DOI: 10.1101/483537Nov 30, 2018Paper

A toolbox of IgG subclass-switched recombinant monoclonal antibodies for enhanced multiplex immunolabeling of brain

BioRxiv : the Preprint Server for Biology
Nicolas P AndrewsJames S Trimmer

Abstract

Generating recombinant monoclonal antibodies (R-mAbs) from mAb-producing hybridomas offers numerous advantages that increase the effectiveness, reproducibility, and transparent reporting of research. We report here the generation of a novel resource in the form of a library of recombinant R-mAbs validated for neuroscience research. We cloned immunoglobulin G (IgG) variable domains from cryopreserved hybridoma cells and input them into an integrated pipeline for expression and validation of functional R-mAbs. To improve efficiency over standard protocols, we eliminated aberrant Sp2/0-Ag14 hybridoma-derived variable light transcripts using restriction enzyme treatment. Further, we engineered a plasmid backbone that allows for switching of the IgG subclasses without altering target binding specificity to generate R-mAbs useful in simultaneous multiplex labeling experiments not previously possible. The method was also employed to rescue IgG variable sequences and generate functional R-mAbs from a non-viable cryopreserved hybridoma. All R-mAb sequences and plasmids will be archived and disseminated from open source suppliers.

Related Concepts

Monoclonal Antibodies
Brain
Guanosine
Hybridomas
Isotope Labeling
Plasmids
Research
Restriction Mapping
Immunolabeling Techniques
Disseminated

Related Feeds

BioRxiv & MedRxiv Preprints

BioRxiv and MedRxiv are the preprint servers for biology and health sciences respectively, operated by Cold Spring Harbor Laboratory. Here are the latest preprint articles (which are not peer-reviewed) from BioRxiv and MedRxiv.

Related Papers

The Western Journal of Medicine
R B Moss
Ceskoslovenská epidemiologie, mikrobiologie, imunologie
R Helm
Bulletin De La Société Des Sciences Médicales Du Grand-Duché De Luxembourg
F Hentges
© 2021 Meta ULC. All rights reserved