Scalable, semi-automated fluorescence reduction neutralization assay for qualitative assessment of Ebola virus-neutralizing antibodies in human clinical samples
Elena N Postnikova, James Pettitt, Collin J Van Ryn, Michael R Holbrook, Laura Bollinger, Shuǐqìng Yú, Yíngyún Caì, Janie Liang, Michael C Sneller, Peter B Jahrling, Lisa E Hensley, Jens H Kuhn, Mosoka P Fallah, Richard S Bennett, Cavan Reilly, Elena N Postnikova, James Pettitt, Collin J Van Ryn, Michael R Holbrook, Laura Bollinger, Shuǐqìng Yú, Yíngyún Caì, Janie Liang, Michael C Sneller, Peter B Jahrling, Lisa E Hensley, Jens H Kuhn, Mosoka P Fallah, Richard S Bennett, Cavan Reilly
Abstract
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: ClinicalTrials.gov NCT02431923.
Conflict of interest statement
The authors have declared that no competing interests exist.
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References
- Bukreyev AA, Chandran K, Dolnik O, Dye JM, Ebihara H, Leroy EM, et al. Discussions and decisions of the 2012–2014 International Committee on Taxonomy of Viruses (ICTV) Filoviridae Study Group, January 2012-June 2013. Arch Virol. 2014;159(4):821–30. Epub 2013/10/15. 10.1007/s00705-013-1846-9
- Kuhn JH. Ebolavirus and marburgvirus infections In: Jameson JL, Fauci AS, Kasper DL, Hauser SL, Longo DL, Loscalzo J, editors. Harrison's Principles of Internal Medicine. 2 20th ed Columbus, USA: McGraw-Hill Education; 2018. p. 1509–15.
- Ebola Outbreak Epidemiology Team. Outbreak of Ebola virus disease in the Democratic Republic of the Congo, April-May, 2018: an epidemiological study. Lancet. 2018;392(10143):213–21. Epub 2018/07/27. 10.1016/S0140-6736(18)31387-4 .
- Dyer O. Ebola: new outbreak appears in Congo a week after epidemic was declared over. Bmj. 2018;362:k3421 Epub 2018/08/09. 10.1136/bmj.k3421 .
- World Health Organization (WHO). Ebola situation report– 30 March 2016. . 2016.
- Meyer M, Malherbe DC, Bukreyev A. Can Ebola virus vaccines have universal immune correlates of protection? Trends Microbiol. 2019;27(1):8–16. Epub 2018/09/12. 10.1016/j.tim.2018.08.008
- Bramble MS, Hoff N, Gilchuk P, Mukadi P, Lu K, Doshi RH, et al. Pan-filovirus serum neutralizing antibodies in a subset of Congolese ebolavirus infection survivors. J Infect Dis. 2018;218(12):1929–36. Epub 2018/08/15. 10.1093/infdis/jiy453
- Rimoin AW, Lu K, Bramble MS, Steffen I, Doshi RH, Hoff NA, et al. Ebola virus neutralizing antibodies detectable in survivors of theYambuku, Zaire outbreak 40 years after infection. J Infect Dis. 2018;217(2):223–31. Epub 2017/12/19. 10.1093/infdis/jix584
- Logue J, Tuznik K, Follmann D, Grandits G, Marchand J, Reilly C, et al. Use of the Filovirus Animal Non-Clinical Group (FANG) Ebola virus immuno-assay requires fewer study participants to power a study than the Alpha Diagnostic International assay. J Virol Methods. 2018;255:84–90. Epub 2018/02/27. 10.1016/j.jviromet.2018.02.018
- Shurtleff AC, Bloomfield HA, Mort S, Orr SA, Audet B, Whitaker T, et al. Validation of the filovirus plaque assay for use in preclinical studies. Viruses. 2016;8(4):113 Epub 2016/04/26. 10.3390/v8040113
- Konduru K, Shurtleff AC, Bavari S, Kaplan G. High degree of correlation between Ebola virus BSL-4 neutralization assays and pseudotyped VSV BSL-2 fluorescence reduction neutralization test. J Virol Methods. 2018;254:1–7. Epub 2018/01/23. 10.1016/j.jviromet.2018.01.003
- Baize S, Pannetier D, Oestereich L, Rieger T, Koivogui L, Magassouba NF, et al. Emergence of Zaire Ebola virus disease in Guinea. N Engl J Med. 2014;371(15):1418–25. Epub 2014/04/18. 10.1056/NEJMoa1404505 .
- Postnikova E, Cong Y, DeWald LE, Dyall J, Yu S, Hart BJ, et al. Testing therapeutics in cell-based assays: Factors that influence the apparent potency of drugs. PLoS One. 2018;13(3):e0194880 Epub 2018/03/23. 10.1371/journal.pone.0194880
- Kilgore N, Nuzum EO. An interagency collaboration to facilitate development of filovirus medical countermeasures. Viruses. 2012;4(10):2312–6. Epub 2012/12/04. 10.3390/v4102312
- Kennedy SB, Bolay F, Kieh M, Grandits G, Badio M, Ballou R, et al. Phase 2 placebo-controlled trial of two vaccines to prevent Ebola in Liberia. N Engl J Med. 2017;377(15):1438–47. Epub 2017/10/12. 10.1056/NEJMoa1614067
Source: PubMed