Human plasma IgG1 repertoires are simple, unique, and dynamic

Bondt, Albert; Hoek, Max; Tamara, Sem; Graaf, Bastiaan de; Peng, Weiwei; Schulte, Douwe; Rijswijck, Danique M.H. van; Boer, Maurits A. den; Greisch, Jean‐François; Varkila, Meri R. J.; Snijder, Joost; Cremer, Olaf L.; Bonten, Marc J. M.; Heck, Albert J. R.

doi:10.1016/j.cels.2021.08.008

Cited by 49 publications

(116 citation statements)

References 56 publications

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“…Stringent analysis revealed that about 7-10% of these distinct ion signals matched to a Thr 109 ‘missed cleavage’, thus leaving still ~400 unique clones. Thus, as earlier observed for IgG1 in plasma ( 14 ), we found that in human milk a limited number of clones dominate the sIgA1 repertoire.…”

Section: Resultssupporting

confidence: 87%

“…This approach indeed provides antibody sequences, but these are likely not identical to the fully matured protein sequence of the Igs that are finally produced and secreted (31). We largely overcome these limitations by analyzing the Ig profiles of human biofluids directly, which we demonstrate here for sIgA1 from human milk and plasma, and have previously shown for IgG1 from human plasma (14).…”

Section: Discussionmentioning

confidence: 97%

“…The same LC-MS and data processing approaches as described by Bondt et al. were applied ( 14 ). In short, the collected Fab samples were separated on a Thermo Scientific Vanquish Flex UHPLC instrument, equipped with a 1 mm x 150 mm MAbPac Reversed Phase HPLC Column.…”

Section: Methodsmentioning

confidence: 99%

“…Recently, we introduced a novel LC-MS based approach to directly monitor the levels of IgG1 in plasma, whereby we could not only monitor the total plasma levels of IgG1 in donors, but were also able to distinguish and quantify the abundance of the 50-500 most abundant individual IgG1 clones present in plasma ( 14 ). Strikingly, we observed that the IgG1 repertoires were unique for each donor and remained quite stable over time in each healthy individual donor when monitored longitudinally.…”

Section: Introductionmentioning

confidence: 99%

“…By collecting these Fab molecules and analyzing them by intact LC-MS we were able to distinguish and quantify distinct IgG1 clones. We demonstrated that this method can be used to monitor the changes in abundance of individual clones in the plasma IgG1 repertoire over time, for instance caused by an infection ( 14 ).…”

Section: Introductionmentioning

confidence: 99%

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A Direct MS-Based Approach to Profile Human Milk Secretory Immunoglobulin A (IgA1) Reveals Donor-Specific Clonal Repertoires With High Longitudinal Stability

et al. 2021

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Recently, a mass spectrometry-based approach was introduced to directly assess the IgG1 immunoglobulin clonal repertoires in plasma. Here we expanded upon this approach by describing a mass spectrometry-based technique to assess specifically the clonal repertoire of another important class of immunoglobulin molecules, IgA1, and show it is efficiently and robustly applicable to either milk or plasma samples. Focusing on two individual healthy donors, whose milk was sampled longitudinally during the first 16 weeks of lactation, we demonstrate that the total repertoire of milk sIgA1 is dominated by only 50-500 clones, even though the human body theoretically can generate several orders of magnitude more clones. We show that in each donor the sIgA1 repertoire only changes marginally and quite gradually over the monitored 16-week period of lactation. Furthermore, the observed overlap in clonal repertoires between the two individual donors is close to non-existent. Mothers provide protection to their newborn infants directly by the transfer of antibodies via breastfeeding. The approach introduced here, can be used to visualize the clonal repertoire transferred from mother to infant and to detect changes in-time in that repertoire adapting to changes in maternal physiology.

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Section: Resultssupporting

confidence: 87%

Section: Discussionmentioning

confidence: 97%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

A Direct MS-Based Approach to Profile Human Milk Secretory Immunoglobulin A (IgA1) Reveals Donor-Specific Clonal Repertoires With High Longitudinal Stability

et al. 2021

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A multiparameter optimization in middle‐down analysis of monoclonal antibodies by LC–MS/MS

et al. 2023

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In antibody-based drug research, a complete characterization of antibody proteoforms covering both the amino acid sequence and all posttranslational modifications remains a major concern. The usual mass spectrometry-based approach to achieve this goal is bottom-up proteomics, which relies on the digestion of antibodies but does not allow the diversity of proteoforms to be assessed. Middle-down and topdown approaches have recently emerged as attractive alternatives but are not yet mastered and thus used in routine by many analytical chemistry laboratories. The work described here aims at providing guidelines to achieve the best sequence coverage for the fragmentation of intact light and heavy chains generated from a simple reduction of intact antibodies using Orbitrap mass spectrometry. Three parameters were found crucial to this aim: the use of an electron-based activation technique, the multiplex selection of precursor ions of different charge states, and the combination of replicates.

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Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top‐down and middle‐down mass spectrometry

et al. 2023

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Monoclonal antibodies (mAbs) have established themselves as the leading biopharmaceutical therapeutic modality. Once the developability of a mAb drug candidate has been assessed, an important step is to check its in vivo stability through pharmacokinetics (PK) studies. The gold standard is ligand‐binding assay (LBA) and liquid chromatography‐mass spectrometry (LC‐MS) performed at the peptide level (bottom‐up approach). However, these analytical techniques do not allow to address the different mAb proteoforms that can arise from biotransformation. In recent years, top‐down and middle‐down mass spectrometry approaches have gained popularity to characterize proteins at the proteoform level but are not yet widely used for PK studies. We propose here a workflow based on an automated immunocapture followed by top‐down and middle‐down liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) approaches to characterize mAb proteoforms spiked in mouse plasma. We demonstrate the applicability of our workflow on a large concentration range using pembrolizumab as a model. We also compare the performance of two state‐of‐the‐art Orbitrap platforms (Tribrid Eclipse and Exploris 480) for these studies. The added value of our workflow for an accurate and sensitive characterization of mAb proteoforms in mouse plasma is highlighted.

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Human plasma IgG1 repertoires are simple, unique, and dynamic

Cited by 49 publications

References 56 publications

A Direct MS-Based Approach to Profile Human Milk Secretory Immunoglobulin A (IgA1) Reveals Donor-Specific Clonal Repertoires With High Longitudinal Stability

A Direct MS-Based Approach to Profile Human Milk Secretory Immunoglobulin A (IgA1) Reveals Donor-Specific Clonal Repertoires With High Longitudinal Stability

A multiparameter optimization in middle‐down analysis of monoclonal antibodies by LC–MS/MS

Monitoring mAb proteoforms in mouse plasma using an automated immunocapture combined with top‐down and middle‐down mass spectrometry

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