Academic Journal
Extending the capabilities of intact-mass analyses to monoclonal immunoglobulins of the E-isotype (IgE)
| العنوان: | Extending the capabilities of intact-mass analyses to monoclonal immunoglobulins of the E-isotype (IgE) |
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| المؤلفون: | Wenhua Yang, Daniil G. Ivanov, Igor A. Kaltashov |
| المصدر: | mAbs, Vol 14, Iss 1 (2022) |
| بيانات النشر: | Taylor & Francis Group, 2022. |
| سنة النشر: | 2022 |
| المجموعة: | LCC:Therapeutics. Pharmacology LCC:Immunologic diseases. Allergy |
| مصطلحات موضوعية: | Immunoglobulin epsilon, mass spectrometry, limited charge reduction, non-covalent complexes, structural heterogeneity, antibody-antigen interactions, Therapeutics. Pharmacology, RM1-950, Immunologic diseases. Allergy, RC581-607 |
| الوصف: | Mass spectrometry (MS) has become an indispensable tool in structural characterization and quality control of monoclonal antibodies (mAbs). Intact-mass analysis is a particularly attractive option that provides a powerful and cost-effective means to not only confirm the structural integrity of the protein, but also probe its interactions with therapeutic targets. To a certain extent, this success can be attributed to relatively modest glycosylation levels exhibited by IgG molecules, which limits their structural heterogeneity and enables straightforward mass measurements at the intact molecule level. The recent surge of interest in expanding the repertoire of mAbs to include other classes of immunoglobulins places a premium on efforts to adapt the IgG-tailored experimental strategies to other classes of antibodies, but their dramatically higher levels of glycosylation may create insurmountable obstacles. The monoclonal murine IgE antibody explored in this work provides a challenging model system, as its glycosylation level exceeds that of conventional IgG mAbs by a factor of nine. The commercial sample, which included various IgE fragments, yields a poorly resolved ionic signal in intact-mass measurements, from which little useful information can be extracted. However, coupling MS measurements with the limited charge reduction of select polycationic species in the gas phase gives rise to well-defined charge ladders, from which both ionic masses and charges can be readily determined. The measurements reveal significant variation of the extent of glycosylation within intact IgE molecules, as well as the presence of low-molecular weight impurities in the commercial IgE sample. Furthermore, incubation of the monoclonal IgE with its antigen (ovalbumin) gives rise to the formation of complexes with varying stoichiometries, which can also be uniquely identified using a combination of native MS, limited charge reduction in the gas phase and data fitting procedures. This work demonstrates that following appropriate modifications, intact-mass analysis measurements can be successfully applied to mAbs beyond the IgG isotype, providing a wealth of information not only on the mass distribution of the intact IgE molecules, but also their large-scale conformational integrity, the integrity of their covalent structure, and their interactions with antigens. |
| نوع الوثيقة: | article |
| وصف الملف: | electronic resource |
| اللغة: | English |
| تدمد: | 19420862 1942-0870 1942-0862 |
| Relation: | https://doaj.org/toc/1942-0862; https://doaj.org/toc/1942-0870 |
| DOI: | 10.1080/19420862.2022.2103906 |
| URL الوصول: | https://doaj.org/article/fedaa0ed52df4655a7b457840cbf76cf |
| رقم الانضمام: | edsdoj.fedaa0ed52df4655a7b457840cbf76cf |
| قاعدة البيانات: | Directory of Open Access Journals |
Full Text Finder | تدمد: | 19420862 19420870 |
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| DOI: | 10.1080/19420862.2022.2103906 |