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MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour
During the SARS-CoV-2 pandemic, many virus-binding monoclonal antibodies have been developed for clinical and diagnostic purposes. This underlines the importance of antibodies as universal bioanalytical reagents. However, little attention is given to the reproducibility crisis that scientific studie...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9036215/ https://www.ncbi.nlm.nih.gov/pubmed/35466280 http://dx.doi.org/10.3390/antib11020027 |
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author | Tscheuschner, Georg Kaiser, Melanie N. Lisec, Jan Beslic, Denis Muth, Thilo Krüger, Maren Mages, Hans Werner Dorner, Brigitte G. Knospe, Julia Schenk, Jörg A. Sellrie, Frank Weller, Michael G. |
author_facet | Tscheuschner, Georg Kaiser, Melanie N. Lisec, Jan Beslic, Denis Muth, Thilo Krüger, Maren Mages, Hans Werner Dorner, Brigitte G. Knospe, Julia Schenk, Jörg A. Sellrie, Frank Weller, Michael G. |
author_sort | Tscheuschner, Georg |
collection | PubMed |
description | During the SARS-CoV-2 pandemic, many virus-binding monoclonal antibodies have been developed for clinical and diagnostic purposes. This underlines the importance of antibodies as universal bioanalytical reagents. However, little attention is given to the reproducibility crisis that scientific studies are still facing to date. In a recent study, not even half of all research antibodies mentioned in publications could be identified at all. This should spark more efforts in the search for practical solutions for the traceability of antibodies. For this purpose, we used 35 monoclonal antibodies against SARS-CoV-2 to demonstrate how sequence-independent antibody identification can be achieved by simple means applied to the protein. First, we examined the intact and light chain masses of the antibodies relative to the reference material NIST-mAb 8671. Already half of the antibodies could be identified based solely on these two parameters. In addition, we developed two complementary peptide mass fingerprinting methods with MALDI-TOF-MS that can be performed in 60 min and had a combined sequence coverage of over 80%. One method is based on the partial acidic hydrolysis of the protein by 5 mM of sulfuric acid at 99 °C. Furthermore, we established a fast way for a tryptic digest without an alkylation step. We were able to show that the distinction of clones is possible simply by a brief visual comparison of the mass spectra. In this work, two clones originating from the same immunization gave the same fingerprints. Later, a hybridoma sequencing confirmed the sequence identity of these sister clones. In order to automate the spectral comparison for larger libraries of antibodies, we developed the online software ABID 2.0. This open-source software determines the number of matching peptides in the fingerprint spectra. We propose that publications and other documents critically relying on monoclonal antibodies with unknown amino acid sequences should include at least one antibody fingerprint. By fingerprinting an antibody in question, its identity can be confirmed by comparison with a library spectrum at any time and context. |
format | Online Article Text |
id | pubmed-9036215 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-90362152022-04-26 MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour Tscheuschner, Georg Kaiser, Melanie N. Lisec, Jan Beslic, Denis Muth, Thilo Krüger, Maren Mages, Hans Werner Dorner, Brigitte G. Knospe, Julia Schenk, Jörg A. Sellrie, Frank Weller, Michael G. Antibodies (Basel) Article During the SARS-CoV-2 pandemic, many virus-binding monoclonal antibodies have been developed for clinical and diagnostic purposes. This underlines the importance of antibodies as universal bioanalytical reagents. However, little attention is given to the reproducibility crisis that scientific studies are still facing to date. In a recent study, not even half of all research antibodies mentioned in publications could be identified at all. This should spark more efforts in the search for practical solutions for the traceability of antibodies. For this purpose, we used 35 monoclonal antibodies against SARS-CoV-2 to demonstrate how sequence-independent antibody identification can be achieved by simple means applied to the protein. First, we examined the intact and light chain masses of the antibodies relative to the reference material NIST-mAb 8671. Already half of the antibodies could be identified based solely on these two parameters. In addition, we developed two complementary peptide mass fingerprinting methods with MALDI-TOF-MS that can be performed in 60 min and had a combined sequence coverage of over 80%. One method is based on the partial acidic hydrolysis of the protein by 5 mM of sulfuric acid at 99 °C. Furthermore, we established a fast way for a tryptic digest without an alkylation step. We were able to show that the distinction of clones is possible simply by a brief visual comparison of the mass spectra. In this work, two clones originating from the same immunization gave the same fingerprints. Later, a hybridoma sequencing confirmed the sequence identity of these sister clones. In order to automate the spectral comparison for larger libraries of antibodies, we developed the online software ABID 2.0. This open-source software determines the number of matching peptides in the fingerprint spectra. We propose that publications and other documents critically relying on monoclonal antibodies with unknown amino acid sequences should include at least one antibody fingerprint. By fingerprinting an antibody in question, its identity can be confirmed by comparison with a library spectrum at any time and context. MDPI 2022-04-14 /pmc/articles/PMC9036215/ /pubmed/35466280 http://dx.doi.org/10.3390/antib11020027 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Tscheuschner, Georg Kaiser, Melanie N. Lisec, Jan Beslic, Denis Muth, Thilo Krüger, Maren Mages, Hans Werner Dorner, Brigitte G. Knospe, Julia Schenk, Jörg A. Sellrie, Frank Weller, Michael G. MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour |
title | MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour |
title_full | MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour |
title_fullStr | MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour |
title_full_unstemmed | MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour |
title_short | MALDI-TOF-MS-Based Identification of Monoclonal Murine Anti-SARS-CoV-2 Antibodies within One Hour |
title_sort | maldi-tof-ms-based identification of monoclonal murine anti-sars-cov-2 antibodies within one hour |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9036215/ https://www.ncbi.nlm.nih.gov/pubmed/35466280 http://dx.doi.org/10.3390/antib11020027 |
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