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A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies

Polyclonal anti–immunoglobulin G (anti-IgG) secondary antibodies are essential tools for many molecular biology techniques and diagnostic tests. Their animal-based production is, however, a major ethical problem. Here, we introduce a sustainable alternative, namely nanobodies against all mouse IgG s...

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Detalles Bibliográficos
Autores principales: Pleiner, Tino, Bates, Mark, Görlich, Dirk
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Rockefeller University Press 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839796/
https://www.ncbi.nlm.nih.gov/pubmed/29263082
http://dx.doi.org/10.1083/jcb.201709115
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author Pleiner, Tino
Bates, Mark
Görlich, Dirk
author_facet Pleiner, Tino
Bates, Mark
Görlich, Dirk
author_sort Pleiner, Tino
collection PubMed
description Polyclonal anti–immunoglobulin G (anti-IgG) secondary antibodies are essential tools for many molecular biology techniques and diagnostic tests. Their animal-based production is, however, a major ethical problem. Here, we introduce a sustainable alternative, namely nanobodies against all mouse IgG subclasses and rabbit IgG. They can be produced at large scale in Escherichia coli and could thus make secondary antibody production in animals obsolete. Their recombinant nature allows fusion with affinity tags or reporter enzymes as well as efficient maleimide chemistry for fluorophore coupling. We demonstrate their superior performance in Western blotting, in both peroxidase- and fluorophore-linked form. Their site-specific labeling with multiple fluorophores creates bright imaging reagents for confocal and superresolution microscopy with much smaller label displacement than traditional secondary antibodies. They also enable simpler and faster immunostaining protocols, and allow multitarget localization with primary IgGs from the same species and of the same class.
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spelling pubmed-58397962018-03-08 A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies Pleiner, Tino Bates, Mark Görlich, Dirk J Cell Biol Research Articles Polyclonal anti–immunoglobulin G (anti-IgG) secondary antibodies are essential tools for many molecular biology techniques and diagnostic tests. Their animal-based production is, however, a major ethical problem. Here, we introduce a sustainable alternative, namely nanobodies against all mouse IgG subclasses and rabbit IgG. They can be produced at large scale in Escherichia coli and could thus make secondary antibody production in animals obsolete. Their recombinant nature allows fusion with affinity tags or reporter enzymes as well as efficient maleimide chemistry for fluorophore coupling. We demonstrate their superior performance in Western blotting, in both peroxidase- and fluorophore-linked form. Their site-specific labeling with multiple fluorophores creates bright imaging reagents for confocal and superresolution microscopy with much smaller label displacement than traditional secondary antibodies. They also enable simpler and faster immunostaining protocols, and allow multitarget localization with primary IgGs from the same species and of the same class. The Rockefeller University Press 2018-03-05 /pmc/articles/PMC5839796/ /pubmed/29263082 http://dx.doi.org/10.1083/jcb.201709115 Text en © 2018 Pleiner et al. https://creativecommons.org/licenses/by/4.0/This article is available under a Creative Commons License (Attribution 4.0 International, as described at https://creativecommons.org/licenses/by/4.0/).
spellingShingle Research Articles
Pleiner, Tino
Bates, Mark
Görlich, Dirk
A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
title A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
title_full A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
title_fullStr A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
title_full_unstemmed A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
title_short A toolbox of anti–mouse and anti–rabbit IgG secondary nanobodies
title_sort toolbox of anti–mouse and anti–rabbit igg secondary nanobodies
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5839796/
https://www.ncbi.nlm.nih.gov/pubmed/29263082
http://dx.doi.org/10.1083/jcb.201709115
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