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Isolated novel coronavirus monoclonal antibody or antigen binding part thereof

A monoclonal antibody, coronavirus technology, applied in the fields of molecular biology and cellular immunology, can solve problems such as complex components

Active Publication Date: 2020-09-04
JIANGSU PROVINCIAL CENT FOR DISEASE CONTROL & PREVENTION PUBLIC HEALTH RES INST OF JIANGSU PROVINCE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the complex composition of plasma and plasma products, there may be potential risk factors

Method used

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  • Isolated novel coronavirus monoclonal antibody or antigen binding part thereof
  • Isolated novel coronavirus monoclonal antibody or antigen binding part thereof
  • Isolated novel coronavirus monoclonal antibody or antigen binding part thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0083] Example 1 Antibody Screening

[0084] 1. Phage library construction

[0085] 1. Collect peripheral blood from patients with COVID-19 during recovery period, and separate mononuclear cells (PBMC) from peripheral blood

[0086]In this project, on February 14, 2020, after informed consent, 20ml of peripheral blood was collected from 5 patients diagnosed with COVID-19 before discharge. The 5 patients belonged to the same transmission chain, and none of them were severe cases. After treatment, they were discharged from the hospital from February 15th to 22nd and were isolated at home. Mononuclear cells (PBMCs) in 20 ml of heparin-anticoagulated blood were separated by density gradient centrifugation using GE's Ficoll-Paque PLUS.

[0087] 2. RNA extraction and cDNA synthesis in PBMC

[0088] Use QIAGEN's RNeasy Mini Kit to extract PBMC cell RNA, and then use Roche's first strand synthesis kit (Transcriptor First Strand cDNA Synthesis Kit, Roche, Cat No.: 04896866001) to re...

Embodiment 2

[0130] Example 2 Indirect ELISA detection antibody binding specificity to S-RBD and S-ECD

[0131] Three human antibodies were selected from the 159 antibodies obtained through screening, and they were constructed into IgG-form human full-molecular antibodies (the three antibodies were named FC05, FC08, and FC11), expressed in 293F cells, and purified with Protein A Backup.

[0132] The FC05 antibody sequence is as follows:

[0133] The CDR1 sequence of the heavy chain variable region is shown in SEQ ID NO.1, the sequence of the CDR2 of the heavy chain variable region is shown in SEQ ID NO.2, and the CDR3 sequence of the heavy chain variable region is shown in SEQ ID NO.3; The CDR1 sequence of the light chain variable region is shown in SEQ ID NO.5, the CDR2 sequence of the light chain variable region is shown in SEQ ID NO.6, and the CDR3 sequence of the light chain variable region is shown in SEQ ID NO.7. The amino acid sequence of the heavy chain variable region is shown i...

Embodiment 3

[0138] Embodiment 3 Immunoprecipitation experiment of antibody, S-RBD and S-ECD

[0139] In the early stage, Western Blot was used to detect the binding specificity of the three antibodies to S-RBD and S-ECD, and it was found that none of the three antibodies reacted with S-RBD and S-ECD after SDS-PAGE, indicating that the three antibodies were all conformational epitope. Therefore, the immunoprecipitation (Immunoprecipitation, IP) method was used to detect the binding specificity of the antibody to the target protein, as follows:

[0140] Combine FC05, FC08 and FC11 triclonal antibodies with 20μL Protein A beads at room temperature for 2min, then wash away unbound antibodies with 20mM sodium phosphate. Then 20 μg of target antigens (S-RBD and S-ECD) were added to the antibody and Protein A gel mixture, and combined at room temperature for 2 minutes. Wash away the unbound antigen with 20mM sodium phosphate, elute the antigen-antibody complex with 30μl Gly-HCl buffer (pH 3.0)...

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Abstract

The invention discloses an isolated novel coronavirus monoclonal antibody or an antigen binding part thereof. The monoclonal antibody is specifically bound to S protein of the novel coronavirus. The monoclonal antibody contains a heavy chain variable region of SEQ ID NO.1-3 amino acid sequences and a light chain variable region of SEQ ID NO.5-7 amino acid sequences. The monoclonal antibody can beused to detect the existence of the novel coronavirus. In addition, the monoclonal antibody has neutralizing activity, so that can be used for the development of drugs for preventing or treating novelcoronavirus infection.

Description

technical field [0001] The invention belongs to the field of cellular immunology and molecular biology, and relates to an isolated novel coronavirus monoclonal antibody or an antigen-binding part thereof. Background technique [0002] The International Committee on Taxonomy of Viruses named the new coronavirus as SARS-CoV-2, and the World Health Organization named the pneumonia caused by this virus as COVID-19. The virus is highly contagious and spreads widely. The virus can quickly adapt to the human environment, and has the ability to spread during the incubation period after infection. At the same time, some asymptomatic infections have been reported, and viral nucleic acids have even been detected in various animals. These factors make the prevention and control of the virus very complicated, and there are currently no effective therapeutic drugs and vaccines on the market. [0003] SARS-CoV-2 belongs to the genus Coronaviridae and is a single-stranded positive-sense R...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10C07K16/46C12N15/13G01N33/577G01N33/569A61K39/42A61P31/14
CPCA61K2039/505A61P31/14C07K16/005C07K16/10C07K2317/31C07K2317/56C07K2317/565C07K2317/76C07K2317/92G01N33/56983G01N33/577G01N2333/165
Inventor 朱凤才张黎潘红星李靖欣郭喜玲王祥喜
Owner JIANGSU PROVINCIAL CENT FOR DISEASE CONTROL & PREVENTION PUBLIC HEALTH RES INST OF JIANGSU PROVINCE
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