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Method for producing tabling monoclonal antibody of anti CD25

A monoclonal antibody and expression vector technology, applied in the field of genetic engineering, can solve the problem that the immunosuppressant does not achieve a very satisfactory effect, etc.

Inactive Publication Date: 2007-12-26
SHANGHAI NEWSUMMIT BIOPHARMA +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the immunosuppressants used so far in clinical practice have not achieved very satisfactory results.
Although the one-year survival rate of transplanted kidneys reaches 80-90%, the incidence of acute rejection is still as high as 50%

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] High expression engineered cells obtained

[0043] COS-7 cells (ATCC No.CRL1651) (5000 cells / ml) were co-transfected with the obtained recombinant expression vector encoding the heavy and light chain of chimeric monoclonal antibody against CD25 with FuGENE6 (Boehringer Manheim) reagent, and cultured for three days , the secreted antibody in the supernatant was detected by ELISA. Obtain high-expressing cell lines.

Embodiment 2

[0045] Choose the best medium

[0046] One serum-free cell line recovered from the seed cell bank was inoculated into a T25 square bottle for cultivation. After 48 hours, the cells were inoculated into three T25 square bottles at a ratio of 1:3, and then the cells could be inoculated at a ratio of 1:3. Carry out expansion culture.

[0047] Media name

[0048] Cells were inoculated into different media at the same density, and the number and expression of cells were investigated after 48 hours. The experimental data showed that the cell growth rate and product expression of the cells reached the highest in the B+E+P medium. Therefore, B+E+P medium was selected as the expansion medium for the cells.

Embodiment 3

[0050] Study on High Density Serum-free Suspension Culture of Engineering Cells

[0051] We investigated the adaptation process of engineered cells expressing chimeric monoclonal antibody against CD25 in BDCHO, BD+excel620+primitone, excell 620+DOMA+primitone and Excell 620 medium.

[0052] culture medium

[0053] In summary, BD+excel 620+primitone medium is planned to be used as the serum-free medium for cell expansion, and the cells are expanded and cultured in a 250ml spinner to obtain a certain amount of culture supernatant, which will be purified for purification process research , and obtain a certain amount of pure product to provide quality control for structural confirmation.

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PUM

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Abstract

This invention provides a method for producing anti-CD25 chimeric monoclonal antibody, and corresponding expression vector and host cells. The method has such advantages as high expression level, and simple separation and purification. This invention also provides corresponding expression vector and engineering cells.

Description

technical field [0001] The present invention relates to the field of genetic engineering, and more specifically relates to a method for producing an anti-CD25 chimeric monoclonal antibody, as well as an expression vector and a host cell used in the method. Background technique [0002] Acute rejection after organ transplantation is common, which may cause necrosis of the transplanted kidney or decline in the function of the transplanted kidney. Therefore, drugs that reduce and attenuate these reactions are critical to the success of organ transplantation. Clinically, immunosuppressants are generally used to treat organ transplant patients to inhibit the occurrence of transplant rejection or reduce the degree of reaction. Therefore, kidney transplant recipients have to take immunosuppressive drugs for life to achieve: 1. Reasonable application to prevent and treat rejection (protect the alien kidney); 2. Maintain the patient's moderate immune resistance (protect the patient ...

Claims

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Application Information

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IPC IPC(8): C12P21/08
Inventor 黄阳滨张翊孙九如任军游磊陈子珺徐晓燕刘勇张甘良
Owner SHANGHAI NEWSUMMIT BIOPHARMA
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