A novel in-cell ELISA assay allows rapid and automated quantification of SARS-CoV-2 to analyse neutralizing antibodies and antiviral compounds

  1. Lara Schöler
  2. Vu Thuy Khanh Le-Trilling
  3. Mareike Eilbrecht
  4. Denise Mennerich
  5. Olympia E. Anastasiou
  6. Adalbert Krawczyk
  7. Anke Herrmann
  8. Ulf Dittmer
  9. Mirko Trilling

Reviewed by

Read the full article See related articles

Discuss this preprint

Start a discussion What are Sciety discussions?

Listed in

Log in to save this article

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, determination of virus-neutralizing antibodies (NAbs) is indispensable for convalescent plasma selection, vaccine candidate evaluation, and immunity certificates. In contrast to standard serology 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. Using commercially available nucleocapsid protein-specific antibodies, viral infection could easily be quantified in human and highly permissive Vero E6 cells by 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 icNT was superior to PRNT in discriminating convalescent sera with high from those with intermediate neutralizing capacities.

The SARS-CoV-2 icELISA test allows rapid (<48h in total, read-out in seconds) and automated quantification of virus infection in cell culture to evaluate the efficacy of NAbs as well as antiviral drugs, using reagents and equipment present in most routine diagnostics departments. We propose the icELISA and the icNT for COVID-19 research and diagnostics.

Article activity feed

  1. ScreenIT

    SciScore for 10.1101/2020.06.05.135806: (What is this?)

    Please note, not all rigor criteria are appropriate for all manuscripts.

    Table 1: Rigor

    Institutional Review Board StatementIRB: The collection of serum samples has been approved by the ethics committee of the medical faculty of the University of Duisburg-Essen (20-9208-BO).
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Anti-SARS-CoV-2 IgG antibodies were detected using ELISA detecting SARS-CoV-2 Spike protein (Euroimmun Medizinische Labordiagnostika, Germany) according to manufacturer’s instructions SARS-CoV-2 icELISA and icNT: Defined doses of SARS-CoV-2 were incubated with different serum dilutions for 1 h at 37°C prior to Vero E6 infection.
    Anti-SARS-CoV-2 IgG
    suggested: None
    The α-N mAb1 (ABIN6952435), α-N mAb2 (ABIN6952433), α-S Ab (ABIN1031551), and POD-coupled secondary antibodies (Dianova) were used.
    α-N
    suggested: None
    α-S
    suggested: None
    POD-coupled secondary antibodies (Dianova)
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Cells, viruses, interferons, and sera: Caco-2 (ATCC HTB-37) and Vero E6 (ATCC CRL-1586) were cultivated in Roswell Park Memorial Institute (RPMI) and Dulbecco’s minimal essential medium (DMEM), respectively, supplemented with 10% (v/v) FCS, penicillin, and streptomycin at 37°C in an atmosphere of 5% CO.
    Caco-2
    suggested: None
    SARS-CoV-2 was isolated from a patient sample using Vero E6 and confirmed by SARS-CoV-2 diagnostic qRT-PCR.
    Vero E6
    suggested: RRID:CVCL_XD71)

    Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

    Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

    Results from TrialIdentifier: No clinical trial numbers were referenced.

    Results from Barzooka: We found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

    Results from JetFighter: We did not find any issues relating to colormaps.

    Results from rtransparent:
    • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
    • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
    • No protocol registration statement was detected.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1101/2020.06.05.135806 on bioRxiv