Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy
Monoclonal antibodies (mAbs) are often needed and applied in high concentration solutions, >100 mg/mL. Due to close intermolecular distances between mAbs at high concentrations (~10−20 nm at 200 mg/mL), intermolecular interactions between mAbs and mAbs and solvent/co-solute molecule...
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MDPI AG
2019-07-01
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| Series: | Molecules |
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| Online Access: | https://www.mdpi.com/1420-3049/24/14/2528 |
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| author | Haleh H. Haeri Jacob Blaffert Florian A. Schöffmann Michaela Blech Josef Hartl Patrick Garidel Dariush Hinderberger |
| author_facet | Haleh H. Haeri Jacob Blaffert Florian A. Schöffmann Michaela Blech Josef Hartl Patrick Garidel Dariush Hinderberger |
| author_sort | Haleh H. Haeri |
| collection | DOAJ |
| description | Monoclonal antibodies (mAbs) are often needed and applied in high concentration solutions, >100 mg/mL. Due to close intermolecular distances between mAbs at high concentrations (~10−20 nm at 200 mg/mL), intermolecular interactions between mAbs and mAbs and solvent/co-solute molecules become non-negligible. Here, EPR spectroscopy is used to study the high-concentration solutions of mAbs and their effect on co-solvated small molecules, using EPR “spin probing” assay in aqueous and buffered solutions. Such, information regarding the surrounding environments of mAbs at high concentrations were obtained and comparisons between EPR-obtained micro-viscosities (rotational correlation times) and macroscopic viscosities measured by rheology were possible. In comparison with highly viscous systems like glycerol-water mixtures, it was found that up to concentrations of 50 mg/mL, the mAb-spin probe systems have similar trends in their macro- (rheology) and micro-viscosities (EPR), whereas at very high concentrations they deviate strongly. The charged spin probes sense an almost unchanged aqueous solution even at very high concentrations, which in turn indicates the existence of large solvent regions that despite their proximity to large mAbs essentially offer pure water reservoirs for co-solvated charged molecules. In contrast, in buffered solutions, amphiphilic spin probes like TEMPO interact with the mAb network, due to slight charge screening. The application of EPR spectroscopy in the present work has enabled us to observe and discriminate between electrostatic and hydrophobic kinds of interactions and depict the potential underlying mechanisms of network formation at high concentrations of mAbs. These findings could be of importance as well for the development of liquid-liquid phase separations often observed in highly concentrated protein solutions. |
| first_indexed | 2024-04-14T06:16:58Z |
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| language | English |
| last_indexed | 2024-04-14T06:16:58Z |
| publishDate | 2019-07-01 |
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| record_format | Article |
| series | Molecules |
| spelling | doaj.art-8d0f2cf081444ac0850acb431e7453c92022-12-22T02:08:10ZengMDPI AGMolecules1420-30492019-07-012414252810.3390/molecules24142528molecules24142528Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR SpectroscopyHaleh H. Haeri0Jacob Blaffert1Florian A. Schöffmann2Michaela Blech3Josef Hartl4Patrick Garidel5Dariush Hinderberger6Institute of Chemistry, Martin Luther University Halle-Wittenberg, D-06120 Halle (Saale), GermanyInstitute of Chemistry, Martin Luther University Halle-Wittenberg, D-06120 Halle (Saale), GermanyInstitute of Chemistry, Martin Luther University Halle-Wittenberg, D-06120 Halle (Saale), GermanyBoehringer Ingelheim Pharma GmbH & Co. KG, Protein Science, D-88397 Biberach an der Riss, GermanyInstitute of Chemistry, Martin Luther University Halle-Wittenberg, D-06120 Halle (Saale), GermanyInstitute of Chemistry, Martin Luther University Halle-Wittenberg, D-06120 Halle (Saale), GermanyInstitute of Chemistry, Martin Luther University Halle-Wittenberg, D-06120 Halle (Saale), GermanyMonoclonal antibodies (mAbs) are often needed and applied in high concentration solutions, >100 mg/mL. Due to close intermolecular distances between mAbs at high concentrations (~10−20 nm at 200 mg/mL), intermolecular interactions between mAbs and mAbs and solvent/co-solute molecules become non-negligible. Here, EPR spectroscopy is used to study the high-concentration solutions of mAbs and their effect on co-solvated small molecules, using EPR “spin probing” assay in aqueous and buffered solutions. Such, information regarding the surrounding environments of mAbs at high concentrations were obtained and comparisons between EPR-obtained micro-viscosities (rotational correlation times) and macroscopic viscosities measured by rheology were possible. In comparison with highly viscous systems like glycerol-water mixtures, it was found that up to concentrations of 50 mg/mL, the mAb-spin probe systems have similar trends in their macro- (rheology) and micro-viscosities (EPR), whereas at very high concentrations they deviate strongly. The charged spin probes sense an almost unchanged aqueous solution even at very high concentrations, which in turn indicates the existence of large solvent regions that despite their proximity to large mAbs essentially offer pure water reservoirs for co-solvated charged molecules. In contrast, in buffered solutions, amphiphilic spin probes like TEMPO interact with the mAb network, due to slight charge screening. The application of EPR spectroscopy in the present work has enabled us to observe and discriminate between electrostatic and hydrophobic kinds of interactions and depict the potential underlying mechanisms of network formation at high concentrations of mAbs. These findings could be of importance as well for the development of liquid-liquid phase separations often observed in highly concentrated protein solutions.https://www.mdpi.com/1420-3049/24/14/2528ESR/EPR spectroscopymonoclonal antibody (mAb)fluid interfacenitroxide spin probessolvationprotein network |
| spellingShingle | Haleh H. Haeri Jacob Blaffert Florian A. Schöffmann Michaela Blech Josef Hartl Patrick Garidel Dariush Hinderberger Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy Molecules ESR/EPR spectroscopy monoclonal antibody (mAb) fluid interface nitroxide spin probes solvation protein network |
| title | Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy |
| title_full | Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy |
| title_fullStr | Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy |
| title_full_unstemmed | Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy |
| title_short | Concentration Effects in the Interaction of Monoclonal Antibodies (mAbs) with their Immediate Environment Characterized by EPR Spectroscopy |
| title_sort | concentration effects in the interaction of monoclonal antibodies mabs with their immediate environment characterized by epr spectroscopy |
| topic | ESR/EPR spectroscopy monoclonal antibody (mAb) fluid interface nitroxide spin probes solvation protein network |
| url | https://www.mdpi.com/1420-3049/24/14/2528 |
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