{Reference Type}: Journal Article
{Title}: Flow Cytometry-Based Measurement of Antibodies Specific for Cell Surface-Expressed Folded SARS-CoV-2 Receptor-Binding Domains.
{Author}: Sehgal ANA;Safran J;Kratzer B;Gattinger P;Stieger RB;Musiejovsky L;Trapin D;Ettel P;Körmöczi U;Rottal A;Borochova K;Dorofeeva Y;Tulaeva I;Weber M;Grabmeier-Pfistershammer K;Perkmann T;Wiedermann U;Valenta R;Pickl WF;
{Journal}: Vaccines (Basel)
{Volume}: 12
{Issue}: 4
{Year}: 2024 Apr 1
{Factor}: 4.961
{DOI}: 10.3390/vaccines12040377
{Abstract}: <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.