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Mouse anti-human IgG monoclonal antibody

A monoclonal antibody, a variable technology, applied in the field of cellular immunology and genetic engineering, which can solve the problems of high background, unclear immune background, false positives, etc.

Active Publication Date: 2020-12-01
南京妙迪生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Human IgG antibodies are composed of heavy chains and light chains (kappa or lambda). After immunization of rabbits, the animal immune system can be stimulated to produce antibodies against the heavy chains and light chains of human IgG antibodies, while the IgM, IgA, IgE and other antibodies in human serum The light chain is the same as the light chain of IgG, that is, kappa or lambda, so when indirect ELISA is used to detect a certain coated antigen-specific human IgG, due to the impurity of the secondary antibody (containing both heavy and light chains Antibodies) will cause false positive results or high background
Second, because the immune background of the animal (such as rabbit) for which the secondary antibody was prepared is not very clear, the rabbit itself has antibodies against the coated antigen (caused by recessive infection). Antibodies to the coated antigen can directly bind to the coated antigen, resulting in false positive results or high background

Method used

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  • Mouse anti-human IgG monoclonal antibody
  • Mouse anti-human IgG monoclonal antibody
  • Mouse anti-human IgG monoclonal antibody

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0065] The preparation of embodiment 1 mouse anti-human IgG monoclonal antibody

[0066] 1. Immunization of Balb / c mice

[0067] The human IgG antibody obtained after human serum was purified by Protein G affinity chromatography column was dialyzed into PBS, and the concentration was measured. Human IgG was used to immunize 5-week-old female Balb / c mice (100 μg / mouse). In the first immunization, 100 μg antigen was mixed with an equal volume of Freund’s complete adjuvant and intraperitoneally injected; after the third week, an equal amount of antigen was mixed with Freund’s incomplete adjuvant. Intraperitoneal immunization after mixing the doses; the third immunization in the 5th week without adjuvant.

[0068] 2. Fusion of splenocytes and myeloma cells

[0069] One week before fusion, resuscitate mouse myeloma cells sp2 / 0 into OPTI-MEM medium (containing 10% fetal bovine serum) and place at 37°C, 5% CO 2 The cells were cultured in an incubator, and the cells were passaged o...

Embodiment 2

[0105] The application of embodiment 2 monoclonal antibody

[0106] 1. Detection of dengue virus (DENV) IgG antibody in humans

[0107] 1. After 1 μl of convalescent serum (14 parts in total) of type II dengue virus infection was mixed with 100 μl of sample diluent, it was added to a microtiter plate (coated with recombinant dengue virus serum type II NS1 antigen, 100ng / well), Set positive and negative controls at the same time, seal the microplate plate with a sealing film, and incubate in a 37°C incubator for 30 minutes.

[0108] 2. Take out the enzyme-labeled plate and place it on the plate washer, wash it with washing solution for 5 times, then shake off the residual liquid in the well or flip the enzyme-labeled reaction plate upside down and pat the residual liquid in the well on absorbent paper.

[0109] 3. Add 100 μl / well of 0070-HRP working solution to each well, and incubate in a 37°C incubator for 30 minutes.

[0110] 4. Repeat step 2.

[0111] 5. Mix the chromoge...

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Abstract

The invention discloses a mouse anti-human IgG monoclonal antibody. The monoclonal antibody has amino acid sequences of heavy chain variable regions CDR1, CDR2 and CDR3 as shown in SEQ ID NO.26, 28 and 30, and amino acid sequences of light chain variable regions CDR1, CDR2 and CDR3 as shown in SEQ ID NO.42, 44 and 46. The invention also provides a nucleic acid molecule for encoding the monoclonalantibody, an expression vector containing the nucleic acid molecule and a host cell. The monoclonal antibody disclosed by the invention has relatively high sensitivity and specificity when being applied to immunodetection.

Description

technical field [0001] The invention belongs to the fields of cellular immunology and genetic engineering, and relates to a mouse anti-human IgG monoclonal antibody. Background technique [0002] Enzyme Linked Immunosorbent Assay (ELISA) is a commonly used immunoenzyme technique. The main method is to adsorb the known antigen or antibody on the surface of the solid phase carrier, then incubate with the enzyme-labeled antibody or antigen, add a chromogenic agent to develop the color, and measure the difference between the color of the test object and the color of the standard object by a microplate reader. difference to obtain qualitative or quantitative results. According to different analytes (detection antigen or detection antibody) and different determination principles, ELISA can be divided into many types, such as direct method, indirect method, competition method, sandwich method, etc. [0003] Indirect ELISA is an effective method for detecting unknown antibodies wi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K16/42C12N15/13G01N33/68G01N33/577G01N33/569
CPCC07K16/4283C07K2317/56C07K2317/565C07K2317/567G01N33/56983G01N33/577G01N33/6854G01N2333/165G01N2333/185G01N2469/20
Inventor 姬艺洪
Owner 南京妙迪生物科技有限公司
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