Abstract
Out of all human antibodies, IgG4 stands out by its ability to dissociate in half-molecules and reassemble forming novel bivalent antibodies. Although this so-called Fab-arm exchange mechanism is acknowledged, detailed analysis of whether all IgG4 clones in serum are involved in such processes is not known. Here, by introducing a liquid chromatography-mass spectrometry-based approach enabling the analysis of serum IgG4 clonal repertoires, we show that widespread Fab-arm exchange occurs in serum of healthy donors and leads to a massive explosion in the molecular diversity of the IgG4 clonal repertoire. These findings provide new insight into IgG4, which plays a critical role in allergy, autoimmunity and vaccination settings, and may also impact the use of IgG4 as scaffold fo…
Abstract
Out of all human antibodies, IgG4 stands out by its ability to dissociate in half-molecules and reassemble forming novel bivalent antibodies. Although this so-called Fab-arm exchange mechanism is acknowledged, detailed analysis of whether all IgG4 clones in serum are involved in such processes is not known. Here, by introducing a liquid chromatography-mass spectrometry-based approach enabling the analysis of serum IgG4 clonal repertoires, we show that widespread Fab-arm exchange occurs in serum of healthy donors and leads to a massive explosion in the molecular diversity of the IgG4 clonal repertoire. These findings provide new insight into IgG4, which plays a critical role in allergy, autoimmunity and vaccination settings, and may also impact the use of IgG4 as scaffold for therapeutics.
Data availability
All data needed to evaluate the conclusions in the paper are present in the paper and/or the Supplementary Materials. The SEC-UV traces, raw spectra of the LC-MS data, and search algorithm output have been deposited to the MassIVE repository with identifier MSV000097341 [https://doi.org/10.25345/C5CC0V585]55. Source data underlying the figures are provided with this paper. Source data are provided with this paper.
Code availability
The data analysis code used to generate all Figures has been deposited to the MassIVE repository with identifier MSV000097341 [https://doi.org/10.25345/C5CC0V585]55.
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Acknowledgements
This research received funding by the Netherlands Organization for Scientific Research (NWO) through CHEMIE.PGT.2023.009 and the Spinoza Award SPI.2017.028 to AJRH. Additionally, continuous support from Genmab (Utrecht, NL) is appreciated, through the gift of recombinant mAbs (team lead Aran Labrijn) and financial support for our work. We thank Arjan Barendregt and Nadia Mokiem for their technical support during the method development and Gestur Vidarsson (Sanquin Research) for providing us with serum samples. We would further like to express our gratitude to Aran Labrijn for critically evaluating the manuscript, and to Oscar Dekker and Jessica van Bokkum (Leiden University Medical Center) for valuable discussions on how best to purify IgG4. M.G.H. receives financial support from the LUMC Gisela Thier 2021 fellowship, PPS Health Holland match call 2021, a Dutch incentive grant and the European Union through a ERC starter grant IgG4-START no.: 101163002.
Author information
Authors and Affiliations
Biomolecular Mass Spectrometry and Proteomics, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, CH, Utrecht, The Netherlands
Linus Wollenweber, Albert Bondt, Gestur Vidarsson & Albert J. R. Heck 1.
Netherlands Proteomics Center, CH, Utrecht, The Netherlands
Linus Wollenweber, Albert Bondt, Gestur Vidarsson & Albert J. R. Heck 1.
Immunoglobulin Research Laboratory, Sanquin Research, CX, Amsterdam, The Netherlands
Gestur Vidarsson 1.
Department of Human Genetics, Leiden University Medical Center, ZA, Leiden, The Netherlands
Maartje G. Huijbers 1.
Department of Neurology, Leiden University Medical Center, ZA, Leiden, The Netherlands
Maartje G. Huijbers
Authors
- Linus Wollenweber
- Albert Bondt
- Maartje G. Huijbers
- Albert J. R. Heck
Contributions
L.W., A.B. and A.J.R.H. conceived the project. G.V. advised on immunoglobulin biology. M.G.H. advised on IgG4 biology and purification of IgG4 from serum. A.B. and A.J.R.H. supervised the study. L.W. performed all experiments and analyzed the data. All drafted and edited the manuscript. A.J.R.H. provided funding for the project.
Corresponding author
Correspondence to Albert J. R. Heck.
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Competing interests
M.G.H. and G.V. are consultants for argenx. M.G.H. is a co-inventor on MuSK-related pending patents and LUMC and MGH receive royalties over these. LUMC receives royalties over a MuSK ELISA. All other authors declare no conflict of interest.
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Wollenweber, L., Bondt, A., Vidarsson, G. et al. Widespread Fab-arm exchange affects all endogenous serum IgG4. Nat Commun (2025). https://doi.org/10.1038/s41467-025-67105-9
Received: 06 April 2025
Accepted: 22 November 2025
Published: 08 December 2025
DOI: https://doi.org/10.1038/s41467-025-67105-9