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Preparation method of magnetic beads for protein purification

A protein purification and magnetic bead technology, applied in the biological field, can solve problems such as protein denaturation, and achieve rapid separation, convenient separation, and easy operation

Active Publication Date: 2017-01-04
RUBYBERRIES BIOTECHNOLOGIES CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This treatment method is more effective for the recovery and adsorption of heavy metal ions in industrial wastewater pollution, but in metal chelation affinity chromatography, too many metal ions will lead to protein denaturation, so it is not applicable

Method used

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  • Preparation method of magnetic beads for protein purification
  • Preparation method of magnetic beads for protein purification

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] (1) Take 9g of chitosan, add 900ml of pure water, add 3ml of glacial acetic acid under stirring, fully stir to dissolve, and cool to 4°C to prepare a chitosan solution; (2) Add 750g of magnetic powder to the 10L reaction system , 1875ml of distilled water, add 750ml of chitosan solution prepared in step (1), add 37.5ml of cold 4% glyoxal solution under high-speed stirring conditions, and mix well; (3) add 0.2mol / L K 2 HPO 4 Solution 220ml, stirred at high speed for 2h; (4) Add (NH 4 ) 2 SO 4 40g, while dynamically adjusting and maintaining the pH of the solution at about 8.5 with 1mol / L NaOH solution, stirring for 2 hours; (5) Add NaBH 4 10g, stirred for 2h, removed the supernatant, and washed twice with 5L of distilled water; (6) Added 66g of sodium chloroacetate, and stirred for 2h at 85°C and pH8.5; (7) The mixture obtained by the above steps, Wash twice with distilled water to obtain magnetic beads for protein purification.

[0023] Take 0.3 g (wet) of the a...

Embodiment 2

[0024] Example 2mol / L

[0025] (1) Take 9g of chitosan, add 900ml of pure water, add 3ml of glacial acetic acid under stirring, fully stir to dissolve, and cool to 4°C to prepare a chitosan solution; (2) Add 750g of magnetic powder to the 10L reaction system , 1875ml of distilled water, add 750ml of chitosan solution prepared in step 1, add 37.5ml of cold 4% glutaraldehyde solution under high-speed stirring conditions, and fully mix; (3) add 0.2m / L of K 2 HPO 4 Solution 220ml, stirred at high speed for 2h; (4) Add (NH 4 ) 2 SO 4 40g, while dynamically adjusting and maintaining the pH of the solution at about 8.5 with 1mol / L NaOH solution, stirring for 2 hours; (5) Add NaBH 4 10g, stirred for 2h, removed the supernatant, and washed twice with 5L distilled water; (6) Added 66g of sodium chloroacetate at 85°C and pH8.5, stirred for 2h; (7) The mixture obtained by the above steps, Wash twice with distilled water to obtain magnetic beads for protein purification.

[0026] T...

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Abstract

The invention provides a preparation method of magnetic beads for protein purification. Chitosan and magnetic powder are used as raw materials, core-shell structure particles of crosslinked chitosan coated magnetic powder are formed under the effect of a cross-linking agent, and the magnetic beads for protein purification are formed under reduction and carboxymethylation effects. The magnetic beads for protein purification are used for separation and purification of protein with his-tag, and the separation efficiency is greatly improved. The problem of protein denaturation possibly occurs during elution is avoided, and the protein separating efficiency can be improved. In addition, the preparation method is convenient and quick to operate, is suitable for separation of a turbid protein solution and is especially suitable for great industrial production.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for preparing magnetic beads used in protein separation and purification. Background technique [0002] The separation and purification of proteins mainly include methods such as salting out, centrifugation, gel electrophoresis and affinity chromatography. Among them, salting out, centrifugation and gel electrophoresis are limited by factors such as high cost and low efficiency, and their applications are relatively limited. Affinity chromatography is to make an affinity molecule with a special structure into a solid-phase adsorbent and place it in a chromatography column. When the protein mixture to be separated passes through the chromatography column, the protein with affinity with the adsorbent will be It will be adsorbed and retained in the chromatographic column, and can be separated and purified after being eluted. At present, for the separation and pu...

Claims

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Application Information

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IPC IPC(8): B01J13/14C07K1/14
CPCB01J13/14C07K1/14
Inventor 金彩科郑诚
Owner RUBYBERRIES BIOTECHNOLOGIES CO LTD
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