Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2019 Feb 4;9(1):1390.
doi: 10.1038/s41598-018-37747-5.

Development and Evaluation of a Multiplexed Immunoassay for Simultaneous Detection of Serum IgG Antibodies to Six Human Coronaviruses

Suvang U Trivedi  1 , Congrong Miao  2 , Joseph E Sanchez  2 , Hayat Caidi  2 , Azaibi Tamin  2 , Lia Haynes  2 , Natalie J Thornburg  3
Affiliations

Development and Evaluation of a Multiplexed Immunoassay for Simultaneous Detection of Serum IgG Antibodies to Six Human Coronaviruses

Suvang U Trivedi et al. Sci Rep. .

Abstract

Known human coronaviruses (hCoV) usually cause mild to moderate upper-respiratory tract illnesses, except SARS-CoV and MERS-CoV, which, in addition to mild illness can also be associated with severe respiratory diseases and high mortality rates. Well-characterized multiplexed serologic assays are needed to aid in rapid detection and surveillance of hCoVs. The present study describes development and evaluation of a multiplexed magnetic microsphere immunoassay (MMIA) to simultaneously detect immunoglobulin G (IgG) antibodies specific for recombinant nucleocapsid proteins (recN) from hCoVs 229E, NL63, OC43, HKU1, SARS-CoV, and MERS-CoV. We used paired human sera to screen for IgG with reactivity against six hCoVs to determine assay sensitivity, specificity and reproducibility. We found no signal interference between monoplex and multiplex assay formats (R2 range = 0.87-0.97). Screening of paired human sera using MMIA, resulted in 92 of 106 (sensitivity: 86%) as positive and 68 of 80 (specificity: 84%) as negative. This study serves as a proof of concept that it is feasible to develop and use a multiplexed microsphere immunoassay as a next generation screening tool for use in large scale seroprevalence studies of hCoVs.

PubMed Disclaimer

Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Titration of recombinant nucleocapsid (recN) proteins of hCoVs (A) 229E, (B) NL63, (C) OC43, (D) HKU1, (E) SARS-CoV and (F) MERS-CoV at concentrations of 1.0 µg (□), 2.5 µg (∆), and 5.0 µg (○) per conjugation reaction. Positive and negative controls for each hCoV were tested at dilutions 1:200, 1:400 and 1:800. Mean Fluorescence Intensity (MFI) values of negative control antigens (pET E. coli and His-BoNT) were subtracted from the MFI values of corresponding positive control antigen for each antigen.
Figure 2
Figure 2
Correlation of Mean Fluorescence Intensity (MFI) values between monoplex and multiplex microsphere immunoassays using positive control serum for (A) 229E, (B) NL63, (C) OC43, (D) HKU1, (E) SARS-CoV and (F) MERS-CoV.
Figure 3
Figure 3
Receiver Operating Characteristic curve (ROC) analysis to determine area under the curve (AUC) values for (A) 229E, (B) NL63, (C) OC43, (D) HKU1, (E) SARS-CoV and (F) MERS-CoV recN in MMIA using real-time RT-PCR as the reference diagnostic test.
Figure 4
Figure 4
Cross reactivity of hCoV recN conjugated beads with Positive control serum samples for all six hCoVs in a multiplex microsphere immunoassay (MMIA) @ 1:400 dilution.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Cabeca TK, Granato C, Bellei N. Epidemiological and clinical features of human coronavirus infections among different subsets of patients. Influenza Other Respir. Viruses. 2013;7:1040–1047. doi: 10.1111/irv.12101. - DOI - PMC - PubMed
    1. Su S, et al. Epidemiology, Genetic Recombination, and Pathogenesis of Coronaviruses. Trends Microbiol. 2016;24:490–502. doi: 10.1016/j.tim.2016.03.003. - DOI - PMC - PubMed
    1. Zumla A, Chan JF, Azhar EI, Hui DS, Yuen KY. Coronaviruses - drug discovery and therapeutic options. Nat. Rev. Drug Discov. 2016;15:327–347. doi: 10.1038/nrd.2015.37. - DOI - PMC - PubMed
    1. Wevers BA, van der Hoek L. Recently discovered human coronaviruses. Clin. Lab. Med. 2009;29:715–724. doi: 10.1016/j.cll.2009.07.007. - DOI - PMC - PubMed
    1. de Wit E, van Doremalen N, Falzarano D, Munster VJ. SARS and MERS: recent insights into emerging coronaviruses. Nat. Rev. Microbiol. 2016;14:523–534. doi: 10.1038/nrmicro.2016.81. - DOI - PMC - PubMed

Publication types

LinkOut - more resources

-