A High Through-put Assay for Circulating Antibodies Directed against the S Protein of Severe Acute Respiratory Syndrome Corona virus 2

Svenja Weiss; Jeromine Klingler; Catarina Hioe; Fatima Amanat; Ian Baine; Erna Milunka Kojic; Jonathan Stoever; Sean Liu; Denise Jurczyszak; Maria Bermudez-Gonzalez; Viviana Simon; Florian Krammer; Susan Zolla-Pazner

The reverse transcription polymerase chain reaction (RT-PCR) test is currently being used for the qualitative detection of SARS-CoV-2 nucleic acids in specimens from the upper and lower respiratory tract. 1,2 Molecular testing is well established and has been used in clinical laboratories throughout the world for two decades. In contrast, testing of serum and other bodily fluids for antibodies (Abs) to infectious diseases such as syphilis, typhoid and diphtheria have been used for over a century. 3, 4 Antibody (Ab) assays are most useful for identifying individuals who have been infected with a particular pathogen and seroconverted. As such, they can be particularly valuable, for example, for identification of subjects who have had asymptomatic viral infections and those who have recovered and would no longer be positive in tests for viral nucleic acids. They would also be particularly useful for serosurveillance, to identify donors for COVID-19 plasma therapy, and to identify individuals who are potentially immune to reinfection. Antibody assays thus fill an essential gap both during and after the current SARS-CoV-2 pandemic. In fact, in one study, depending on the time of testing postinfection, the combined use of RT-PCR and Ab positivity provided an advantage over either test alone. 5 We and others 5-7 have described tests for assessing the presence of Abs to SARS-CoV-2 in serum and plasma using the enzyme-linked immunosorbent assay (ELISA) platform with a recombinant form of the S protein of the virus and/or the central portion of this molecule identified as the receptor binding domain (RBD), consisting of amino acids 319-541. 7-9 We report here a modification of the ELISA assay in which beads labeled with a particular fluorochrome signature are coated with the soluble recombinant S protein or RBD, incubated with serum, biotinylated anti-human total Ig Abs, and phycoerythrin (PE)-labeled streptavidin. The readout is performed with a laser-based instrument. This is a high through-put assay that offers the advantages of being able to prepare the antigen-coated beads for thousands of tests in a single day and using at least 20-fold less antigen than is required for . CC-BY-NC-ND 4.0 International license It is made available under a author/funder, who has granted medRxiv a license to display the preprint in perpetuity.