Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras

Our aim was to identify conformational epitopes, recognized by monoclonal antibodies (mAbs) made against human (h) interferon (IFN)-γ. Based on the mAbs' (n = 12) ability to simultaneously bind hIFN-γ in ELISA, 2 epitope clusters with 5 mAbs in each were defined; 2 mAbs recognized unique epitop...

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Main Authors:	Zuber, Bartek, Rudström, Karin, Ehrnfelt, Cecilia, Ahlborg, Niklas
Format:	Online
Language:	English
Published:	Mary Ann Liebert, Inc. 2016
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5011633/

id	pubmed-5011633
recordtype	oai_dc
spelling	pubmed-50116332016-09-13 Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras Zuber, Bartek Rudström, Karin Ehrnfelt, Cecilia Ahlborg, Niklas Research Reports Our aim was to identify conformational epitopes, recognized by monoclonal antibodies (mAbs) made against human (h) interferon (IFN)-γ. Based on the mAbs' (n = 12) ability to simultaneously bind hIFN-γ in ELISA, 2 epitope clusters with 5 mAbs in each were defined; 2 mAbs recognized unique epitopes. Utilizing the mAbs' lack of reactivity with bovine (b) IFN-γ, epitopes were identified using 7 h/bIFN-γ chimeras where the helical regions (A-F) or the C terminus were substituted with bIFN-γ residues. Chimeras had a N-terminal peptide tag enabling the analysis of mAb recognition of chimeras in ELISA. The 2 mAb clusters mapped to region A and E, respectively; the epitopes of several mAbs also involved additional regions. MAbs in cluster A neutralized, to various degrees, IFN-γ-mediated activation of human cells, in line with the involvement of region A in the IFN-γ receptor interaction. MAbs mapping to region E displayed a stronger neutralizing capacity although this region has not been directly implicated in the receptor interaction. The results corroborate earlier studies and provide a detailed picture of the link between the epitope specificity and neutralizing capacity of mAbs. They further demonstrate the general use of peptide-tagged chimeric proteins as a powerful and straightforward method for efficient mapping of conformational epitopes. Mary Ann Liebert, Inc. 2016-09-01 2016-09-01 /pmc/articles/PMC5011633/ /pubmed/27336613 http://dx.doi.org/10.1089/jir.2016.0017 Text en © Bartek Zuber, et al., 2016; Published by Mary Ann Liebert, Inc. This Open Access article is distributed under the terms of the Creative Commons License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly 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	Zuber, Bartek Rudström, Karin Ehrnfelt, Cecilia Ahlborg, Niklas
spellingShingle	Zuber, Bartek Rudström, Karin Ehrnfelt, Cecilia Ahlborg, Niklas Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras
author_facet	Zuber, Bartek Rudström, Karin Ehrnfelt, Cecilia Ahlborg, Niklas
author_sort	Zuber, Bartek
title	Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras
title_short	Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras
title_full	Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras
title_fullStr	Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras
title_full_unstemmed	Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras
title_sort	epitope mapping of neutralizing monoclonal antibodies to human interferon-γ using human-bovine interferon-γ chimeras
description	Our aim was to identify conformational epitopes, recognized by monoclonal antibodies (mAbs) made against human (h) interferon (IFN)-γ. Based on the mAbs' (n = 12) ability to simultaneously bind hIFN-γ in ELISA, 2 epitope clusters with 5 mAbs in each were defined; 2 mAbs recognized unique epitopes. Utilizing the mAbs' lack of reactivity with bovine (b) IFN-γ, epitopes were identified using 7 h/bIFN-γ chimeras where the helical regions (A-F) or the C terminus were substituted with bIFN-γ residues. Chimeras had a N-terminal peptide tag enabling the analysis of mAb recognition of chimeras in ELISA. The 2 mAb clusters mapped to region A and E, respectively; the epitopes of several mAbs also involved additional regions. MAbs in cluster A neutralized, to various degrees, IFN-γ-mediated activation of human cells, in line with the involvement of region A in the IFN-γ receptor interaction. MAbs mapping to region E displayed a stronger neutralizing capacity although this region has not been directly implicated in the receptor interaction. The results corroborate earlier studies and provide a detailed picture of the link between the epitope specificity and neutralizing capacity of mAbs. They further demonstrate the general use of peptide-tagged chimeric proteins as a powerful and straightforward method for efficient mapping of conformational epitopes.
publisher	Mary Ann Liebert, Inc.
publishDate	2016
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5011633/
_version_	1613643570085888000

Epitope Mapping of Neutralizing Monoclonal Antibodies to Human Interferon-γ Using Human-Bovine Interferon-γ Chimeras

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