When monoclonal antibodies are not monospecific: Hybridomas frequently express additional functional variable regions

MAbs
Andrew R M BradburyS Dübel

Abstract

Monoclonal antibodies are commonly assumed to be monospecific, but anecdotal studies have reported genetic diversity in antibody heavy chain and light chain genes found within individual hybridomas. As the prevalence of such diversity has never been explored, we analyzed 185 random hybridomas, in a large multicenter dataset. The hybridomas analyzed were not biased towards those with cloning difficulties or known to have additional chains. Of the hybridomas we evaluated, 126 (68.1%) contained no additional productive chains, while the remaining 59 (31.9%) contained one or more additional productive heavy or light chains. The expression of additional chains degraded properties of the antibodies, including specificity, binding signal and/or signal-to-noise ratio, as determined by enzyme-linked immunosorbent assay and immunohistochemistry. The most abundant mRNA transcripts found in a hybridoma cell line did not necessarily encode the antibody chains providing the correct specificity. Consequently, when cloning antibody genes, functional validation of all possible VH and VL combinations is required to identify those with the highest affinity and lowest cross-reactivity. These findings, reflecting the current state of hybridomas use...Continue Reading

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Citations

Mar 26, 2019·Endocrinology·Luis Leon MercadoLaurent Gautron
Jun 4, 2019·Analytical and Bioanalytical Chemistry·Andreas PoschenriederPeter B Luppa
Aug 5, 2020·MAbs·Jan L A VoskuilSimon L Goodman
May 14, 2019·Frontiers in Neuroanatomy·Laurent Gautron
Aug 25, 2020·Biospektrum : Zeitschrift Der Gesellschaft Für Biologishe Chemie (GBCH) Und Der Vereinigung Für Allgemeine Und Angewandte Mikrobiologie (VAAM)·Esther Veronika WenzelStefan Dübel
Jun 20, 2019·Toxins·Leslie NaranjoAndrew R M Bradbury
Oct 14, 2020·Nature Biotechnology·Alison GrayStefan Dübel
Nov 6, 2020·Current Protocols in Neuroscience·James S Trimmer
Nov 13, 2020·EMBO Reports·René Custers, Jan Steyaert
Jan 13, 2021·Molecular Imaging and Biology : MIB : the Official Publication of the Academy of Molecular Imaging·Mitra Sadat TabatabaeeFrederic Menard
Feb 16, 2021·Protein Engineering, Design & Selection : PEDS·Nileena VelappanAndrew R M Bradbury
Feb 19, 2021·Journal for Immunotherapy of Cancer·Daniele CaraccioloPierfrancesco Tassone
Aug 8, 2018·New Biotechnology·Michael J TaussigJames S Trimmer
Mar 25, 2021·Biological Chemistry·Bernhard ValldorfStefan Zielonka
Mar 30, 2021·Trends in Biotechnology·Andreas H LaustsenTimothy P Jenkins
Jul 27, 2021·Frontiers in Cellular and Infection Microbiology·Kristian Daniel Ralph RothMichael Hust
Aug 22, 2021·Analytical and Bioanalytical Chemistry·Riikka PeltomaaMaría C Moreno-Bondi
Jan 1, 2022·ELife·Keith F DeLucaJennifer G DeLuca

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Methods Mentioned

BETA
enzyme-linked immunosorbent assay
ELISA
PCR
phage display
transfections

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