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Purification method of recombinant human vascular endothelial growth factor receptor-antibody fusion protein

A technology of growth factor receptor and vascular endothelium, which is applied in the field of purification of recombinant human vascular endothelial growth factor receptor-antibody fusion protein, which can solve the problems of complex structure and difficult purification work

Active Publication Date: 2015-12-23
QILU PHARMA CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Recombinant human vascular endothelial growth factor receptor-antibody fusion protein is a fusion protein with a relatively complex structure. It has many disulfide bonds and glycosylation sites, and contains many basic charge isomers, which brings difficulties to downstream purification work. come very difficult

Method used

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  • Purification method of recombinant human vascular endothelial growth factor receptor-antibody fusion protein

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Embodiment 1

[0053] A method for purifying recombinant human vascular endothelial growth factor receptor-antibody fusion protein, the specific steps of which are:

[0054] Affinity chromatography

[0055] (1) Equilibrate the rProteinAFastFlow affinity chromatography column (5.0×20cm) with 1000mL disodium hydrogen phosphate-sodium dihydrogen phosphate equilibrium buffer containing 1mol / L NaCl at a concentration of 10mmol / L and a pH value of 7.5, and the recombinant The cell culture supernatant of human vascular endothelial growth factor receptor-antibody fusion protein was loaded as a sample, so that the recombinant human vascular endothelial growth factor receptor-antibody fusion protein was adsorbed on the affinity chromatography medium, and after loading, continue to use 1mol / LNaCl concentration of 10mmol / L, pH value of 7.5 disodium hydrogen phosphate-sodium dihydrogen phosphate equilibrium buffer for washing, and then citric acid-sodium citrate buffer solution with a concentration of 2...

Embodiment 2

[0070] A method for purifying recombinant human vascular endothelial growth factor receptor-antibody fusion protein, the specific steps of which are:

[0071] Affinity chromatography

[0072] (1) Equilibrate the Mabselect affinity chromatography column (5.0×20cm) with 1000mL disodium hydrogen phosphate-sodium dihydrogen phosphate equilibration buffer containing 1mol / LNaCl at a concentration of 10mmol / L and a pH value of 7.0, and the recombinant The cell culture supernatant of human vascular endothelial growth factor receptor-antibody fusion protein was loaded as a sample, so that the recombinant human vascular endothelial growth factor receptor-antibody fusion protein was adsorbed on the affinity chromatography medium, and after loading, continue to use 1mol / LNaCl concentration of 10mmol / L, pH value of 7.0 disodium hydrogen phosphate-sodium dihydrogen phosphate equilibrium buffer for washing, and then use concentration of 10mmol / L, pH value of 5.5 citric acid-sodium citrate b...

Embodiment 3

[0087] A method for purifying recombinant human vascular endothelial growth factor receptor-antibody fusion protein, the specific steps of which are:

[0088] Affinity chromatography

[0089] (1) MabselectSure affinity chromatography column (5.0 × 20cm) is equilibrated with 1000mL disodium hydrogen phosphate-sodium dihydrogen phosphate equilibrium buffer solution containing 0.9mol / LNaCl concentration of 20mmol / L and pH value of 7.5, and The cell culture supernatant of the recombinant human vascular endothelial growth factor receptor-antibody fusion protein was loaded as a sample, so that the recombinant human vascular endothelial growth factor receptor-antibody fusion protein was adsorbed on the affinity chromatography medium. The concentration of 0.9mol / LNaCl is 20mmol / L, the pH value is 7.5 disodium hydrogen phosphate-sodium dihydrogen phosphate equilibrium buffer solution for washing, and then the concentration is 30mmol / L, the pH value is 6.0 citric acid-sodium citrate Bu...

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Abstract

The invention relates to production of a genetic engineering drug by a recombinant DNA technology, belongs to the biotechnical field, and particularly relates to a purification method of a recombinant human vascular endothelial growth factor receptor-antibody fusion protein, that is to say, a cell supernatant containing the recombinant human vascular endothelial growth factor receptor-antibody fusion protein is successively subjected to affinity chromatography, anion exchange chromatography, cation exchange chromatography, hydrophobic chromatography and desalination chromatography to obtain the high-quality recombinant human vascular endothelial growth factor receptor-antibody fusion protein. The purification method has the characteristics of low cost, high purity of the target protein, easily controlled technological process, easily large-scale enlarged production and the like.

Description

technical field [0001] The invention relates to the production of genetic engineering drugs by using recombinant DNA technology, in particular to a purification method of recombinant human vascular endothelial growth factor receptor-antibody fusion protein, which belongs to the field of biotechnology. Background technique [0002] Vascular endothelial growth factor (vascular endothelial growth factor, VEGF) stimulates the formation of new blood vessels by combining with a specific receptor - vascular endothelial growth factor receptor (vascular endothelial growth factor receptor, VEGFR). In the process of tumor growth, the number of VEGFs surges, and the regulation imbalance between VEGFs and angiogenesis inhibitors greatly promotes the division, proliferation and migration of endothelial cells, improves vascular permeability, and provides a good microenvironment for tumor growth and metastasis. . [0003] Recombinant human vascular endothelial growth factor receptor-antibo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C07K1/22C07K1/20C07K1/18C07K1/16
Inventor 孙丽霞陈俊良孙波邢婷婷贾国栋朱鹏飞王庆民郑焕兰丁仁志石干
Owner QILU PHARMA CO LTD
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