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Anti-Ebola virus neutralizing monoclonal antibody

A technology of antibodies and carriers, applied in antiviral immunoglobulins, antibodies, chemical instruments and methods, etc., can solve problems such as vaccines and drugs that have not been approved for marketing

Inactive Publication Date: 2017-12-01
INST PASTEUR OF SHANGHAI CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] For such a highly contagious and pathogenic virulent virus, there are still no vaccines and drugs approved for marketing, and the WHO allows some drugs and vaccines to be used in emergencies

Method used

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  • Anti-Ebola virus neutralizing monoclonal antibody
  • Anti-Ebola virus neutralizing monoclonal antibody
  • Anti-Ebola virus neutralizing monoclonal antibody

Examples

Experimental program
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Effect test

preparation example Construction

[0162] Preparation of monoclonal antibodies

[0163] Antibodies of the present invention can be prepared by various techniques known to those skilled in the art. For example, an antigen of the invention may be administered to an animal to induce the production of monoclonal antibodies. For monoclonal antibodies, hybridoma technology can be used to prepare (see Kohler et al., Nature 256; 495, 1975; Kohler et al., Eur.J.Immunol.6:511, 1976; Kohler et al., Eur.J.Immunol. 6:292,1976; Hammerling et al., In Monoclonal Antibodies and T Cell Hybridomas, Elsevier, N.Y., 1981) or can be prepared by recombinant DNA methods (US Patent No. 4,816,567).

[0164] Representative myeloma cells are those that fuse efficiently, support stable high-level production of antibody by selected antibody-producing cells, and are sensitive to culture medium (HAT medium matrix), including myeloma cell lines, such as murine Myeloma cell lines, including those derived from MOPC-21 and MPC-11 mouse tumors (...

Embodiment 1

[0175] The construction of embodiment 1 Ebola DNA vaccine

[0176] 1.1 Acquisition of GP gene

[0177] There are 5 subtypes of Ebola virus, the Zaire subtype that broke out in most parts of West Africa in 2014, and a small number of Sudan subtypes, so the present invention hopes to obtain a strain with cross-neutralizing activity against Zaire subtype and Sudan subtype antibodies. First, Ebola virus EBOV subtype Zaire (Ref ID: gb|U23187.1) and Ebola virus EBOV subtype Sudan (Ref ID: lcl|Query_182515) were purchased from Sino Biological Inc. gene cloning. Primers were designed according to the corresponding sequence information on NCBI, and the full length of the GP gene of the two subtypes was obtained by PCR amplification, and the full length of the GP gene was 2031bp ( figure 1 ), the genes with the correct size were recovered and then sequenced.

[0178] 1.2 Construction of Ebola DNA vaccine

[0179]Because there is a signal peptide region of 1-32aa on the GP protein, ...

Embodiment 2

[0181] Example 2 Establishment of Mammalian Biotinylated GP Protein Screening System

[0182] 2.1 Construction of biotinylated GP protein particles

[0183] Because there is a 1-32aa signal peptide (SP) region on the GP protein, and the pIRES-BirA-FLAG vector (purchased from Clonetech) used for biotinylation also has its own IgE secretion signal peptide, it cannot be guaranteed whether the two exist at the same time. It will affect the expression of GP protein, so pIRES-Zaire / Sudan GP-BirA-FLAG SP- and pIRES-Zaire / Sudan GP-BirA-FLAG clones were constructed at the same time. Using the GP gene obtained in 1.1, design primers, add restriction sites for the GP gene, and obtain a target band with a size of about 2000bp ( image 3 ),Recycle.

[0184] At the same time, the GP gene and the pIRES-BirA-FLAG vector were double-digested, ligated, transformed, and positive clones were sequenced after recovery. Successfully obtained pIRES-Zaire / Sudan GP-BirA-FLAG SP- and pIRES-Zaire / Suda...

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Abstract

The invention discloses a neutralizing monoclonal antibody against Ebola virus. Specifically, the invention discloses a monoclonal antibody against Ebola virus. The antibody has significant binding activity to Ebola virus antigen, and Has cross-neutralizing activity against Zaire and Sudan subtypes.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular, the invention relates to a neutralizing monoclonal antibody against Ebola virus. Background technique [0002] On March 23, 2014, the World Health Organization (WHO) reported a new outbreak of Ebola virus (EBOV). The Ebola outbreak first started in Guinea in December 2013 and spread to other West African countries shortly thereafter, including Nigeria, Sierra Leone, Liberia and Senegal. Since then, with the flow of people, the epidemic has been brought to the United States, Spain, Mali and other countries. A small outbreak of Ebola virus infection began in Guinea in February 2014, and was reported in Liberia in April, before spreading to Sierra Leone. Soon, in July 2014, Nigeria reported the first death case of Ebola infection. In August 2014, it was introduced to Senegal. In September and October 2014, patients infected with Ebola virus were found in the United States and Spain resp...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10C12N15/13G01N33/68G01N33/569A61K39/395
CPCC07K16/10C07K2317/52C07K2317/56C07K2317/565C07K2317/64G01N33/56983G01N33/6854G01N2469/10
Inventor 孙兵赵磊凌志洋伊春艳
Owner INST PASTEUR OF SHANGHAI CHINESE ACADEMY OF SCI
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