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Anti-human adenovirus type 7 antibody 2-1H and application thereof

A monoclonal antibody, sequence technology, used in antiviral agents, antiviral immunoglobulins, applications, etc.

Active Publication Date: 2019-07-02
ACADEMY OF MILITARY MEDICAL SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, there is no specific therapeutic drug against adenovirus for clinical use in the world, and the research and development of drugs is mostly limited to chemical drugs—nucleoside analogs against DNA viruses, while the research on specific biological drugs for adenovirus is still blank

Method used

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  • Anti-human adenovirus type 7 antibody 2-1H and application thereof
  • Anti-human adenovirus type 7 antibody 2-1H and application thereof
  • Anti-human adenovirus type 7 antibody 2-1H and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Embodiment 1, the discovery of antibody

[0070] The peripheral blood of convalescent human adenovirus type 7 infected patients was collected, and peripheral blood mononuclear lymphocytes were separated and collected. Collected peripheral blood mononuclear lymphocytes were added with anti-cell surface marker antibodies (BD, #555332, #555415, #555441, #560677, #555622) or control antibodies (BD, #555748, #555742, #555751, # 555749, #557872) were labeled and flow sorted, and the sorted cells were used for antibody gene amplification. Using the isolated single B cell as a template, the antibody gene was amplified, and the amplified positive clone was sequenced and expressed, and the purified sample of adenovirus type 7 was used as the antigen. After a large number of screening, analysis, and verification, an antibody sequence was obtained, named as 2-1H antibody.

[0071] The amino acid sequence of the heavy chain variable region of the 2-1H antibody is shown in Sequence...

Embodiment 2、2-1

[0073] The preparation of embodiment 2, 2-1H antibody

[0074] 1. Construction of recombinant plasmids

[0075] 1. Replace the small fragment between the SalI and PmlI sites of the pTSE-G1n vector (Beijing Baite Meibo Biological Co., Ltd.) with the DNA molecule shown in Sequence 1 of the sequence listing to obtain a recombinant expression vector containing the heavy chain variable region (Already sequenced and verified).

[0076] 2. Replace the small fragment between the SalI and PmlI sites of the pTSE-K vector (Beijing Baite Meibo Biological Co., Ltd.) with the DNA molecule shown in sequence 3 of the sequence listing to obtain a recombinant expression vector containing the light chain variable region (Already sequenced and verified).

[0077] 2. Preparation of 2-1H antibody

[0078] 1. The day before transfection, FreeStyle TM HEK 293-F cells (Invitrogen, product number: R79007) were adjusted to a concentration of 1.0×10 6 / ml, inoculated into culture flasks, 37°C, 5% C...

Embodiment 3、2-1

[0083] Example 3, the detection of binding ability of 2-1H antibody

[0084] 1. Preparation of human adenovirus type 7 stock solution, concentrated solution and inactivated virus

[0085] 1. Preparation of adenovirus liquid

[0086] Adenovirus culture: conventional DMEM+10% (volume percentage) FBS medium to culture A549 cells (Beijing Concorde Cell Resource Center, article number: 25), and pass A549 cells to 75cm one day before virus inoculation. 2 In the cell flask, make the cell density reach 75% to 90% when the virus is inoculated the next day; on the day of inoculation, slowly suck out the cell culture medium in the culture flask, add 5ml DMEM to gently wash the cells and discard, and then add 3ml DMEM+2 % (volume percentage content) FBS; use a micropipette to draw an appropriate amount of HAdV7 virus into the cell bottle, and infect according to MOI ≈ 0.001, shake the bottle evenly several times to make the virus evenly dispersed, and place it at 37 ° C, 5% CO 2 Adsorb ...

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Abstract

The invention discloses an anti-human adenovirus type 7 antibody 2-1H and an application thereof. The invention provides a monoclonal antibody, and the monoclonal antibody comprises a heavy chain variable region and a light chain variable region; the heavy chain variable region comprises three complementary determining regions HCDR1, HCDR2 and HCDR3; the light chain variable region comprises threecomplementary determining regions LCDR1, LCDR2 and LCDR3; the HCDR1, the HCDR2 and the HCDR3 are sequentially shown as a 26th-33th position, a 51th-58th position and a 97th-117th position of a sequence 2 of a sequence table from an N end; and the LCDR1, the LCDR2 and the LCDR3 are sequentially shown as a 27th-32th position, a 50th-52th position and a 89th-97th position of a sequence 4 of the sequence table from the N end. Based on clinical needs, the anti-human adenovirus type 7 antibody 2-1H is found, is used for prevention and treatment of adenovirus infection, and has important biologicaland medical significance.

Description

technical field [0001] The invention relates to an anti-human type 7 adenovirus antibody 2-1H and its application. Background technique [0002] Human adenovirus belongs to the Adenoviridae mammalian adenovirus genus. Human adenovirus was first discovered in 1953. It was isolated and cultured from the atrophic tonsil adenoid tissue of healthy people; the genome of the virus is double-stranded DNA with a total length of about 36kb. These double-stranded DNA and viral structural proteins combine to form the core of the virus. The virus has no envelope, and the core is covered with a capsid. The capsid is composed of 252 capsomers, of which 240 are hexon proteins, 12 penton proteins, and the shell It is a regular 20-hedron structure with a diameter of about 70-90nm. [0003] There are 7 subgroups of adenoviruses found so far, A-G, and 67 different serotypes, of which 55 subtypes can infect humans and cause disease. The most common adenovirus infection is to infect the respira...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/08C12N15/13A61K39/42A61P31/20
CPCA61K2039/505A61P31/20C07K16/081C07K2317/565
Inventor 杨志新王荣陆健昇余云舟周权黄莺周晓巍
Owner ACADEMY OF MILITARY MEDICAL SCI
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