Cargando…
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...
Autores principales: | , , |
---|---|
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 |
_version_ | 1783304465027170304 |
---|---|
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. |
format | Online Article Text |
id | pubmed-5839796 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
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 |
work_keys_str_mv | AT pleinertino atoolboxofantimouseandantirabbitiggsecondarynanobodies AT batesmark atoolboxofantimouseandantirabbitiggsecondarynanobodies AT gorlichdirk atoolboxofantimouseandantirabbitiggsecondarynanobodies AT pleinertino toolboxofantimouseandantirabbitiggsecondarynanobodies AT batesmark toolboxofantimouseandantirabbitiggsecondarynanobodies AT gorlichdirk toolboxofantimouseandantirabbitiggsecondarynanobodies |