tetano
Editor, Senior Moderator
Pol J Vet Sci
. 2023 Dec 12;26(4):571-579.
doi: 10.24425/pjvs.2023.148277. Expression of water-soluble nucleocapsid protein of SARS-CoV-2 and analysis of its immunogenicity
Y B Wang[SUP] 1 [/SUP], S W Wang[SUP] 2 [/SUP], Q Y Jin[SUP] 3 [/SUP], L P Chen[SUP] 4 [/SUP], F Q Zhang[SUP] 1 [/SUP], J J Shi[SUP] 1 [/SUP], Y Yin[SUP] 5 [/SUP], Z X Fan[SUP] 1 [/SUP], X Y Liu[SUP] 6 [/SUP], L P Wang[SUP] 6 [/SUP], P Li[SUP] 6 [/SUP]
Affiliations
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.
Keywords: SARS-CoV-2; immunogenicity analysis; optimization of expression conditions; water-soluble nucleocapsid protein.
. 2023 Dec 12;26(4):571-579.
doi: 10.24425/pjvs.2023.148277. Expression of water-soluble nucleocapsid protein of SARS-CoV-2 and analysis of its immunogenicity
Y B Wang[SUP] 1 [/SUP], S W Wang[SUP] 2 [/SUP], Q Y Jin[SUP] 3 [/SUP], L P Chen[SUP] 4 [/SUP], F Q Zhang[SUP] 1 [/SUP], J J Shi[SUP] 1 [/SUP], Y Yin[SUP] 5 [/SUP], Z X Fan[SUP] 1 [/SUP], X Y Liu[SUP] 6 [/SUP], L P Wang[SUP] 6 [/SUP], P Li[SUP] 6 [/SUP]
Affiliations
- PMID: 38088301
- DOI: 10.24425/pjvs.2023.148277
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.
Keywords: SARS-CoV-2; immunogenicity analysis; optimization of expression conditions; water-soluble nucleocapsid protein.