Analysis of SARS-CoV-2 Antibodies in COVID-19 Convalescent Plasma using a Coronavirus Antigen Microarray

Rafael R. de Assis; Aarti Jain; Rie Nakajima; Algis Jasinskas; Jiin Felgner; Joshua M. Obiero; Oluwasanmi Adenaiye; Sheldon Tai; Filbert Hong; Philip Norris; Mars Stone; Graham Simmons; Anil Bagri; Martin Schreiber; Andreas Buser; Andreas Holbro; Manuel Battegay; Donald K. Milton; Huw Davies; Laurence M. Corash; Michael P. Busch; Philip L. Felgner; Saahir Khan

Background COVID-19 caused by the SARS-CoV-2 virus is a worldwide pandemic with significant morbidity and mortality estimates from 1-4% of confirmed cases 1 . The current case definition for confirmed SARS-CoV-2 infection relies on PCR-positive pharyngeal or respiratory specimens, with testing largely determined by presence of fever or respiratory symptoms in an individual at high epidemiologic risk. However, this case definition likely underestimates true prevalence, as individuals who develop subclinical infection that does not produce fever or respiratory symptoms are unlikely to be tested, and testing by PCR of pharyngeal or respiratory specimens is only around 60-80% sensitive depending on sampling location and technique and the patient's viral load 2 . Widespread testing within the United States is also severely limited by the lack of available testing kits and testing capacity limitations of available public and private laboratories. Therefore, the true prevalence of SARS-CoV-2 infection is likely much higher than currently reported case numbers would indicate. Serology can play an important role in defining the true prevalence of COVID-19, particularly for subclinical infection 2 . Early studies of serology demonstrate high sensitivity to detect confirmed SARS-CoV-2 infection, with antibodies to virus detected approximately 1 to 2 weeks after symptom onset 3 . Unlike PCR positivity, SARS-CoV-2 antibodies are detectable throughout the disease course and persist indefinitely 4 . Multiple serologic tests have been developed for COVID-19 5 including a recently FDA-approved lateral flow assay. However, these tests are limited to detection of antibodies against one or two antigens, and cross-reactivity with antibodies to other human coronaviruses that are present in all adults 6 is currently unknown. Prior use of serology for detection of emerging coronaviruses focused on antibodies against the spike (S) protein, particularly the S1 domain, and the nucleocapsid (N) protein 7 . However, the optimal set of antigens to detect strain-specific coronavirus antibodies remains unknown. Protein microarray technology can be used to detect antibodies of multiple isotypes against hundreds of antigens in a high throughput manner 8, 9 so is well suited to serologic surveillance studies. This technology, which has previously been applied to other emerging coronaviruses 10 , is based on detection of binding antibodies, which are wellcorrelated with neutralizing antibodies 11 but do not require viral culture in biosafety level 3 facilities. Recently, our group developed a coronavirus antigen microarray (CoVAM) that includes antigens from SARS-CoV-2 and tested it on human sera collected prior to the pandemic to demonstrate low cross-reactivity with antibodies from human coronaviruses that cause the common cold, particularly for the S1 domain 2 . Here, we further validate this methodology using convalescent blood specimens from COVID-19 cases confirmed by positive SARS-CoV-2 PCR.