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Heterologous antibody of novel coronavirus (2019-nCOV) and preparation method thereof

A 2019-ncov, virus technology, applied in the direction of antibodies, antiviral agents, antiviral immunoglobulins, etc., can solve the problems of large clinical side effects, high content of impurity proteins, and low purity

Inactive Publication Date: 2020-07-31
TANGSHAN YIAN BIOLOGICAL ENG CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The research and development of specific antiviral drugs and vaccines has great uncertainty and long-term nature. The re-development of heterologous serum antibodies has its more urgent needs. Horse serum prepared by traditional antiserum production processes still exists. IgG or Antibody F(ab') 2 Low purity, high impurity protein content, low neutralizing antibody titer, large clinical side effects, etc.

Method used

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  • Heterologous antibody of novel coronavirus (2019-nCOV) and preparation method thereof
  • Heterologous antibody of novel coronavirus (2019-nCOV) and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Embodiment 1: Preparation of 2019-nCOV (SARS-CoV-2) virus S protein immunogen

[0030] Follow the steps below to obtain the S protein immunogen

[0031] (1) Protein expression and inspection: the 293T-2019-nCOV (SARS-CoV-2)-S engineered cell line capable of stably expressing the 2019-nCOV (SARS-CoV-2) S protein was tested in a paper carrier bioreactor cultured at 37°C, 60% DO, 160rpm, and detected the concentration and size of the protein in the culture medium by SDS-PAGE, which was consistent with the expected fragment size of 45kd, see figure 1 .

[0032] (2) Protein harvest and purification: Harvest the culture medium with peak expression of S protein for 3-7 days, filter through 0.65 μm to remove cell debris, concentrate 100 times with a 10kd ultrafiltration concentration system, and add β-propiolactone at a ratio of 1:4000 Inactivate for 24 hours, stirring and mixing several times during this period. After inactivation, hydrolyze at 37°C for 2 hours, and store a...

Embodiment 2

[0034] Example 2: Preparation and collection of horse anti-2019-nCOV (SARS-CoV-2) virus hyperimmune serum

[0035] Quarantine was carried out in accordance with the relevant standards for immune donors. Four 4-5-year-old healthy antibody-negative brown horses were selected for primary vaccination and multiple booster immunizations by subcutaneous injection on the back and intraperitoneal injection respectively. A total of 3 Each time, with an interval of 7-14 days, the dose of immunogen was 1mg, 2mg, 3mg / horse in sequence. Before each immunization and 14 days after the last immunization, blood was collected venously, and the serum was separated, and the antibody level was detected by the agar expansion method.

Embodiment 3

[0036] Example 3: Treatment and potency determination of horse anti-2019-nCOV (SARS-CoV-2) virus hyperimmune serum

[0037] 1. Hyperimmune serum inactivation and pyrogen removal

[0038] Under sterile conditions, inactivate the collected hyperimmune serum at 56°C for 30 minutes, adjust the pH to 4.0, add 10% aluminum potassium sulfate according to 0.5% of the total volume, then adjust the pH to 7.0 with NaOH, stir and adsorb for 1 hour, 2-8 overnight at ℃, centrifuge to take the supernatant, add aseptically treated activated carbon to the supernatant, stir and absorb for 1 hour, and filter for later use.

[0039] 2. Determination of hyperimmune serum titer

[0040] Prepare an agar plate with agarose gel prepared with 8.0% NaCl and 1% agarose purified water, punch 7 plum blossom holes, add 2019-nCOV (SARS-CoV-2) agar amplification antigen in the middle hole, and add in sequence to the surrounding 6 holes 1:2, 1:4, 1:8, 1:16, 1:32 diluted horse serum to be tested, add 0.01mol ...

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Abstract

The present invention provides a non-human animal-derived antiserum and / or non-human animal-derived immunoglobulin against the 2019-nCOV (SARS-CoV-2). The invention is intended to be able to be used in the treatment and prevention of novel coronavirus pneumonia.

Description

【Technical field】 [0001] The present invention relates to an antibody and a method for preparing the antibody, and further relates to a heterologous antibody to the novel coronavirus (2019-nCOV) and a method for preparing the same. 【Background technique】 [0002] 2019-nCOV (SARS-CoV-2) is another new type of coronavirus that causes Novel Coronavirus Pneumonia (Novelcoronavirus pneumonia NCP). Some patients with this type of pneumonia will get worse after 1 week, and may progress to the following Life-threatening conditions, such as acute respiratory distress syndrome, septic shock, metabolic acidosis, coagulation dysfunction, renal failure or other systemic organ failure, etc. At present, no specific therapeutic drugs and vaccines have been successfully developed. The special immune plasma products of convalescent patients are currently the most effective method for the treatment of new coronavirus infection. However, this is a last resort emergency treatment method, and so...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/10A61K39/395A61P31/14
CPCC07K16/10A61P31/14A61K2039/505
Inventor 杨培豫李润耿纯梅张洪军张玉新张子佳王晓辉谷孝玉李艳玲黄韬汪新明裴亚芳
Owner TANGSHAN YIAN BIOLOGICAL ENG CO LTD
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