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Isolated SARS-CoV-2 protein binding molecule against virus infection

A technology that binds molecules, sars-cov-2, and is used in antiviral immunoglobulins, antiviral agents, immunoglobulins, etc.

Inactive Publication Date: 2020-07-17
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

[0004] At present, there is no anti-SARS-CoV-2 antibody on the market at home and abroad. Therefore, the establishment and development of antibody-based detection and diagnosis and treatment supplies with independent intellectual property rights has important practical significance for the intervention of various related diseases.

Method used

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  • Isolated SARS-CoV-2 protein binding molecule against virus infection
  • Isolated SARS-CoV-2 protein binding molecule against virus infection
  • Isolated SARS-CoV-2 protein binding molecule against virus infection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0098] Example 1 Screening of anti-SARS-CoV-2 single-chain antibody

[0099] 1. Virus culture (all operations are carried out in BSL-3 laboratory)

[0100] The BetaCoV / JS03 / human / 2020 strain was isolated by the patent applicant from a throat swab of a COVID-19 patient in Jiangsu Province in 2020. The virus was serially diluted and inoculated into Vero E6 cells in 5% CO 2 , and incubated at 37° C. to determine the 50% tissue culture infectious dose (50% tissue culture infective dose, TCID50).

[0101] 2. Recombinantly expressed SARS-CoV-2 S protein (SARS-CoV-2-S protein), commercially purchased, the sequence is shown in SEQ ID NO:49.

[0102] 3. Construction of ScFv human antibody library and screening of anti-SARS-CoV-2-S protein single-chain antibody

[0103] 1. Materials

[0104] Primers: Family-specific light chain (Vκ and Vλ), IgG heavy chain (VH) and overlap-PCR primers were designed according to the book "Phage Display", including 12 pairs of Vκ, 24 pairs of Vλ, 6 pa...

Embodiment 2

[0159] Detection of neutralizing activity of the single-chain antibody obtained by screening in Example 2

[0160] Microneutralization experiments of 7 scFv monoclonal antibody clones (all operations were performed in BSL-3 laboratory).

[0161] 1. Preparation of phage antibody

[0162] Pick seven single colonies that were positive in the Phage-ELISA test and place them in 2ml 2×YT medium (containing 100 μg / mL ampicillin, 12.5 μg / mL tetracycline, and 1 g / mL glucose), culture them overnight at 37°C with shaking, and the next day 1:10 were inoculated into 10ml of 2×YT medium (containing 100 μg / mL ampicillin and 30 μg / mL tetracycline) and cultured with shaking at 37°C for 6 hours, and the helper phage VCSM13 (final concentration of 1×10 9PFU / mL), incubate at 37°C for 1 h, add kanamycin (final concentration is 50 μg / mL), shake at 30°C overnight, centrifuge at 900 g for 30 minutes, discard the precipitate, add 5×PEG / NaCl to the supernatant, mix After uniformity, place on ice for ...

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Abstract

The invention discloses an isolated SARS-CoV-2 protein binding molecule against virus infection, and the binding molecule is a monoclonal antibody of S protein of SARS-CoV-2. The amino acid sequence of the heavy chain variable region of the binding molecule is selected from SEQ ID NOs:4, 12, and 20, and the amino acid sequence of the light chain variable region is selected from SEQ ID NOs:8, 16 and 24. The binding molecule provided by the invention can specifically bind to the S protein of the SARS-CoV-2 and neutralize the SARS-CoV-2 to exert an antiviral effect.

Description

technical field [0001] The invention belongs to the fields of antiviral therapy and molecular immunology, and relates to an isolated anti-virus infection SARS-CoV-2 protein binding molecule, and the binding molecule is a specific antibody for the protein. Background technique [0002] Corona Virus Disease 2019 (COVID-19), referred to as "New Coronary Pneumonia", is a pneumonia caused by SARS-CoV-2 infection. Respiratory droplet transmission is the main route of transmission. In a relatively closed environment for a long time There is the possibility of aerosol transmission when exposed to high-concentration aerosols, and other transmission routes have yet to be clarified; the population is generally susceptible, and the incubation period is 1 to 14 days, generally 3 to 7 days. The clinical types include asymptomatic infection, mild, common, severe, and critical. The prognosis of the elderly and those with underlying diseases is poor, and the symptoms of children are relative...

Claims

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Application Information

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IPC IPC(8): C07K16/10C12N15/13A61K39/42A61P31/14G01N33/569
CPCA61K2039/505A61P31/14C07K16/10C07K2317/56C07K2317/565G01N33/56983G01N2333/165
Inventor 张文帅郭喜玲焦永军曾晓燕朱凤才朱宝立
Owner JIANGSU PROVINCIAL CENT FOR DISEASE CONTROL & PREVENTION PUBLIC HEALTH RES INST OF JIANGSU PROVINCE