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Abstract

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to be a major public health concern. Nucleocapsid (N) protein is the most abundant structural protein on SARS-CoV-2 virions and induces the production of antibodies at the early stage of infection. Large-scale preparation of N protein is essential for the development of immunoassays to detect antibodies to SARS-CoV-2 and the control of virus transmission. In this study, expression of water-soluble N protein was achieved through inducing protein expression at 25°C with 0.5 mM IPTG for 12 h. Western blot and ELISA showed that recombinant N protein could be recognized by sera collected from subjects immunized with Sinovac inactivated SARS-CoV-2 vaccine. Four monoclonal antibodies namely 2B1B1, 4D3A3, 5G1F8, and 7C6F5 were produced using hybridoma technology. Titers of all four monoclonal antibodies in ELISA reached more than 1.28×10 6.0. Moreover, all monoclonal antibodies could react specifically with N protein expressed by transfection of pcDNA3.1-N into BHK-21 cells in IPMA and IFA. These results indicated that water-soluble N protein retained high immunogenicity and possessed the same epitopes as that of native N protein on virions. In addition, the preparation of water-soluble N protein and its monoclonal antibodies laid the basis for the development of immunoassays for COVID-19 detection.
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Authors and Affiliations

Y.B. Wang
1
S.W. Wang
2
Q.Y. Jin
3
L.P. Chen
4
F.Q. Zhang
1
J.J. Shi
1
Y. Yin
5
Z.X. Fan
1
X.Y. Liu
6
L.P. Wang
6
P. Li
6

  1. School of Public Health, Xinxiang Medical University, Xinxiang 453003, P.R. China
  2. School of Basic Medical Sciences, Xinxiang Medical University, Xinxiang 453003, P.R. China
  3. Henan Provincial Key Laboratory of Animal Immunology, Henan Academy of Agricultural Sciences, Zhengzhou 450002, P.R. China
  4. Gushi County Center for Animal Disease Control and Prevention, Xinyang 465200, P.R. China
  5. Mingde College of Xinxiang Medical University, Xinxiang 453003, P.R. China
  6. School of Biological Engineering, Xinxiang University, Xinxiang 453003, P.R. China

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