Generation and diversification of recombinant monoclonal antibodies
- Colorado State University, United States
- University of Geneva, Switzerland
- University of Colorado Anschutz Medical Campus, United States
- Northwestern University, United States
- University of Copenhagen, Denmark
Abstract
Antibodies are indispensable tools used for a large number of applications in both foundational and translational bioscience research; however, there are drawbacks to using traditional antibodies generated in animals. These include a lack of standardization leading to problems with reproducibility, high costs of antibodies purchased from commercial sources, and ethical concerns regarding the large number of animals used to generate antibodies. To address these issues, we have developed practical methodologies and tools for generating low-cost, high-yield preparations of recombinant monoclonal antibodies and antibody fragments directed to protein epitopes from primary sequences. We describe these methods here, as well as approaches to diversify monoclonal antibodies, including customization of antibody species specificity, generation of genetically encoded small antibody fragments, and conversion of single chain antibody fragments (e.g. scFv) into full-length, bivalent antibodies. This study focuses on antibodies directed to epitopes important for mitosis and kinetochore function; however, the methods and reagents described here are applicable to antibodies and antibody fragments for use in any field.
Data availability
All data generated during this study are included in the manuscript. We will also deposit the plasmid text files and maps on our institutional repository and AddGene.
Article and author information
Author details
Funding
National Institute of General Medical Sciences (R35GM130365)
- Jennifer G DeLuca
National Institute of General Medical Sciences (MIRA R35GM119728)
- Timothy J Stasevich
National Institute of General Medical Sciences (K99GM141453)
- Ning Zhao
National Institute of General Medical Sciences (R01GM135391)
- Dileep Varma
National Science Foundation (MCB-1845761)
- Timothy J Stasevich
National Cancer Institute (P30CA046934)
- Lori Sherman
- Steven M Anderson
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Silke Hauf, Virginia Tech, United States
Publication history
- Received: July 10, 2021
- Preprint posted: September 11, 2021 (view preprint)
- Accepted: December 20, 2021
- Accepted Manuscript published: December 31, 2021 (version 1)
- Accepted Manuscript updated: January 7, 2022 (version 2)
- Version of Record published: January 17, 2022 (version 3)
Copyright
© 2021, DeLuca et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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