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Novel method for preparing magnetic microspheres and separating antibody of magnetic microspheres

A technology of magnetic microspheres and a new method, applied in the biological field, can solve the problems of high control requirements of equipment and process parameters, complicated operation in the expanded bed mode, etc. Effect

Active Publication Date: 2015-12-23
南京佰抗生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the expanded bed mode can also avoid the removal of solid particles from the feed liquid, the operation of the expanded bed mode is complex and requires high control of equipment and process parameters

Method used

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  • Novel method for preparing magnetic microspheres and separating antibody of magnetic microspheres
  • Novel method for preparing magnetic microspheres and separating antibody of magnetic microspheres

Examples

Experimental program
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Effect test

Embodiment 1

[0022] Weigh 2g of agarose, 1g of nanometer ferric oxide, 1g of sodium chloride and 50g of deionized water, ultrasonically mix and heat to melt the agarose to obtain an aqueous phase; mix the aqueous phase with 324g of vacuum pump oil and 10.8g of Tween Mix at 80°C, stir at high speed at 70°C for 2 hours, place in an ice-water bath to cool down after emulsification, and collect the microspheres; mix 10g of microspheres with 10g of 1M sodium hydroxide and 2g of epichlorohydrin, and shake at room temperature for 1 hour , followed by adding 1g of sodium borohydride, shaking at room temperature for 8 hours to obtain a crosslinked magnetic microsphere matrix; weighing 10g of crosslinked magnetic microspheres, adding 10g of deionized water, 5g of allyl bromide and 5g of sodium hydroxide, Activate by shaking at room temperature for 24 hours, wash with water, 5g N-bromosuccinimide and 30g deionized water, shake at room temperature for 1 hour, wash with water to obtain activated magneti...

Embodiment 2

[0024] Weigh 2g of agarose, 1g of nanometer ferric oxide, 1g of sodium chloride and 50g of deionized water, ultrasonically mix and heat to melt the agarose to obtain an aqueous phase; mix the aqueous phase with 324g of vacuum pump oil and 10.8g of Tween Mix at 80°C, stir at high speed at 70°C for 2 hours, place in an ice-water bath to cool down after emulsification, and collect the microspheres; mix 10g of microspheres with 10g of 1M sodium hydroxide and 2g of epichlorohydrin, and shake at room temperature for 1 hour , followed by adding 1g of sodium borohydride, shaking at room temperature for 8 hours to obtain a crosslinked magnetic microsphere matrix; weighing 10g of crosslinked magnetic microspheres, adding 10g of deionized water, 5g of allyl bromide and 5g of sodium hydroxide, Activate by shaking at room temperature for 24 hours, wash with water, 5g N-bromosuccinimide and 30g deionized water, shake at room temperature for 1 hour, wash with water to obtain activated magneti...

Embodiment 3

[0026]Weigh 2g of agarose, 1g of nanometer ferric oxide, 1g of sodium chloride and 50g of deionized water, ultrasonically mix and heat to melt the agarose to obtain an aqueous phase; mix the aqueous phase with 324g of vacuum pump oil and 10.8g of Tween Mix at 80°C, stir at high speed at 70°C for 2 hours, place in an ice-water bath to cool down after emulsification, and collect the microspheres; mix 10g of microspheres with 10g of 1M sodium hydroxide and 2g of epichlorohydrin, and shake at room temperature for 1 hour , followed by adding 1g of sodium borohydride, shaking at room temperature for 8 hours to obtain a crosslinked magnetic microsphere matrix; weighing 10g of crosslinked magnetic microspheres, adding 10g of deionized water, 5g of allyl bromide and 5g of sodium hydroxide, Activate by shaking at room temperature for 24 hours, wash with water, 5g N-bromosuccinimide and 30g deionized water, shake at room temperature for 1 hour, wash with water to obtain activated magnetic...

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Abstract

The invention discloses a novel method for preparing magnetic microspheres and separating an antibody of the magnetic microspheres. The novel method comprises the specific steps that 1, a magnetic microsphere substrate is prepared; 2, the magnetic microsphere substrate is activated; 3, functional ligands are coupled; 4, the microspheres are added into monoclonal antibody cell sap and are mixed with and absorb an antibody through oscillation; 5, a magnet is used for attracting the microspheres on the outer wall of a triangular flask, the cell sap is poured out, and flushing is performed by using an equilibration buffer solution; 6, a de-absorption buffer solution is added, full oscillation is performed, the microspheres are attracted from the outer wall of the triangular flask by using the magnet, and the de-absorption buffer solution containing a product is collected. The novel magnetic microspheres prepared by means of the method and having hydrophobic charge inductive effect integrate the hydrophobic charge inductive effect and magnetic response, have the advantages of being high in protein carrying capacity, moderate in de-absorption condition, simple and convenient to separate and the like and can be used for direct and efficient separation of the antibody from feed liquid containing solid impurities.

Description

technical field [0001] The invention relates to a new method for preparing magnetic microspheres and antibody separation, and belongs to the field of biotechnology. Background technique [0002] Antibodies are an important class of biological macromolecules that can specifically recognize corresponding antigens. At present, antibodies have been widely used in the fields of disease treatment, in vitro diagnosis and detection, tumor localization imaging and biological separation. [0003] Whether it is separating antibodies from blood, ascites, colostrum or cell culture fluid, it is usually necessary to remove cells and solid impurities in the feed solution by centrifugation or filtration, and then combine steps such as affinity chromatography for purification. There are many steps in the whole process, and the operation is cumbersome. At the same time, the affinity chromatography packing has the disadvantages that the ligand is easy to fall off and the price is expensive. T...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J13/02G01N33/531G01N33/543G01N33/553
Inventor 顾佳黎童红飞
Owner 南京佰抗生物科技有限公司
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