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Anti-monocytic leukemia-associated antigen mla-34 fully human monoclonal single-chain antibody scfv

A mononuclear cell and single-chain antibody technology, applied in the direction of anti-receptor/cell surface antigen/cell surface determinant immunoglobulin, anti-animal/human immunoglobulin, immunoglobulin, etc., can solve the problem of non-panning And identified problems such as fully human monoclonal antibody, to achieve the effect of small molecular weight, strong penetrating power, and favorable application

Active Publication Date: 2019-05-14
THE SECOND AFFILIATED HOSPITAL OF XIAN JIAOTONG UNIV
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Problems solved by technology

MLAA-34 is a new leukemia anti-apoptotic gene expressed on the surface of the cell membrane and related to the onset and prognosis of M5, so it can be a target for monoclonal antibody therapy, but so far in the prior art, there is no such gene in the whole human source. Panned and identified anti-MLAA-34 fully human monoclonal antibody in phage antibody library

Method used

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  • Anti-monocytic leukemia-associated antigen mla-34 fully human monoclonal single-chain antibody scfv
  • Anti-monocytic leukemia-associated antigen mla-34 fully human monoclonal single-chain antibody scfv
  • Anti-monocytic leukemia-associated antigen mla-34 fully human monoclonal single-chain antibody scfv

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Embodiment 1

[0059] 1.1.1 Construction of prokaryotic expression vector

[0060] The total RNA in U937 cells was extracted, reverse-transcribed into cDNA, using cDNA as a template strand, primers were designed, and the coding region of MLAA-34 was amplified by PCR. The target carrier is digested, and the digested products are recovered by electrophoresis and exchanged, and the products are transformed into bacterial competent cells. Colony PCR identification was performed on the grown clones first, and then sequencing and comparative analysis were performed on the positive clones identified by PCR. The correct comparison was the successful construction of the target plasmid ( Figure 3-1 ). figure 1 Flowchart for the construction of the prokaryotic expression vector for the target gene, figure 2 A map of the expression vector.

[0061] 1) Enzyme digestion of the vector.

[0062] Enzyme digestion reaction temperature: 37°C, enzyme digestion reaction time: 2h,

[0063] Table 1 enzyme d...

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Abstract

The invention provides a monocytic leukemia associated antigen MLAA-34 resistant fully human monoclonal single-chain antibody ScFv. The monoclonal single-chain antibody ScFv of special anti-MLAA-34 is obtained by screening and identifying in a phage antibody library; MLAA-34 protein is firstly expressed and purified, and antigen protein is provided for monoclonal antibody preparation. A method beneficial to genetic engineering is used for screening and identifying anti-MLAA-34 fully human monoclonal antibody. The fully human monoclonal single-chain antibody ScFv for specially combining and efficiently expressing tumor cell strains of MLAA-34 is obtained. The fully human monoclonal single-chain antibody ScFv can be used for solving the immunogenicity problem of a mouse original antibody, and the single-chain antibody ScFv is formed by connecting a heavy chain and a light chain of an antibody variable region, is small in molecular weight, is high in penetrating power and is more beneficial to application.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to an anti-monocytic leukemia-associated antigen MLAA-34 fully human monoclonal single-chain antibody ScFv. Background technique [0002] Acute myeloid leukemia (AML) has a poor prognosis, especially those with adverse chromosomal or molecular abnormalities, mainly due to the high relapse rate after chemotherapy-induced remission. The eradication of minimal residual disease (MRD) after remission remains a huge challenge. Allogeneic hematopoietic stem cell transplantation (hematopoietic stem cell transplantation, HSCT) shows effective anti-leukemia effect, but because of the difficulty of matching, high cost, and more importantly, some life-threatening complications, such as graft-versus-host disease ( graftversus host disease, GVHD), only suitable for a small number of young and suitable patients. Therefore, there is an urgent need to develop new therapeutic approaches bas...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/30C12N15/13
CPCC07K16/30C07K2317/622
Inventor 张王刚张扬曹星梅陈银霞何爱丽刘捷赵万红马肖荣杨云张鹏宇
Owner THE SECOND AFFILIATED HOSPITAL OF XIAN JIAOTONG UNIV
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