Abstract
The coronavirus disease 2019 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently the most pressing medical and socioeconomic challenge. Constituting important correlates of protection, the determination of virus-neutralizing antibodies (NAbs) is indispensable for convalescent plasma selection, vaccine candidate evaluation, and immunity certificates. In contrast to standard serological ELISAs, plaque reduction neutralization tests (PRNTs) are laborious, time-consuming, expensive, and restricted to specialized laboratories. To replace microscopic counting-based SARS-CoV-2 PRNTs by a novel assay exempt from genetically modified viruses, which are inapplicable in most diagnostics departments, we established a simple, rapid, and automated SARS-CoV-2 neutralization assay employing an in-cell ELISA (icELISA) approach. After optimization of various parameters such as virus-specific antibodies, cell lines, virus doses, and duration of infection, SARS-CoV-2-infected cells became amenable as direct antigen source for quantitative icELISA. Antiviral agents such as human sera containing NAbs or antiviral interferons dose dependently reduced the SARS-CoV-2-specific signal. Applying increased infectious doses, the icELISA-based neutralization test (icNT) was superior to PRNT in discriminating convalescent sera with high from those with intermediate neutralizing capacities. In addition, the icNT was found to be specific, discriminating between SARS-CoV-2-specific NAbs and those raised against other coronaviruses. Altogether, the SARS-CoV-2 icELISA test allows rapid (<48 h in total, read-out in seconds) and automated quantification of virus infection in cell culture to evaluate the efficacy of NAbs and antiviral drugs using reagents and equipment present in most routine diagnostics departments.
Highlights
By the time of writing, more than 29 million people experienced a laboratory confirmed infection with the severe acute respiratory syndrome coronavirus (SARS-CoV)-2 and more than 930,000 people died while having coronavirus infectious disease 19 (COVID-19)
The actual data acquisition is conducted within seconds, using standard ELISA plate readers present in most routine diagnostics departments
Since the detection antibodies can be applied in in-cell ELISA (icELISA) and icELISA-based neutralization test (icNT) in relatively high dilutions (1/5,000 to 1/10,000 and 1/2,000 of primary and secondary antibody, respectively), the assay is relatively cheap
Summary
By the time of writing, more than 29 million people experienced a laboratory confirmed infection with the severe acute respiratory syndrome coronavirus (SARS-CoV)-2 and more than 930,000 people died while having coronavirus infectious disease 19 (COVID-19). Together with four seasonal human CoVs (HCoVs) and the two other emerging HCoVs SARS-CoV-1 and MERSCoV, SARS-CoV-2 is the seventh HCoV causing widespread human diseases [2]. In addition to its genetic similarity, SARS-CoV-2 shares some clinical characteristics with SARS-CoV-1 [4] and exhibits some highly relevant particularities such as an increased spreading efficacy and the length of the course of disease [5]. On January 31, the WHO declared the SARS-CoV-2 outbreak a Public Health Emergency of International Concern. Even countries that managed the first wave must prepare in terms of diagnostics capacities for potential future re-emergences
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