Method for purifying VEGF capture agent fusion protein

A purification method and fusion protein technology, applied in the field of purification of human vascular endothelial growth factor receptor-antibody fusion protein, can solve problems such as increasing the cost of drug production

Active Publication Date: 2019-06-25
SHANDONG BIOANTY BIOLOGICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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And these complicated purification processes will

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  • Method for purifying VEGF capture agent fusion protein
  • Method for purifying VEGF capture agent fusion protein

Examples

Experimental program
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Embodiment 1

[0028] Take an appropriate amount of VEGF capture agent fusion protein fermentation broth, and purify according to the following steps and conditions, specifically including the following steps:

[0029] (1) Protein A affinity chromatography

[0030] Equilibrate buffer with 1L phosphate (pH7.0, 20mM NaH 2 PO 4 , 110mM NaCl) to equilibrate the affinity chromatography column MabSuReLX, and load the cell culture supernatant containing the VEGF capture agent fusion protein, so that the VEGF capture agent fusion protein is adsorbed on the protein A affinity medium. After loading the samples, the above equilibration buffer was used to wash, and then the protein was eluted with pH 2.90 and 100 mM Glycine buffer, and the eluate was collected. Then use 1mol / L phosphoric acid to acidify the eluate to pH 3.45 to inactivate the virus.

[0031] (2) Anion exchange chromatography

[0032] Equilibrate the anion-exchange chromatography column QFF with 1L Tris equilibrium buffer (pH8.40, 50...

Embodiment 2

[0037] Similar to Example 1, an appropriate amount of VEGF capture agent fusion protein fermentation broth was taken again, and purified according to the following steps and conditions, specifically as follows:

[0038] (1) Protein A affinity chromatography

[0039] Equilibrate buffer with phosphate (pH7.2, 20mM NaH 2 PO 4 , 110mM NaCl) to equilibrate the affinity chromatography column MabSuReLX, and load the cell culture supernatant containing the VEGF capture agent fusion protein encoded by SEQ ID NO:1, so that the VEGF capture agent is adsorbed on the protein A affinity medium . After loading the sample, wash with equilibration buffer, then use pH 3.00 Glycine (100 mM) eluent to elute the protein, and collect the eluate. The eluate was then acidified to pH 3.45 with phosphoric acid for virus inactivation.

[0040] (2) Anion exchange chromatography

[0041] Equilibrate the anion-exchange chromatography column QFF with Tris equilibrium buffer (pH8.50, 50mM Tris, 50mM NaC...

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Abstract

The invention relates to a method for purifying VEGF capture agent fusion protein. The method comprises the following steps: (1) affinity chromatography; (2) anion exchange chromatography; and (3) cation exchange chromatography. The purification process of the invention is simple and can effectively reduce the preparation cost, which is very beneficial for mass production.

Description

technical field [0001] The invention relates to the production of genetic engineering medicine by recombinant DNA technology, especially a purification method of human vascular endothelial growth factor receptor-antibody fusion protein. Background technique [0002] Proteins, often referred to as "biologics," play an important role in today's medicine. In order to ensure the safety of biopharmaceuticals to humans, impurities such as nucleic acids, viruses, and host cell proteins that may cause serious harm must be specifically removed. In addition, the Food and Drug Administration has very high standards for the quality of human protein. One of the greatest challenges in human biologics is the development of cost-effective and efficient protein purification methods at commercial scale. [0003] Typically, a mammalian or bacterial cell line that has been genetically engineered to produce the protein of interest is used to produce the protein through cell culture by insertin...

Claims

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Application Information

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IPC IPC(8): C07K19/00C07K1/22C07K1/18
Inventor 赵燕燕陶文杰刘丽丽王鑫王广珺
Owner SHANDONG BIOANTY BIOLOGICAL TECH CO LTD
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