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- Title
Flow Cytometry-Based Measurement of Antibodies Specific for Cell Surface-Expressed Folded SARS-CoV-2 Receptor-Binding Domains.
- Authors
Sehgal, Al Nasar Ahmed; Safran, Jera; Kratzer, Bernhard; Gattinger, Pia; Stieger, Robert B.; Musiejovsky, Laszlo; Trapin, Doris; Ettel, Paul; Körmöczi, Ulrike; Rottal, Arno; Borochova, Kristina; Dorofeeva, Yulia; Tulaeva, Inna; Weber, Milena; Grabmeier-Pfistershammer, Katharina; Perkmann, Thomas; Wiedermann, Ursula; Valenta, Rudolf; Pickl, Winfried F.
- Abstract
Background: COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has now become endemic and is currently one of the important respiratory virus infections regularly affecting mankind. The assessment of immunity against SARS-CoV-2 and its variants is important for guiding active and passive immunization and SARS-CoV-2-specific treatment strategies. Methods: We here devised a novel flow cytometry-based diagnostic platform for the assessment of immunity against cell-bound virus antigens. This platform is based on a collection of HEK-293T cell lines which, as exemplified in our study, stably express the receptor-binding domains (RBDs) of the SARS-CoV-2 S-proteins of eight major SARS-CoV-2 variants, ranging from Wuhan-Hu-1 to Omicron. Results: RBD-expressing cell lines stably display comparable levels of RBD on the surface of HEK-293T cells, as shown with anti-FLAG-tag antibodies directed against a N-terminally introduced 3x-FLAG sequence while the functionality of RBD was proven by ACE2 binding. We exemplify the usefulness and specificity of the cell-based test by direct binding of IgG and IgA antibodies of SARS-CoV-2-exposed and/or vaccinated individuals in which the assay shows a wide linear performance range both at very low and very high serum antibody concentrations. In another application, i.e., antibody adsorption studies, the test proved to be a powerful tool for measuring the ratios of individual variant-specific antibodies. Conclusion: We have established a toolbox for measuring SARS-CoV-2-specific immunity against cell-bound virus antigens, which may be considered as an important addition to the armamentarium of SARS-CoV-2-specific diagnostic tests, allowing flexible and quick adaptation to new variants of concern.
- Subjects
SARS-CoV-2; IMMUNOGLOBULINS; FLOW measurement
- Publication
Vaccines, 2024, Vol 12, Issue 4, p377
- ISSN
2076-393X
- Publication type
Article
- DOI
10.3390/vaccines12040377