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. 2021 Aug:294:114183.
doi: 10.1016/j.jviromet.2021.114183. Epub 2021 May 11.

Development of an immunochromatographic kit to detect severe acute respiratory syndrome coronavirus 2

Satoshi Oshiro  1 , Yoko Tabe  2 , Keiji Funatogawa  3 , Kaori Saito  2 , Tatsuya Tada  1 , Tomomi Hishinuma  1 , Naeko Mizutani  1 , Makoto Akiwa  4 , Jun-Ichiro Sekiguchi  4 , Takashi Miida  2 , Teruo Kirikae  5
Affiliations

Development of an immunochromatographic kit to detect severe acute respiratory syndrome coronavirus 2

Satoshi Oshiro et al. J Virol Methods. 2021 Aug.

Abstract

Background: The novel severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for the worldwide coronavirus disease-19 (COVID-19) pandemic, starting in late 2019. The standard diagnostic methods to detect SARS-CoV-2 are PCR-based genetic assays. Antigen-antibody-based immunochromatographic assays are alternative methods of detecting this virus. Rapid diagnosis kits to detect SARS-CoV-2 are urgently needed.

Study design: Three monoclonal antibodies against SARS-CoV-2 nucleocapsid (N) protein were used to develop an antigen-antibody-based immunochromatographic kit to detect SARS-CoV-2. These assays were evaluated using nasopharyngeal swab specimens collected from patients suspected of having COVID-19.

Results: These assays detected recombinant SARS-CoV-2 N protein at concentrations >0.2 ng/mL within 10 min after protein loading, but did not detect the N proteins of Middle East respiratory syndrome coronavirus (MERS-CoV), human coronaviruses OC43 (HCoV-OC43) and 299E (HCoV-229E) and other pathogens causing respiratory infections. Nasopharyngeal swab specimens obtained 1～3, 4～9, and ≥ 10 days after symptom onset from COVID-19 patients diagnosed by RT-PCR showed positivity rates of 100 %, >80 %, and <30 %, respectively.

Conclusions: Kits using this immunochromatographic assay may be a rapid and useful tool for point-of-care diagnosis of COVID-19 when samples are obtained from patients 1～9 days after symptom onset.

Keywords: Coronavirus disease-19 (COVID-19); Immunochromatographic kit; Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

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Conflict of interest statement

The authors report no declarations of interest.

Figures

Fig. 1
Fig. 1
Determination of epitopes by direct ELISA. a) The amino acid structure of SARS-CoV-2 nucleocapsid (N) protein and the regions of recombinant SARS-CoV-2 N protein. b) Competition assay using full- and partial-length recombinant SARS-CoV-2 N proteins. Assays were performed using three mAbs (N1J7, N1K1 and N1K2) as primary antibodies and horseradish peroxidase-conjugated anti-rat IgG as secondary antibody.
Fig. 2
Fig. 2
Determination of the optimal mABs combination by sandwich ELISA. The primary (capture) antibodies consisted of N1J7, N1K1, N1K2, and their combinations (N1J7 and N1K1, N1J7 and N1K2 and N1K1 and N1K2). The antigen was recombinant SARS-CoV-2 N protein, and the secondary (detection) antibodies consisted of horseradish peroxidase-conjugated N1J7, N1K1 and N1K2.
Fig. 3
Fig. 3
Details of the immunochromatographic assay (KBM Linecheck nCoV) developed using the mAbs N1J7, N1K2 and N1K2. a) Samples showing a single line at the control position were negative for N protein, whereas samples showing two lines, one each at the test and control positions, were positive for N protein. b) Antigen detection limits of KBM Linecheck nCoV. Recombinant SARS-CoV-2 N protein was firstly diluted in PBS-0.1 % Tween 20, and further diluted in Tris-based buffer (pH7.6) to 12.8, 6.4, 3.2, 1.6, 0.8, 0.4, 0.2 and 0.1 ng/mL, and 200 μL of each solution were used to determine detection limits. Color development was measured by a C10066-10 immunochromato-reader (Hamamatsu Photonics Co., Hamamatsu, Japan).
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References

    1. Baek Y.H., Um J., Antigua K.J.C., et al. Development of a reverse transcription-loop-mediated isothermal amplification as a rapid early-detection method for novel SARS-CoV-2. Emerg. Microbes Infect. 2020;9(1):998–1007. doi: 10.1080/22221751.2020.1756698. - DOI - PMC - PubMed
    1. Guruprasad L. Human SARS CoV-2 spike protein mutations. Proteins. 2021 doi: 10.1002/prot.26042. - DOI - PMC - PubMed
    1. Kawachi S., Matsushita T., Sato T., et al. Multicenter prospective evaluation of a novel rapid immunochromatographic diagnostic kit specifically detecting influenza A H1N1 2009 virus. J. Clin. Virol. 2011;51(1):68–72. - PubMed
    1. Kogaki H., Uchida Y., Fujii N., et al. Novel rapid immunochromatographic test based on an enzyme immunoassay for detecting nucleocapsid antigen in SARS-associated coronavirus. J. Clin. Lab. Anal. 2005;19(4):150–159. - PMC - PubMed
    1. Leung K., Shum M.H., Leung G.M., et al. Early transmissibility assessment of the N501Y mutant strains of SARS-CoV-2 in the United Kingdom, October to November 2020. Euro Surveill. 2021;26(1) - PMC - PubMed

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