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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 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 thirty-five monoclonal antibodies against SARS-CoV-2 to demonstrate how sequence-independent antibody identification can be achieved by simple means applied onto 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 twoDuring 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 thirty-five monoclonal antibodies against SARS-CoV-2 to demonstrate how sequence-independent antibody identification can be achieved by simple means applied onto 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 45 minutes 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 (https://gets.shinyapps.io/ABID/). 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.zeige mehrzeige weniger

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Metadaten
Autor*innen:Georg TscheuschnerORCiD, Melanie N. Kaiser, Jan LisecORCiD, D. Beslic, Thilo MuthORCiD, M. Krüger, H.-W. Mages, B. G. Dorner, J. Knospe, J. A. Schenk, F. Sellrie, Michael G. WellerORCiD
Dokumenttyp:Sonstiges
Veröffentlichungsform:Graue Literatur
Sprache:Englisch
Titel des übergeordneten Werkes (Englisch):Preprints
Jahr der Erstveröffentlichung:2022
Organisationseinheit der BAM:1 Analytische Chemie; Referenzmaterialien
1 Analytische Chemie; Referenzmaterialien / 1.5 Proteinanalytik
1 Analytische Chemie; Referenzmaterialien / 1.7 Organische Spuren- und Lebensmittelanalytik
VP Vizepräsident
VP Vizepräsident / VP.1 eScience
Veröffentlichende Institution:Bundesanstalt für Materialforschung und -prüfung (BAM)
Verlag:MDPI
Verlagsort:Basel
Aufsatznummer:2022030229
Erste Seite:1
Letzte Seite:24
DDC-Klassifikation:Naturwissenschaften und Mathematik / Chemie / Analytische Chemie
Freie Schlagwörter:ABID; Acidic cleavage; Antibody identification; Antibody light chain; Antibody subclass; Database; Identity; MALDI-TOF-MS; Monoclonal antibody; Online software; Peak overlap; Peptide coverage; Peptide mass fingerprinting; Reproducibility crisis; SARS-CoV-2 antibody; Sequencing; Sulfuric acid; Traceability; Trypsin
Themenfelder/Aktivitätsfelder der BAM:Chemie und Prozesstechnik
DOI:10.20944/preprints202203.0229.v1
URN:urn:nbn:de:kobv:b43-545182
ISSN:2310-287X
Verfügbarkeit des Dokuments:Datei für die Öffentlichkeit verfügbar ("Open Access")
Lizenz (Deutsch):License LogoCreative Commons - CC BY - Namensnennung 4.0 International
Datum der Freischaltung:23.03.2022
Referierte Publikation:Nein
Schriftenreihen ohne Nummerierung:Arbeitspapiere der BAM
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