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Magnetic agarose composite microspheres as well as preparation method and application thereof

A technology of composite microspheres and agarose, which is applied in the preparation of microspheres, microcapsule preparations, etc., can solve problems such as difficult to wash off, long incubation time, mechanical and biological loss, etc., and achieve improved magnetic response and targeting , The effect of improving the extraction efficiency

Active Publication Date: 2018-07-20
湖北新纵科病毒疾病工程技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
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Problems solved by technology

However, this porous structure also brings some problems, such as: 1) Antibodies are trapped in the pores and are difficult to wash off, so a lot of washing is required to reduce the background; 2) The washing of immunoprecipitation is carried out in microcentrifuge tubes Yes, the exchange between liquids is done through a pipette gun, it is easy to lose samples
3) The contact between the antibody on the microsphere and the target protein is slow, requiring a long incubation time
4) Long incubation time and extensive washing lead to mechanical and biological (protease hydrolysis) loss of the target protein
For a long time, people have been using ferric oxide as the magnetic core to prepare agarose magnetic microspheres, but when the particle size is small, its magnetic response is not ideal, and its purification effect is not ideal.

Method used

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  • Magnetic agarose composite microspheres as well as preparation method and application thereof

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Embodiment 1

[0029] This embodiment provides a method for preparing magnetic agarose composite microspheres, including:

[0030] (1) 0.1g ferric oxide is dispersed in 16mL aqueous solution containing 0.5g agarose;

[0031] (2) Take 4.0g Span80 and dissolve it in 100mL liquid paraffin;

[0032] (3) Under the condition of 80°C, mix the two solutions, and vigorously mechanically stir (750-800 rpm) for 30 minutes, after cooling to room temperature, wash with ultrapure water and ethanol three times to obtain ferric oxide / agar Sugar complex microspheres;

[0033] (4) Disperse 5g of ferric oxide / agarose composite microspheres in 5mL of ultrapure water, add 0.5mL of epichlorohydrin and 6mL of 5M aqueous sodium hydroxide solution and react at 60°C for 120min to obtain cross-linked trioxide Iron / agarose composite microspheres.

[0034] (5) Take 5 g of cross-linked ferric oxide / agarose composite microspheres and disperse them in 6.5 mL of 2M aqueous sodium hydroxide solution, add 2 mL of epichloro...

Embodiment 2

[0036] This embodiment provides a method for preparing magnetic agarose composite microspheres, including:

[0037] (1) 0.1g ferric oxide is dispersed in 16mL aqueous solution containing 0.5g agarose;

[0038] (2) Take 4.0g Span80 and dissolve it in 100mL liquid paraffin;

[0039] (3) Under the condition of 80°C, mix the two solutions, and vigorously mechanically stir (750-800 rpm) for 30 minutes, after cooling to room temperature, wash with ultrapure water and ethanol three times to obtain ferric oxide / agar Sugar complex microspheres;

[0040] (4) Disperse 5g of ferric oxide / agarose composite microspheres in 5mL of ultrapure water, add 0.6mL of epichlorohydrin and 6mL of 5M aqueous sodium hydroxide solution and react at 60°C for 120min to obtain cross-linked trioxide Iron / agarose composite microspheres.

[0041] (5) Take 5 g of cross-linked ferric oxide / agarose composite microspheres and disperse them in 6.5 mL of 2M aqueous sodium hydroxide solution, add 2.4 mL of epichlo...

Embodiment 3

[0043] This embodiment provides a method for preparing magnetic agarose composite microspheres, including:

[0044] (1) 0.1g ferric oxide is dispersed in 16mL aqueous solution containing 0.5g agarose;

[0045] (2) Take 4.0g Span80 and dissolve it in 100mL liquid paraffin;

[0046] (3) Under the condition of 80°C, mix the two solutions, and vigorously mechanically stir (750-800 rpm) for 30 minutes, after cooling to room temperature, wash with ultrapure water and ethanol three times to obtain ferric oxide / agar Sugar complex microspheres;

[0047](4) Disperse 5g of ferric oxide / agarose composite microspheres in 5mL of ultrapure water, add 0.3mL of epichlorohydrin and 6mL of 5M aqueous sodium hydroxide solution and react at 60°C for 120min to obtain cross-linked trioxide Iron / agarose composite microspheres.

[0048] (5) Take 5g of cross-linked ferric oxide / agarose composite microspheres and disperse them in 6.5mL of 2M sodium hydroxide aqueous solution, add 1.2mL of epichlorohy...

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Abstract

The invention provides magnetic agarose composite microspheres as well as a preparation method and an application thereof. The preparation method of the magnetic agarose composite microspheres comprises the steps as follows: Fe3O4 / agarose composite microspheres are sequentially crosslinked and activated with epichlorohydrin as a crosslinking agent and an activating agent and 1,4-dioxane as a spacer arm. The agarose composite microspheres obtained with the preparation method have improved magnetic responsiveness, targeting and purification efficiency for target protein in cell lysate.

Description

technical field [0001] The invention relates to the technical field of polymer microspheres, in particular to a magnetic agarose composite microsphere and its preparation method and application. Background technique [0002] Traditional agarose microspheres are porous and have a high surface area where proteins can come into contact with each other and can hold large volumes of liquid. When purifying a large amount of protein, traditional agarose microspheres are the most ideal material. However, this porous structure also brings some problems, such as: 1) Antibodies are trapped in the pores and are difficult to wash off, so a lot of washing is required to reduce the background; 2) The washing of immunoprecipitation is carried out in microcentrifuge tubes Yes, the exchange between liquids is done through a pipette, and it is easy to lose samples. 3) The contact between the antibody on the microsphere and the target protein is slow, requiring a long incubation time. 4) Lon...

Claims

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Application Information

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IPC IPC(8): B01J13/14C08J3/24C08L5/12C08K3/22
CPCB01J13/14C08J3/24C08J2305/12C08K2003/2275
Inventor 王华林张涛张科登
Owner 湖北新纵科病毒疾病工程技术有限公司
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