Chin, David Y.
Mahler, Stephen M.
Targeting membrane proteins for antibody discovery using phage display
title Targeting membrane proteins for antibody discovery using phage display
title_full Targeting membrane proteins for antibody discovery using phage display
title_fullStr Targeting membrane proteins for antibody discovery using phage display
title_full_unstemmed Targeting membrane proteins for antibody discovery using phage display
title_short Targeting membrane proteins for antibody discovery using phage display
title_sort targeting membrane proteins for antibody discovery using phage display
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4870581/
https://www.ncbi.nlm.nih.gov/pubmed/27189586
http://dx.doi.org/10.1038/srep26240
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