A Novel In-Cell ELISA Assay Allows Rapid and Automated Quantification of SARS-CoV-2 to Analyze Neutralizing Antibodies and Antiviral Compounds

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.

The Utility of Specific Antibodies Against SARS-CoV-2 in Laboratory Diagnosis

“…A diagnostic specificity of 100.0% and sensitivity of 98.31% was achieved in a newly established ELISA using 59 sera of infected or vaccinated animals including ferrets, raccoon dogs, hamsters, rabbits, chickens, cattle, and a cat and a total of 220 antibody-negative sera of the same animal species ( Wernike et al, 2020 ). What’s more, Schöler et al (2020) established a SARS-CoV-2 neutralization assay employing an in-cell ELISA (icELISA) approach. It allows rapid (<48 h in total, read-out in seconds) and automated quantification of COVID-19 infection in cell culture to evaluate the efficacy of NAbs and antiviral drugs using reagents and equipment present in most routine diagnostics departments ( Schöler et al, 2020 ).…”

“…What’s more, Schöler et al (2020) established a SARS-CoV-2 neutralization assay employing an in-cell ELISA (icELISA) approach. It allows rapid (<48 h in total, read-out in seconds) and automated quantification of COVID-19 infection in cell culture to evaluate the efficacy of NAbs and antiviral drugs using reagents and equipment present in most routine diagnostics departments ( Schöler et al, 2020 ). A report on a microfluidic, multiplexed POC test based on multiple SARS-CoV-2 antigens—S, N, and RBD—shows good concordance with a live virus microneutralization assay and successfully tracked the longitudinal evolution of the antibody response in infected individuals ( Heggestad et al, 2020 ).…”

Universally available herbal teas based on sage and perilla elicit potent antiviral activity against SARS-CoV-2 variants of concern by HMOX-1 upregulation in human cells

“…1d). To confirm, that the experimental setup allows reporting of antiviral activity affecting post-entry steps, we included a SARS-CoV-2 convalescent serum sample with shown neutralizing capacity 23 in our analysis. We observed that the perilla and sage infusions outperformed the effect of the convalescent serum (Fig.…”

NeutrobodyPlex—monitoring SARS‐CoV‐2 neutralizing immune responses using nanobodies

“…Compared with other assays, the Neutro-bodyPlex can be performed in a fully automated, high-throughput manner and can be readily used for large cohort screening. Since it requires only non-living and non-infectious viral material, costs and safety conditions can be significantly reduced (Muruato et al, 2020;Scholer et al, 2020;Tan et al, 2020). In addition, the Neutro-bodyPlex is highly sensitive, as low dilutions of serum (tested dilution: 1:400) are sufficient for analysis, significantly reducing patient material compared with standard assays.…”