Flow Cytometry-Based Measurement of Antibodies Specific for Cell Surface-Expressed Folded SARS-CoV-2 Receptor-Binding Domains

Al Nasar Ahmed Sehgal, Jera Safran, Bernhard Kratzer, Pia Gattinger, Robert B Stieger, Laszlo Musiejovsky, Doris Trapin, Paul Ettel, Ulrike Körmöczi, Arno Rottal, Kristina Borochova, Yulia Dorofeeva, Inna Tulaeva, Milena Weber, Katharina Grabmeier-Pfistershammer, Thomas Perkmann, Ursula Wiedermann, Rudolf Valenta, Winfried F Pickl

Publikation: Beitrag in Fachzeitschrift (peer-reviewed)Artikel in Fachzeitschrift


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.

PublikationsstatusVeröffentlicht - 01 Apr. 2024


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