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A kind of monoclonal antibody zk2c2 and its application

A technology of antibodies and sequence lists, applied in the direction of antibodies, applications, antiviral agents, etc., can solve problems such as increasing virus entry, aggravating the disease, and aggravating the severity of the disease, achieving significant application value and far-reaching social significance

Active Publication Date: 2021-02-09
GUANGZHOU EIGHTH PEOPLES HOSPITAL +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Studies have shown that people who have been infected with different serotypes of dengue virus successively have the risk of aggravating the severity of the disease, that is, the antibodies produced by the pre-infected serotype of dengue virus can bind to the different serotypes of dengue virus reinfected, but cannot neutralize Viruses, on the contrary, increase the entry of viruses and aggravate the condition

Method used

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  • A kind of monoclonal antibody zk2c2 and its application
  • A kind of monoclonal antibody zk2c2 and its application
  • A kind of monoclonal antibody zk2c2 and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Example 1. Discovery of Antibodies

[0040] 1. Preparation of protein

[0041] 1. Construction of recombinant plasmids

[0042] (1) Synthesize the double-stranded DNA molecule shown in SEQ ID NO: 2 of the Sequence Listing.

[0043] The double-stranded DNA molecule shown in the sequence 2 of the sequence listing encodes the protein shown in the sequence 1 of the sequence listing, wherein the open reading frame is the 13-1296th nucleotide from the 5' end of the sequence 2.

[0044] In Sequence 1 of the sequence listing, the signal peptide is composed of the 1st to 19th amino acid residues from the N-terminal, the 20th to 421st amino acid residues form the E protein of Zika virus, and the 422nd to 427th amino acid residues form His6 Label. The protein shown in Sequence 1 of the Sequence Listing was named E-His 6 Fusion protein with an expected molecular weight of 50kDa.

[0045] (2) The double-stranded DNA molecule obtained in step 1 was double-digested with restrictio...

Embodiment 2

[0073] Example 2. Preparation of ZK2C2 antibody

[0074] 1. Construction of recombinant plasmids

[0075] The double-stranded DNA molecule shown in SEQ ID NO: 4 of the sequence listing was inserted into the PMD18-T vector to obtain a heavy chain expression vector. In Sequence 4 of the Sequence Listing, nucleotides 1 to 888 constitute the CMV promoter, nucleotides 889 to 2280 encode the full-length heavy chain shown in Sequence 3 of the Sequence Listing, and nucleotides 2333 to 2478 is the ployA fragment.

[0076] The double-stranded DNA molecule shown in SEQ ID NO: 6 of the sequence listing was inserted into the PMD18-T vector to obtain a light chain expression vector. In Sequence 6 of the Sequence Listing, nucleotides 1 to 888 constitute the CMV promoter, nucleotides 889 to 1596 encode the full-length light chain shown in Sequence 5 of the Sequence Listing, and nucleotides 1597 to 1742 is the ployA fragment.

[0077] 2. Preparation of Antibodies

[0078] 1. 293T cells we...

Embodiment 3

[0086] Example 3. Neutralizing activity of ZK2C2 antibody to virus

[0087] 1. Preparation of virus

[0088] Zika virus was inoculated into C6 / 36 cells (in DMEM medium containing 10% FBS) and placed at 28°C, 5% CO 2 The cells were cultured under static conditions for 4 days, and then centrifuged at 3000 rpm for 5 min, and the supernatant was collected, which was the Zika virus solution.

[0089] 2. Detection of neutralizing activity of monoclonal antibodies

[0090] 1. Take the ZK2C2 solution prepared in Example 2 and dilute it with PBS buffer (pH 7.2, 10 mM) to obtain an antibody dilution.

[0091] 2. Seed Vero cells in a six-well plate (4 × 10 per well). 5 cells), and cultured overnight, the cell density grew to 90% and evenly covered the well plate.

[0092]3. Mix the Zika virus solution prepared in step 1 with the antibody diluent prepared in step 1 to obtain each mixture (in each milliliter of mixture, the virus content is 100 pfu, and the antibody concentration is 40...

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Abstract

The invention discloses a monoclonal antibody ZK2C2 and its application. The present invention provides an IgG antibody named ZK2C2, which is composed of a light chain and a heavy chain; CDR1, CDR2 and CDR3 in the heavy chain variable region in the heavy chain are sequence 3 in the sequence table in sequence from the N-terminal 45-52 amino acid residues, 70-77 amino acid residues and 116-122 amino acid residues; CDR1, CDR2 and CDR3 in the light chain variable region in the light chain are the sequences in the sequence listing successively 5 From the N-terminal amino acid residues 45-52, amino acid residues 70-72 and amino acid residues 109-119. The present invention also protects the application of the IgG antibody in the preparation of a medicine for inhibiting Zika virus. The present invention has great application value for the prevention and control of Zika virus disease, and will have far-reaching social significance.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a monoclonal antibody ZK2C2 and its application. Background technique [0002] Zika virus, dengue virus, West Nile virus and yellow fever virus belong to the genus Flavivirus. Zika virus disease is a mosquito-borne infectious disease caused by Zika virus with mild or no symptoms. The main manifestations are mild fever, rash (mostly maculopapular rash), headache, arthralgia, muscle pain, weakness, and non-purulent conjunctivitis. In recent years, Zika virus broke out in French Polynesia and Brazil, and continued to spread rapidly around the world, especially in Latin America and the Caribbean. It has become an international public health emergency. [0003] Zika virus infection causes only mild symptoms. However, in the epidemic of Polynesia, adults have serious neurological complications - Guillain-Barré syndrome (GBS), and its incidence is as high as 1 / 4000. More int...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/10C12N15/13A61K39/42A61P31/14
CPCA61K2039/505C07K16/1081C07K2317/565C07K2317/76Y02A50/30
Inventor 张复春庾蕾张林琦王若珂高菲
Owner GUANGZHOU EIGHTH PEOPLES HOSPITAL