Recombinant rat anti-mouse CD4 monoclonal antibody, preparation method and application

A monoclonal antibody, mouse technology, applied in the fields of botanical equipment and methods, biochemical equipment and methods, applications, etc., can solve the problems of lack of mouse monoclonal antibody, species limitation, large batch difference, etc. High bit recognition, low production cost, and small batch-to-batch variation of antibodies

Active Publication Date: 2020-11-13
WUHAN CLOUD CLONE CORP
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  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although mouse models are widely used at present, the lack of monoclonal antibodies against mice has caused great difficulties in research
In order to solve this problem, the present invention uses genetic engineering antibody technology to prepare recombinant rat anti-mouse CD4 monoclonal antibody, which solves the problems of species limitation, low yield and large batch difference of traditional monoclonal antibody index technology

Method used

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  • Recombinant rat anti-mouse CD4 monoclonal antibody, preparation method and application
  • Recombinant rat anti-mouse CD4 monoclonal antibody, preparation method and application
  • Recombinant rat anti-mouse CD4 monoclonal antibody, preparation method and application

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Experimental program
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Embodiment 1

[0056] Embodiment 1 Preparation of recombinant rat anti-mouse CD4 monoclonal antibody

[0057] Figure 16 and 17 Flowchart for the preparation of recombinant rat anti-mouse CD4 monoclonal antibody.

[0058] 1. Recombinant rat anti-mouse CD4 monoclonal antibody variable region gene fishing

[0059] 1. Hybridoma RNA extraction and reverse transcription results

[0060] Rats were first immunized with mouse CD4 antigen to obtain rat spleen cells, which were fused with SP20 to obtain hybridoma cells. RNA was extracted from hybridoma cells by Trizol method, and clear 28S and 18S bands were seen by agarose gel electrophoresis , indicating that the integrity of the RNA is good. The RNA concentration and purity measurement results are D(260nm) / D(280nm)=1.85, which can meet the requirements of this experiment. Using RNA as a template to reverse transcribe and synthesize cDNA, using cDNA as a template and the rat internal reference gene β-actin as a primer for PCR amplification, the...

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Abstract

The invention discloses a recombinant rat anti-mouse CD4 monoclonal antibody, its preparation method and application, comprising a heavy chain constant region, a heavy chain variable region, a light chain constant region and a light chain variable region, wherein the heavy chain is variable The region contains CDR1, CDR2, and CDR3, and the coding gene sequence is as shown in SEQ ID NO.7~9, and the translated amino acid sequence is shown in sequence as SEQ ID NO.1~3. The light chain variable region contains CDR1, CDR2, and CDR3, and the coding gene The sequences are shown in SEQ ID NO.10, 11, and 12 in sequence, and the translated amino acid sequences are shown in SEQ ID NO.4, 5, and 6 in sequence. The recombinant monoclonal antibody has clear structure, high potency and better stability.

Description

technical field [0001] The invention relates to the technical field of preparation of monoclonal antibodies in bioengineering, in particular to a recombinant rat anti-mouse CD4 monoclonal antibody, its preparation method and application. Background technique [0002] Antibodies are the workhorse of the biological sciences, but their reliability has repeatedly put researchers in a quandary. Nowadays, the most common problems encountered by researchers are: the purchased antibody for detecting protein X preferentially binds to protein Y (maybe even not binding to X at all); the reproducibility is poor, repeat previous experiments with new antibodies, and find that Results cannot be repeated. A good project was stranded because of this, and the unreliability of antibodies is very alarming. Different results of the same antibody in several parallel experiments will produce disastrous results, which may be related to the production process of the antibody. Insufficient specifi...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/28C12N15/13G01N33/68G01N33/577
CPCC07K16/2812C07K2317/35C07K2317/56C07K2317/565G01N33/577G01N33/68G01N2333/70514
Inventor 杨娟姚新欣何峰容李华渊刘勇李庆严小丽姚雯
Owner WUHAN CLOUD CLONE CORP
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