Monoclonal antibody of anti-human BAP31 protein, preparation method and application thereof

A monoclonal antibody and protein technology, applied in anti-animal/human immunoglobulin, botanical equipment and methods, biochemical equipment and methods, etc. Check the effect of the tool

Inactive Publication Date: 2010-03-03
TIANJIN MEDICAL UNIV CANCER HOSPITAL
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

The disadvantage of using this method to prepare monoclonal antibodies is that the workload of preparing antibodies for one fusion is large, and antigen identification is required in the later stage. It is difficult to predict what kind of antibodies will be produced at the be

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  • Monoclonal antibody of anti-human BAP31 protein, preparation method and application thereof
  • Monoclonal antibody of anti-human BAP31 protein, preparation method and application thereof
  • Monoclonal antibody of anti-human BAP31 protein, preparation method and application thereof

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Embodiment Construction

[0027] 1. Preparation of MCF-7 cell monoclonal antibody library

[0028] (1) Material method

[0029] 1. Source of experimental materials

[0030] BALB / c mice, MCF-7 cells and mouse myeloma cells (SP2 / 0) were provided by Tianjin Cancer Hospital. Freund's complete adjuvant and incomplete Freund's adjuvant, HAT, and HT conditioned medium components were purchased from Invitrigen. DMEM medium, polyethylene glycol (PEG), and RPMI 1640 medium were purchased from Hyclone Company. Horseradish peroxidase (HRP)-labeled goat anti-mouse Ig, o-phenylenediamine (OPD), diaminobenzidine (DAB), and 8-azaguanine (8-AG) were purchased from Sigma.

[0031] 2. Animal immunization

[0032] Take 1×10 7 MCF-7 cells were collected by centrifugation, added 200ul normal saline and repeatedly frozen and thawed 5 times, and the supernatant was discarded by centrifugation. Add 300 ul of lysate (0.05M Tris-HCl pH=8.0, 1% Triton X100) to the precipitate, centrifuge at 12,000 rpm at 4°C, and take the ...

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Abstract

The invention discloses a monoclonal antibody of anti-human BAP31 protein, a preparation method and the application thereof, belonging to the preparation of antibody-containing medical preparation or the monoclonal antibody. The monoclonal antibody of the anti-human BAP31 protein is generated by a hybridoma cell with the collection number of CGMCC No. 3271. The breast adenocarcinoma MCF-7 immuno-BALB/c mouse is adopted, spleen and myeloma cell of the mouse are taken to make cell fusion, the MCF-7 cell is utilized for screening, and then a monoclonal antibody library is formed by the obtained positive clones; and the antigen of the monoclonal antibody derived from the antibody library can be identified to be BAP31 by immuno-precipitation and a mass spectrum method. A variable region sequence for identifying the antibody can be determined, and the antibody can distinguish the BAP31 protein in a specific way and has specific binding capacity with the BAP31 protein. The invention is applicable to the detection of immunohistochemistry, immuno-fluorescence, immuno-blot and immuno-precipitation of the molecule.

Description

technical field [0001] The present invention relates to the preparation of pharmaceutical preparations containing antibodies, or monoclonal antibodies, specifically a monoclonal antibody against human BAP31 protein, Background technique [0002] In 1975, Kohler and Milstein discovered that the fusion of mouse myeloma cells and mouse spleen cells immunized with sheep red blood cells could produce antibodies and proliferate indefinitely, thus creating the monoclonal antibody hybridoma technology. [0003] The monoclonal antibody library is a collection of various monoclonal antibodies against different proteins obtained by immunizing mice with mixed antigens such as cells or cell extracts. The disadvantage of using this method to prepare monoclonal antibodies is that the workload of preparing antibodies for one fusion is large, and antigen identification is required in the later stage, and it is difficult to predict what kind of antibodies will be produced at the beginning. T...

Claims

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Application Information

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IPC IPC(8): C07K16/18C12N15/06C12P21/08C12N5/20G01N33/577
Inventor 张霖罗艺牛瑞芳张宁
Owner TIANJIN MEDICAL UNIV CANCER HOSPITAL
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