Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes
Abstract Background To stop the spread of the COVID-19 disease, it is crucial to create molecular tools to investigate and diagnose COVID-19. Current efforts focus on developing specific neutralizing monoclonal antibodies (NmAbs) elicited against the receptor-binding domain (RBD). Methods In the pre...
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| Format: | Article |
| Language: | English |
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BMC
2024-02-01
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| Series: | Virology Journal |
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| Online Access: | https://doi.org/10.1186/s12985-024-02304-2 |
| _version_ | 1797275955079675904 |
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| author | Seyed Mostafa Mostafavi Zadeh Ali Ahmad Bayat Hosein Shahsavarani Feridoun Karimi-Busheri Jafar Kiani Roya Ghods Zahra Madjd |
| author_facet | Seyed Mostafa Mostafavi Zadeh Ali Ahmad Bayat Hosein Shahsavarani Feridoun Karimi-Busheri Jafar Kiani Roya Ghods Zahra Madjd |
| author_sort | Seyed Mostafa Mostafavi Zadeh |
| collection | DOAJ |
| description | Abstract Background To stop the spread of the COVID-19 disease, it is crucial to create molecular tools to investigate and diagnose COVID-19. Current efforts focus on developing specific neutralizing monoclonal antibodies (NmAbs) elicited against the receptor-binding domain (RBD). Methods In the present study, recombinant RBD (rRBD) protein was produced in E. coli, followed by immunizing mice with purified rRBD. ELISA was applied to screen the hybridomas for positive reactivity with rRBD protein. The linear and conformational epitopes of the mAbs were subsequently identified using western blot. Finally, the reactivity, affinity, and neutralization activity of the purified mAbs were evaluated using ELISA. Results All mAbs exhibited similar reactivity trends towards both eukaryotic RBD and prokaryotic rRBD in ELISA. Among them, 2E7-D2 and 2B4-G8 mAbs demonstrated higher reactivity than other mAbs. Additionally, in western blot assays, these two mAbs could detect reducing and non-reducing rRBD, indicating recognition of linear epitopes. Notably, five mAbs effectively blocked rRBD- angiotensin-converting enzyme 2 (ACE2) interaction, while two high-affinity mAbs exhibited potent neutralizing activity against eukaryotic RBD. Conclusion In the current study, we generated and characterized new RBD-specific mAbs using the hybridoma technique that recognized linear and conformational epitopes in RBD with neutralization potency. Our mAbs are novel candidates for diagnosing and treating SARS-CoV-2. |
| first_indexed | 2024-03-07T15:20:28Z |
| format | Article |
| id | doaj.art-f1373fe9d97743d9833da193ce701695 |
| institution | Directory Open Access Journal |
| issn | 1743-422X |
| language | English |
| last_indexed | 2024-03-07T15:20:28Z |
| publishDate | 2024-02-01 |
| publisher | BMC |
| record_format | Article |
| series | Virology Journal |
| spelling | doaj.art-f1373fe9d97743d9833da193ce7016952024-03-05T17:41:10ZengBMCVirology Journal1743-422X2024-02-0121111310.1186/s12985-024-02304-2Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopesSeyed Mostafa Mostafavi Zadeh0Ali Ahmad Bayat1Hosein Shahsavarani2Feridoun Karimi-Busheri3Jafar Kiani4Roya Ghods5Zahra Madjd6Oncopathology Research Center, Iran University of Medical SciencesMonoclonal Antibody Research Center, Avicenna Research Institute, ACECRLaboratory of Regenerative Medicine and Biomedical Innovations, Pasteur Institute of Iran, National Cell BankDepartment of Oncology, Faculty of Medicine, University of AlbertaOncopathology Research Center, Iran University of Medical SciencesOncopathology Research Center, Iran University of Medical SciencesOncopathology Research Center, Iran University of Medical SciencesAbstract Background To stop the spread of the COVID-19 disease, it is crucial to create molecular tools to investigate and diagnose COVID-19. Current efforts focus on developing specific neutralizing monoclonal antibodies (NmAbs) elicited against the receptor-binding domain (RBD). Methods In the present study, recombinant RBD (rRBD) protein was produced in E. coli, followed by immunizing mice with purified rRBD. ELISA was applied to screen the hybridomas for positive reactivity with rRBD protein. The linear and conformational epitopes of the mAbs were subsequently identified using western blot. Finally, the reactivity, affinity, and neutralization activity of the purified mAbs were evaluated using ELISA. Results All mAbs exhibited similar reactivity trends towards both eukaryotic RBD and prokaryotic rRBD in ELISA. Among them, 2E7-D2 and 2B4-G8 mAbs demonstrated higher reactivity than other mAbs. Additionally, in western blot assays, these two mAbs could detect reducing and non-reducing rRBD, indicating recognition of linear epitopes. Notably, five mAbs effectively blocked rRBD- angiotensin-converting enzyme 2 (ACE2) interaction, while two high-affinity mAbs exhibited potent neutralizing activity against eukaryotic RBD. Conclusion In the current study, we generated and characterized new RBD-specific mAbs using the hybridoma technique that recognized linear and conformational epitopes in RBD with neutralization potency. Our mAbs are novel candidates for diagnosing and treating SARS-CoV-2.https://doi.org/10.1186/s12985-024-02304-2Linear epitopeMonoclonal antibodyNeutralizing antibodyRBDSARS-CoV-2 |
| spellingShingle | Seyed Mostafa Mostafavi Zadeh Ali Ahmad Bayat Hosein Shahsavarani Feridoun Karimi-Busheri Jafar Kiani Roya Ghods Zahra Madjd Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes Virology Journal Linear epitope Monoclonal antibody Neutralizing antibody RBD SARS-CoV-2 |
| title | Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes |
| title_full | Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes |
| title_fullStr | Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes |
| title_full_unstemmed | Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes |
| title_short | Novel neutralizing SARS-CoV-2-specific mAbs offer detection of RBD linear epitopes |
| title_sort | novel neutralizing sars cov 2 specific mabs offer detection of rbd linear epitopes |
| topic | Linear epitope Monoclonal antibody Neutralizing antibody RBD SARS-CoV-2 |
| url | https://doi.org/10.1186/s12985-024-02304-2 |
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