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Universal Ebola virus disease immunoglobulin as well as preparation method and application thereof

An Ebola virus, protein technology, applied in the direction of antiviral immunoglobulin, immunoglobulin, serum immunoglobulin, etc.

Active Publication Date: 2020-08-11
ACAD OF MILITARY SCI PLA CHINA ACAD OF MILITARY MEDICAL SCI INST OF MILITARY VETERINARY MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Currently, there is no effective preventive and curative drug for the disease

Method used

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preparation example Construction

[0044] The invention provides a safe, efficient and reliable refined universal Ebola virus disease immunoglobulin and a preparation method thereof, in particular a complete set of immunogens for producing refined universal Ebola virus disease immunoglobulin.

[0045] The set of immunogens provided by the present invention for refining general-purpose Ebola virus disease immunoglobulins consists of Ebola virus virus-like particle vaccine antigens and / or vaccinia virus recombinant live vector vaccine antigens.

[0046] First, analyze the Zaire, Sudan and Côte d'Ivoire Ebola virus genomes and antigenic epitopes, design and synthesize Ebola virus GP and VP40 genes and optimize cell-preferred codons; secondly, use insect-bad Virus expression system and / or vaccinia virus recombinant live vector vaccine system to express, purify and prepare chimeric Ebola virus-like particle vaccine antigen; again, immunize healthy animals, obtain hyperimmune plasma, obtain high-purity anti- Ebola vi...

Embodiment 1

[0062] Embodiment 1 horse anti-Ebola virus immunoglobulin F (ab') 2 and its preparation method

[0063] Equine Anti-Ebola Virus Immunoglobulin F(ab’) 2 The preparation method comprises the following steps:

[0064] 1. Design and synthesis of Ebola virus GP and VP40: download the Ebola virus sequence from NCBI, and use DNAMAN and other bioinformatics software to analyze biological information such as gene homology, antigenic sites, and preferred codons; Structural characteristics of Ebola virus, optimize the GP and VP40 gene sequences of Ebola virus, so that they can fully cover the main epitopes of Ebola virus of Zaire type, Sudan type, and Côte d'Ivoire type, and optimize the cell preferred codons according to the expression system, The GP nucleotide sequence is shown in SEQ ID NO:1, and the VP40 nucleotide sequence is shown in SEQ ID NO:2.

[0065] 2. Preparation of VLP protein immunogen:

[0066] The two genes GP and VP40 of Ebola virus were cloned on the same insect ba...

Embodiment 2

[0070] Embodiment 2 Ebola virus disease purified immunoglobulin and preparation method thereof

[0071] The preparation method of Ebola virus disease refined immunoglobulin comprises the following steps:

[0072] 1. Design and synthesis of Ebola virus GP and VP40: download the Ebola virus sequence from NCBI, and use DNAMAN and other bioinformatics software to analyze biological information such as gene homology, antigenic sites, and preferred codons; Structural characteristics of Ebola virus, optimize the GP and VP40 gene sequences of Ebola virus, so that they can fully cover the main epitopes of Ebola virus of Zaire type, Sudan type, and Côte d'Ivoire type, and optimize the cell preferred codons according to the expression system, The nucleotide sequence of GP is shown in SEQ ID NO:1, and the nucleotide sequence of VP40 is shown in SEQ ID NO:2.

[0073] 2. Preparation of immunogen:

[0074] The transfer plasmid was constructed based on the upper (L) and lower (R) homology a...

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Abstract

The invention provides universal Ebola virus disease immunoglobulin as well as a preparation method and application thereof. The method comprises the following steps of firstly, analyzing Zaire type,Sudan type and CotedIvoire type Ebola virus genomes, and optimizing Ebola virus GP and VP40 genetic sequences to form chimeric virus particle type sequences, so that the chimeric virus particle type sequences can completely cover main epitopes of viruses III; then, expressing, purifying and preparing Ebola chimeric virus-like particle vaccine antigens by using an insect-rhabdovirus expression system and a vaccinia virus recombinant living vector vaccine system; and finally, immunizing an animal to obtain high-immunity plasma, carrying out pepsin digestion, ammonium sulfate precipitation and chromatographic column purification to obtain the high-purity anti-Ebola virus disease immunoglobulin F(ab')2, and carrying out safety and effectiveness evaluation on the high-purity anti-Ebola virus disease immunoglobulin. The universal Ebola virus disease immunoglobulin provided by the invention can prevent and treat hemorrhagic fever caused by Ebola virus III infection, and provides a powerful tool for Ebola epidemic prevention and control.

Description

technical field [0001] The invention relates to the field of biopharmaceuticals, in particular to a general Ebola virus disease immunoglobulin and its preparation method and application. Background technique [0002] Ebola hemorrhagic fever is caused by Ebola virus (EBOV), and the case fatality rate can be as high as 90%. It belongs to the biosafety level IV virus. Since the disease was discovered in the Ebola River Basin in Africa in 1976, it has been distributed every year. WHO has confirmed it as a global public health emergency and called for the global strengthening of drug research and development. [0003] EBOV includes five subtypes, Ebola-Zaire (Z-EBOV), Ebola-Sudan (S-EBOV), Ebola-Coted'Ivoire , C-EBOV), Ebola-Reston (R-EBOV) and Ebola-bundibugyo (B-EBOLA). Among them, Z-EBOV is the most virulent, and the mortality rate after human infection is the highest; S-EBOV is second; C-EBOV is pathogenic to chimpanzees, but the virulence to humans is the weakest; R and B-...

Claims

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Application Information

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IPC IPC(8): C12N15/40C07K14/08C12N15/866C12N15/863A61K39/12A61K39/42A61P31/14C07K16/10C07K16/06
CPCA61K39/12A61K2039/505A61K2039/5256A61K2039/5258A61P31/14C07K14/005C07K16/06C07K16/10C07K2317/33C07K2317/54C07K2317/76C07K2317/94C12N15/86C12N2710/14043C12N2710/24043C12N2710/24243C12N2760/14022C12N2760/14034C12N2760/14052C12N2800/105C12N2800/30
Inventor 赵永坤赵忠鹏张颖杨松涛高玉伟闫飞虎郑学星王化磊李敏杨晓岚夏咸柱
Owner ACAD OF MILITARY SCI PLA CHINA ACAD OF MILITARY MEDICAL SCI INST OF MILITARY VETERINARY MEDICINE
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