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Mouse anti-human ZNF580 monoclonal antibody

A monoclonal antibody, mouse anti-human technology, applied in the field of genetic engineering, can solve the problem that the pathogenesis has not been fully elucidated

Inactive Publication Date: 2014-07-09
LOGISTICS UNIV OF CAPF
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, as a complex polygenic disease, AS involves a huge gene regulatory network, and its pathogenesis has not yet been fully elucidated. With the completion of genome sequencing of humans and various model organisms, an important goal in the future is to elucidate The function of about 35,000 genes in the human body

Method used

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  • Mouse anti-human ZNF580 monoclonal antibody
  • Mouse anti-human ZNF580 monoclonal antibody
  • Mouse anti-human ZNF580 monoclonal antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Based on the human ZNF580cDNA, according to the codon preference of Escherichia coli, and while ensuring that the amino acid sequence of the expressed protein remains unchanged, the codon modification was carried out, and the human ZNF580 gene with rare codon modification was obtained, which is SEQ ID Shown in No.1.

Embodiment 2

[0019] 1. Materials and methods

[0020] 1.1 Materials

[0021] PCR Kit, Agarose Gel DNA Extraction Kit, Escherichia coli BL21 (DE3), T 4 DNA ligase and restriction endonuclease were purchased from Dalian Bao Biological Engineering Co., Ltd. The high-purity plasmid mini-extraction kit and enterokinase were purchased from TIANGEN BIOTECH, and the His·band protein purification kit was purchased from Novagen, Germany.

[0022] 1.2 Method

[0023] 1.2.1 Construction of expression plasmids

[0024] According to the characteristics of the expression plasmid pET30a used, the corresponding enterokinase sequence and restriction endonuclease site were designed, and the whole sequence was synthesized by Shanghai Sangon Biotechnology Service Co., Ltd. and connected to the plasmid pET30a, named pET30a-ZNF580. The prokaryotic expression vector pET30a-ZNF580 was successfully constructed.

[0025] 1.2.2 Expression of target protein

[0026] 1.2.2.1 Preparation of Competent Escherichia c...

Embodiment 3

[0056] One, the preparation method of mouse anti-human ZNF580 monoclonal antibody

[0057] 1.1 Preparation of immune B lymphocytes

[0058] Immunization of Balb / c mice and determination of serum antibody titers:

[0059] The purified ZNF580 protein was diluted to 0.5 mg / ml with 0.01 M PBS, pH=7.4.

[0060] Antigen emulsification adopts double syringe mutual pushing method. Six Balb / c mice were taken for immunization, and the immunization plan: the first immunization on 0d, 100μg / mouse (0.2ml) plus an equal volume of Freund’s complete adjuvant, multi-point intradermal injection and intraperitoneal injection; the second immunization on the 14th and 28th days The second and third immunizations: the adjuvant was changed to Freund's incomplete adjuvant, and the amount of antigen, injection volume and route remained unchanged; after the third immunization, the titer was determined by indirect ELISA. Booster immunization was carried out 3 days before fusion, without adjuvant, and ...

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Abstract

The invention discloses a mouse anti-human ZNF580 monoclonal antibody. The mouse anti-human ZNF580 monoclonal antibody is produced by a hybridoma cell having an accession number of CGMCC No. 8608. A rare codon in a human ZNF580 gene is modified and the modified gene is subjected to prokaryotic expression so that a large amount of ZNF580 proteins are produced and thus the problem that based on the prior art, a ZNF580 protein gene is not expressed in a prokaryotic cell is solved, expression and purification of the human ZNF580 protein are successfully realized, and the human ZNF580 protein is used as an antigen for preparation of the mouse anti-human ZNF580 monoclonal antibody.

Description

technical field [0001] The invention belongs to the field of genetic engineering, in particular to a monoclonal antibody. Background technique [0002] Atherosclerosis (AS) is a common disease that seriously endangers human health and is an important pathological basis of ischemic cardiovascular and cerebrovascular diseases. The main features of AS plaques are the accumulation of intracellular and extracellular lipids, infiltration of monocytes / macrophages, formation of foam cells, hyperplasia of aortic smooth muscle, and accumulation of connective tissue components in the lesion. In recent years, with the changes in people's lifestyle and diet structure, the incidence of cardiovascular and cerebrovascular diseases caused by AS is on the rise in my country. AS is a polygenic disease induced by environmental factors and genetic factors, and there is still a lack of very effective treatment measures. Applying molecular biology techniques to a large number of studies on patie...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/18
Inventor 张文成牟心红李海生
Owner LOGISTICS UNIV OF CAPF
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