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Application of protein A affinity chromatography medium

A chromatographic medium and affinity technology, applied in chemical instruments and methods, alkali metal oxides/hydroxides, inorganic chemistry, etc., can solve problems such as washing with high-concentration NaOH, and achieve good column packing repeatability, Greatly improved, soft and hard effects

Inactive Publication Date: 2017-12-01
苏州纳微分离纯化技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, a fatal flaw in the structure of glass beads is that it cannot withstand the gold standard of high-concentration NaOH cleaning, which limits its application in large-scale biological samples.

Method used

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  • Application of protein A affinity chromatography medium
  • Application of protein A affinity chromatography medium
  • Application of protein A affinity chromatography medium

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Polyacrylate microspheres prepared by Suzhou Nawei Technology Co., Ltd. were used. When the microspheres are prepared and polymerized, the monovinyl monomer contains part of glycidyl methacrylate. At the same time, commercial hyperbranched polyvinyl alcohol (Hyperbranched G5-PEG6k-OH, ALDRICH) was used. First weigh 20 g of polymer microspheres (with a particle size of 50 μm and a pore size of about 100 nm) in a dry tray, and put them into a 100 ml three-neck bottle. Then, take a 100mL beaker and prepare 20mL 0.6M NaOH solution containing 2mg / mL sodium borohydride, while adding 20g hyperbranched polyvinyl alcohol (Hyperbranched G5-PEG6k-OH, ALDRICH) and make it dispersed and dissolved. Then, the dissolved polyvinyl alcohol solution was added to the above-mentioned flask. Under the condition of mechanical stirring, react at 37°C for 3.5h. After the reaction, the sand core was drained and washed twice with 50mL deionized water, once with 50mL absolute ethanol, and then ...

Embodiment 2

[0044] The reaction device is the same as in Example 1, using the Vinyl polymer microspheres (monodisperse microspheres with a particle size of 15 μm and a pore size of about ). During the preparation and polymerization of microspheres, the monovinyl monomer also contains part of glycidyl methacrylate. Reaction control and ball washing steps are the same as in Example 1.

[0045] Referring to Example 1, 1 g of the product microspheres prepared in this example was taken, and loaded into a 50×2.1 mm stainless steel chromatographic column with a slurry method under a pressure of 1450 psi. Then the column is installed on the Waters high performance liquid chromatograph (e2695), with 0.05mol / L sodium dihydrogen phosphate+0.1mol / L sodium acetate+0.1mol / L sodium chloride as balance liquid, 0.05mol / L glycine , PH=3.0 is the eluent. The chromatographic analysis conditions and process are shown in the table below.

[0046]

[0047]

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Abstract

The invention provides an application of a protein A chromatography medium. The protein A chromatography medium is obtained by surface-hydrophilic dendrimer modification, activation and protein A coupling by taking a polymer microsphere containing a vinyl monomer as a raw material. When being used as an antibody and Fc fusion protein separation and purification medium, the protein A chromatography medium provided by the invention is capable of greatly increasing the flow rate, short in separation time, good in separation effect and low in cost so as to be suitable for rapid separation and purification of antibodies and Fc fusion proteins.

Description

technical field [0001] The invention relates to the fields of chemical material synthesis and biomedical downstream technology, and in particular to the application of a protein A affinity chromatography medium. Background technique [0002] In recent years, antibody drugs have developed rapidly. In 2016, 6 of the top 10 drugs in global sales were antibody drugs, and all of the top 5 were antibody drugs. The purification of antibody drugs usually adopts the classic three-step or two-step chromatography method, and the protein A affinity medium can be used as an antibody capture step due to the unique and specific adsorption ability of multiple regions on the protein A ligand for the Fc region of the antibody. The one-step process can remove most of the impurities in the cell fermentation broth, and the purity can reach more than 99%. With the development of antibody drugs, the affinity chromatography media used for antibody capture has also achieved rapid development. [...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J20/30B01J20/26B01J20/28C07K1/22
CPCB01J20/265B01J20/28019C07K1/22
Inventor 谢岩生王永向朱祺陈荣姬吴俊成江必旺
Owner 苏州纳微分离纯化技术有限公司
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