Optimization of Zika virus envelope protein production for ELISA and correlation of antibody titers with virus neutralization in Mexican patients from an arbovirus endemic region
BACKGROUND: Zika virus (ZIKV) has become a global threat with immediate need for accurate diagnostics, efficacious vaccines and therapeutics. Several ZIKV envelope (Env)-based vaccines have been developed recently. However, many commercially available ZIKV Env are based on the African lineage and produced in insect cells. Here, we sought to produce Asian-lineage ZIKV Env in mammalian cells for research and clinical applications.
METHODS: We designed various gene expression constructs to optimize the production of ZIKV using prM-Env and full or C-terminal truncations of Env; with or without a rat CD4 fusion partner to allow large-scale production of soluble protein in mammalian HEK293 cells. Protein expression was verified by mass spectrometry and western-blot with a pan-flavivirus antibody, a ZIKV Env monoclonal antibody and with immune sera from adenoviral (ChAdOx1) ZIKV Env-vaccinated mice. The resulting Env-CD4 was used as a coating reagent for immunoassay (ELISA) using both mouse and human seropositive sera.
RESULTS: Replacement of the C-terminus transmembrane Env domain by a rat CD4 and addition of prM supported optimal expression and secretion of Env. Binding between the antigens and the antibodies was similar to binding when using commercially available ZIKV Env reagents. Furthermore, antibodies from ZIKV patients bound ZIKV Env-CD4 in ELISA assays, whereas sera from healthy blood donors yielded minimal OD background. The serological outcomes of this assay correlated also with ZIKV neutralisation capacity in vitro.
CONCLUSIONS: Results obtained from this study indicate the potential of the Asian-lineage Zika Env-CD4 and Env proteins in ELISA assays to monitor humoral immune responses in upcoming clinical trials as well as a sero-diagnostic tool in ZIKV infection.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2018 |
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| Erschienen: |
2018 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:15 |
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| Enthalten in: |
Virology journal - 15(2018), 1 vom: 27. Dez., Seite 193 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Kim, Young Chan [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 21.01.2019 Date Revised 29.01.2022 published: Electronic Citation Status MEDLINE |
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| doi: |
10.1186/s12985-018-1104-6 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| PPN (Katalog-ID): |
NLM292164904 |
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| 100 | 1 | |a Kim, Young Chan |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Optimization of Zika virus envelope protein production for ELISA and correlation of antibody titers with virus neutralization in Mexican patients from an arbovirus endemic region |
| 264 | 1 | |c 2018 | |
| 336 | |a Text |b txt |2 rdacontent | ||
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| 338 | |a ƒa Online-Ressource |b cr |2 rdacarrier | ||
| 500 | |a Date Completed 21.01.2019 | ||
| 500 | |a Date Revised 29.01.2022 | ||
| 500 | |a published: Electronic | ||
| 500 | |a Citation Status MEDLINE | ||
| 520 | |a BACKGROUND: Zika virus (ZIKV) has become a global threat with immediate need for accurate diagnostics, efficacious vaccines and therapeutics. Several ZIKV envelope (Env)-based vaccines have been developed recently. However, many commercially available ZIKV Env are based on the African lineage and produced in insect cells. Here, we sought to produce Asian-lineage ZIKV Env in mammalian cells for research and clinical applications | ||
| 520 | |a METHODS: We designed various gene expression constructs to optimize the production of ZIKV using prM-Env and full or C-terminal truncations of Env; with or without a rat CD4 fusion partner to allow large-scale production of soluble protein in mammalian HEK293 cells. Protein expression was verified by mass spectrometry and western-blot with a pan-flavivirus antibody, a ZIKV Env monoclonal antibody and with immune sera from adenoviral (ChAdOx1) ZIKV Env-vaccinated mice. The resulting Env-CD4 was used as a coating reagent for immunoassay (ELISA) using both mouse and human seropositive sera | ||
| 520 | |a RESULTS: Replacement of the C-terminus transmembrane Env domain by a rat CD4 and addition of prM supported optimal expression and secretion of Env. Binding between the antigens and the antibodies was similar to binding when using commercially available ZIKV Env reagents. Furthermore, antibodies from ZIKV patients bound ZIKV Env-CD4 in ELISA assays, whereas sera from healthy blood donors yielded minimal OD background. The serological outcomes of this assay correlated also with ZIKV neutralisation capacity in vitro | ||
| 520 | |a CONCLUSIONS: Results obtained from this study indicate the potential of the Asian-lineage Zika Env-CD4 and Env proteins in ELISA assays to monitor humoral immune responses in upcoming clinical trials as well as a sero-diagnostic tool in ZIKV infection | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Research Support, Non-U.S. Gov't | |
| 650 | 4 | |a CD4 fusion tag | |
| 650 | 4 | |a ELISA | |
| 650 | 4 | |a Envelope protein | |
| 650 | 4 | |a Mexican patients | |
| 650 | 4 | |a Neutralizing antibodies | |
| 650 | 4 | |a Protein production | |
| 650 | 4 | |a Zika virus | |
| 650 | 7 | |a Antibodies, Neutralizing |2 NLM | |
| 650 | 7 | |a Antibodies, Viral |2 NLM | |
| 650 | 7 | |a CD4 Antigens |2 NLM | |
| 650 | 7 | |a Recombinant Fusion Proteins |2 NLM | |
| 650 | 7 | |a Viral Envelope Proteins |2 NLM | |
| 700 | 1 | |a Lopez-Camacho, Cesar |e verfasserin |4 aut | |
| 700 | 1 | |a Nettleship, Joanne E |e verfasserin |4 aut | |
| 700 | 1 | |a Rahman, Nahid |e verfasserin |4 aut | |
| 700 | 1 | |a Hill, Michelle L |e verfasserin |4 aut | |
| 700 | 1 | |a Silva-Reyes, Laura |e verfasserin |4 aut | |
| 700 | 1 | |a Ortiz-Martinez, Georgina |e verfasserin |4 aut | |
| 700 | 1 | |a Figueroa-Aguilar, Gloria |e verfasserin |4 aut | |
| 700 | 1 | |a Mar, María Antonieta |e verfasserin |4 aut | |
| 700 | 1 | |a Vivanco-Cid, Héctor |e verfasserin |4 aut | |
| 700 | 1 | |a Rollier, Christine S |e verfasserin |4 aut | |
| 700 | 1 | |a Zitzmann, Nicole |e verfasserin |4 aut | |
| 700 | 1 | |a Viveros-Sandoval, Martha Eva |e verfasserin |4 aut | |
| 700 | 1 | |a Owens, Raymond J |e verfasserin |4 aut | |
| 700 | 1 | |a Reyes-Sandoval, Arturo |e verfasserin |4 aut | |
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