Scalable, semi-automated fluorescence reduction neutralization assay for qualitative assessment of Ebola virus-neutralizing antibodies in human clinical samples

PLoS One. 2019 Aug 27;14(8):e0221407. doi: 10.1371/journal.pone.0221407. eCollection 2019.


Antibody titers against a viral pathogen are typically measured using an antigen binding assay, such as an enzyme-linked immunosorbent assay (ELISA), which only measures the ability of antibodies to identify a viral antigen of interest. Neutralization assays measure the presence of virus-neutralizing antibodies in a sample. Traditional neutralization assays, such as the plaque reduction neutralization test (PRNT), are often difficult to use on a large scale due to being both labor and resource intensive. Here we describe an Ebola virus fluorescence reduction neutralization assay (FRNA), which tests for neutralizing antibodies, that requires only a small volume of sample in a 96-well format and is easy to automate. The readout of the FRNA is the percentage of Ebola virus-infected cells measured with an optical reader or overall chemiluminescence that can be generated by multiple reading platforms. Using blinded human clinical samples (EVD survivors or contacts) obtained in Liberia during the 2013-2016 Ebola virus disease outbreak, we demonstrate there was a high degree of agreement between the FRNA-measured antibody titers and the Filovirus Animal Non-clinical Group (FANG) ELISA titers with the FRNA providing information on the neutralizing capabilities of the antibodies.

Trial registration: NCT02431923.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, N.I.H., Intramural

MeSH terms

  • Animals
  • Antibodies, Blocking / immunology
  • Antibodies, Neutralizing / immunology*
  • Antibodies, Viral / immunology*
  • Chlorocebus aethiops
  • Disease Outbreaks
  • Ebolavirus / immunology*
  • Ebolavirus / pathogenicity
  • Enzyme-Linked Immunosorbent Assay
  • Hemorrhagic Fever, Ebola / immunology*
  • Hemorrhagic Fever, Ebola / prevention & control
  • Hemorrhagic Fever, Ebola / virology
  • Humans
  • Liberia
  • Neutralization Tests / methods
  • Vero Cells


  • Antibodies, Blocking
  • Antibodies, Neutralizing
  • Antibodies, Viral

Associated data

  • Dryad/10.5061/dryad.4mv6600