%0 Journal Article
%T Flow Cytometry-Based Measurement of Antibodies Specific for Cell Surface-Expressed Folded SARS-CoV-2 Receptor-Binding Domains.
%A Sehgal ANA
%A Safran J
%A Kratzer B
%A Gattinger P
%A Stieger RB
%A Musiejovsky L
%A Trapin D
%A Ettel P
%A Körmöczi U
%A Rottal A
%A Borochova K
%A Dorofeeva Y
%A Tulaeva I
%A Weber M
%A Grabmeier-Pfistershammer K
%A Perkmann T
%A Wiedermann U
%A Valenta R
%A Pickl WF
%J Vaccines (Basel)
%V 12
%N 4
%D 2024 Apr 1
%M 38675759
%F 4.961
%R 10.3390/vaccines12040377
%X <B>BACKGROUND: </B>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.<BR><B>METHODS: </B>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.<BR><B>RESULTS: </B>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.<BR><B>CONCLUSIONS: </B>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.