Abstract
In light of an increasing number of vaccinated and convalescent individuals, there is a major need for the development of robust methods for the quantification of neutralizing antibodies; although, a defined correlate of protection is still missing. Sera from hospitalized COVID-19 patients suffering or not suffering from acute respiratory distress syndrome (ARDS) were comparatively analyzed by plaque reduction neutralization test (PRNT) and pseudotype-based neutralization assays to quantify their neutralizing capacity. The two neutralization assays showed comparable data. In case of the non-ARDS sera, there was a distinct correlation between the data from the neutralization assays on the one hand, and enzyme-linked immune sorbent assay (ELISA), as well as biophysical analyses, on the other hand. As such, surface plasmon resonance (SPR)-based assays for quantification of binding antibodies or analysis of the stability of the antigen–antibody interaction and inhibition of syncytium formation, determined by cell fusion assays, were performed. In the case of ARDS sera, which are characterized by a significantly higher fraction of RBD-binding IgA antibodies, there is a clear correlation between the neutralization assays and the ELISA data. In contrast to this, a less clear correlation between the biophysical analyses on the one hand and ELISAs and neutralization assays on the other hand was observed, which might be explained by the heterogeneity of the antibodies. To conclude, for less complex immune sera—as in cases of non-ARDS sera—combinations of titer quantification by ELISA with inhibition of syncytium formation, SPR-based analysis of antibody binding, determination of the stability of the antigen–antibody complex, and competition of the RBD-ACE2 binding represent alternatives to the classic PRNT for analysis of the neutralizing potential of SARS-CoV-2-specific sera, without the requirement for a BSL3 facility.
Highlights
After its emergence in December 2019, severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) rapidly spread all over the world
We analyzed sera from hospitalized patients suffering from COVID-19 with or without acute respiratory distress syndrome (ARDS) by enzyme-linked immune sorbent assay (ELISA), neutralization assays, surface plasmon resonance spectroscopy and syncytium formation assay (SFA)
ARDS and sera from non-ARDS patients were characterized by plaque reduction neutralization test (PRNT) and by pseudotyped lentiviral vector neutralization assays
Summary
After its emergence in December 2019, severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) rapidly spread all over the world. (source: COVID-19 Dashboard by the Center for Systems Science and Engineering (CSSE). There are more than 5.1 million deaths relating to SARS-CoV-2 infections worldwide (source: COVID-19 Dashboard by the Center for Systems Science and Engineering (CSSE) at Johns Hopkins University (JHU)). The vaccines (mRNA or adenoviral vectors) encode the spike protein (S) of SARS-CoV-2 as an antigen. The S1 domain contains the receptor-binding domain (RBD), which is a major target for neutralizing antibodies [4]. The S protein is the target of almost all neutralizing antibodies found in the convalescent sera or elicited by vaccines. In addition to the RBD-targeting antibodies, there are neutralizing antibodies, which bind to an antigenic “supersite” in the N-terminal domain involving residues 141 to 156 and residues 246 to 260 [5,6]
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