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Preparation method and application of nickel metal chromatography medium based on 6B agarose microspheres

A technology of agarose microspheres and metal layers, which is applied in the field of protein purification, can solve the problems of agarose physical and chemical properties, cumbersome cross-linking steps, and low density of medium-coupling ligands, achieving high hardness, good elution effect, The effect of increasing the surface area

Inactive Publication Date: 2016-04-20
武汉菲恩生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] The purpose of the present invention is to overcome the cumbersome cross-linking steps in the existing agarose microspheres, the low density of the medium-coupling ligand after cross-linking and the influence of the physical and chemical properties of the agarose on the purification effect

Method used

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  • Preparation method and application of nickel metal chromatography medium based on 6B agarose microspheres
  • Preparation method and application of nickel metal chromatography medium based on 6B agarose microspheres

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Embodiment 1

[0036] In order to solve the problems of cumbersome cross-linking steps in the existing agarose microspheres, the low density of the medium-coupling ligand after cross-linking, and the influence of the physical and chemical properties of the agarose on the purification effect, this example provides a 6B agarose microsphere-based The preparation method of nickel metal chromatography medium, comprises the steps:

[0037] 1) Mix epichlorohydrin, 2M sodium hydroxide and 6B agarose microspheres at a volume ratio of 0.2:0.8:1, and react for 3 hours at a reaction temperature of 40°C to prepare activated agarose microspheres. spheres; wherein, 2M sodium hydroxide means that the concentration of sodium hydroxide is 2mol / L; 6B agarose microspheres means that the agarose content accounts for 6% of the total mass.

[0038] 2) Mix 1M sodium hydroxide, 0.3g / mL dextran aqueous solution, and the activated agarose microspheres prepared in step 1) at a volume ratio of 0.8:1:1, and react at a re...

Embodiment 2

[0049] This embodiment provides a kind of preparation method of nickel metal chromatography medium based on 6B agarose microspheres, comprising the following steps:

[0050] 1) Mix epichlorohydrin, 2M sodium hydroxide and 6B agarose microspheres at a volume ratio of 0.1:0.9:1, and react for 3 hours at a reaction temperature of 25°C to prepare activated agarose microspheres. spheres; wherein, 2M sodium hydroxide means that the concentration of sodium hydroxide is 2mol / L; 6B agarose microspheres means that the agarose content accounts for 6% of the total mass.

[0051] 2) Mix 1M sodium hydroxide, 0.1g / mL dextran aqueous solution, and the activated agarose microspheres prepared in step 1) at a volume ratio of 0.3:1:1, and react at a reaction temperature of 25°C , and reacted for 5 hours to obtain dextran-modified 6B agarose microspheres; wherein, the molecular weight of the dextran used was 40,000.

[0052] 3) Mix epichlorohydrin, 2M sodium hydroxide and the dextran-modified 6B ...

Embodiment 3

[0062] This embodiment provides a method for preparing a nickel metal chromatography medium based on 6B agarose microspheres, comprising the following steps:

[0063] 1) Mix epichlorohydrin, 2M sodium hydroxide and 6B agarose microspheres in a volume ratio of 0.5:0.5:1, and react for 6 hours at a reaction temperature of 70°C to prepare activated agarose microspheres. spheres; wherein, 2M sodium hydroxide means that the concentration of sodium hydroxide is 2mol / L; 6B agarose microspheres means that the agarose content accounts for 6% of the total mass.

[0064] 2) Mix 1M sodium hydroxide, 0.3g / mL dextran aqueous solution, and the activated agarose microspheres prepared in step 1) at a volume ratio of 0.9:1:1, and the reaction temperature is 25°C. , and reacted for 10 hours to obtain dextran-modified 6B agarose microspheres; wherein, the molecular weight of the dextran used was 40,000.

[0065] 3) Mix epichlorohydrin, 2M sodium hydroxide and the dextran-modified 6B agarose micr...

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Abstract

The invention provides a preparation method of a nickel metal chromatography medium based on 6B agarose microspheres. The agarose microspheres are activated through epoxy chloropropane to be polymerized with glucan, the superficial area of the agarose microspheres is increased on the condition of not reducing the number of surface hydroxyl groups of the agarose microspheres, the charge effect of the agarose microspheres is effectively reduced through glucan molecules, the activated agarose microspheres are connected with L-asparaginic acid again and subjected to bromoacetic acid treatment, a long lengthened arm is generated, it is ensured that nickel metal can be efficiently combined with recombinant protein, and the purification effect of the medium is improved. The problems that in existing agarose microspheres, the crosslinking step is tedious, the medium coupling ligand density is low after crosslinking, and the agarose physicochemical property affects the purification effect are solved.

Description

technical field [0001] The invention belongs to the technical field of protein purification, and in particular relates to a preparation method and application of a nickel metal chromatography medium based on 6B agarose microspheres. Background technique [0002] The rapid development of modern biomedical industry requires the performance of biomaterials to be gradually improved. Affinity chromatography media prepared from agarose are widely used in protein separation engineering, and the performance of the media directly determines the efficiency of separation and purification. [0003] Agarose is rich in hydroxyl groups and has good biosolubility. It is the most widely used matrix synthesis material in the process of separation and purification. However, due to the low hardness of ordinary agarose microspheres, it is easy to deform under high pressure conditions, reducing the size of the gaps between the microspheres, which directly leads to a decrease in the flow rate of ...

Claims

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Application Information

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IPC IPC(8): B01J20/24B01J20/30C07K1/22B01D15/20
CPCB01J20/24B01D15/203B01J20/0229B01J2220/46B01J2220/4825C07K1/22
Inventor 董垚郑忠亮夏其林
Owner 武汉菲恩生物科技有限公司
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