Preparation method of bovine serum albumin molecular surface imprinted polymer microspheres

A bovine serum albumin and surface imprinting technology, which is applied in the fields of protein engineering and functional polymers, can solve the problems of unsuitable proteins, impact of imprinting effect, competition and destruction, etc., to improve imprinting efficiency, improve imprinting effect, and expand the selection range Effect

Active Publication Date: 2015-02-18
山西铁居环保科技有限公司
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  • Abstract
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  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, there are obviously several problems in western blotting: (1) The traditional embedding blotting method is not suitable for protein blotting
In the highly cross-linked block polymer network, the mobility of protein macromolecules is poor, resulting in low imprinting efficiency and poor binding performance, and imprinted molecules are not easy to elute [Kryscio D R, Peppas N A. Critical review and perspective of macromolecularly imprinted polymers, Acta Biomaterialia, 2012, 8: 461–473]; (2) The choice of solvent is limited
In an organic solvent, although the hydrogen bond interaction between the template protein molecule and the monomer (so far, most protein molecular imprinting mainly relies on the hydrogen bond interaction between the template molecule and the functional monomer) is not dis

Method used

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preparation example Construction

[0018] A method for preparing bovine serum albumin molecular surface imprinted polymer microspheres, comprising the steps of:

[0019] Step 1: Soak 3~8% cross-linked polyvinyl alcohol microspheres in acetone solution to make them fully swell, then add 3~10% methacryloyl chloride and a small amount of acid-binding agent sodium carbonate, keep the temperature at 25~50℃ React for 5-10 hours to prepare modified microspheres with a large number of methacrylate groups bonded to the surface;

[0020] Step two, in neutral aqueous solution, relying on the strong electrostatic interaction, many cationic monomers acryloyloxyethyltrimethylammonium chloride automatically bind around the macromolecules of bovine serum albumin: that is, in neutral phosphate buffer Add 0.5~5% acryloyloxyethyltrimethylammonium chloride to the liquid, and add according to the molar ratio of acryloyloxyethyltrimethylammonium chloride to bovine serum albumin is 100:1~500:1 Bovine serum albumin, after stirring th...

Embodiment 1

[0024] Soak 2 g of cross-linked polyvinyl alcohol microspheres in 40 mL of acetone solution to make them fully swell, then add 2 mL of methacryloyl chloride and a small amount of acid-binding agent sodium carbonate, and react at a constant temperature of 40 °C for 10 h to obtain the modified sex microspheres.

[0025] Dissolve 3.1 g template molecule bovine serum albumin in 95 mL pH=7.4 phosphate buffer, add 5 mL monomer acryloyloxyethyltrimethylammonium chloride, let them interact for a period of time, and dissolve the solution Transfer to a four-necked flask equipped with a reflux condenser, a stirrer and a thermometer; add 1 g of modified microspheres, soak for 12 h, then add 0.16 g of cross-linking agent N,N′-methylenebisacrylamide, and pass nitrogen gas For 30 minutes, remove the air in the system; raise the temperature of the system to 35°C, add 0.058 g of ammonium persulfate and 0.029 g of sodium bisulfite under nitrogen protection and stirring conditions, and make the ...

Embodiment 2

[0027]Soak 4 g of cross-linked polyvinyl alcohol microspheres in 50 mL of acetone solution to make them fully swell, then add 5 mL of methacryloyl chloride and a small amount of acid-binding agent sodium carbonate, and react at a constant temperature of 50 °C for 5 h to obtain the modified sex microspheres.

[0028] Dissolve 1.8 g template molecule bovine serum albumin in 95 mL pH=7.4 phosphate buffer, add 5 mL monomer acryloyloxyethyltrimethylammonium chloride, let them interact for a period of time, and dissolve the solution Transfer to a four-neck flask equipped with a reflux condenser, a stirrer and a thermometer; add 3g of modified microspheres, soak for 12 h, then add 0.1g of cross-linking agent N,N′-methylenebisacrylamide, and pass nitrogen for 30min , Exclude the air in the system; raise the temperature of the system to 25°C, add 0.072 g of ammonium persulfate and 0.039 g of sodium bisulfite under the protection of nitrogen and stir to make the surface graft cross-link...

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Abstract

The invention provides a preparation method of bovine serum albumin molecular surface imprinted polymer microspheres. The method comprises the following steps: 1, adding methacryloyl chloride into an acetone solution containing crosslinked polyvinyl alcohol microspheres to form modified microspheres; 2, combining acryloyloxyethyl trimethyl ammonium chloride with bovine serum albumin molecules in a neutral aqueous solution by virtue of strong electrostatic interactions so as to form a host-guest complex; 3, carrying out a crosslinking copolymerization reaction on the metacrylate modified microspheres, the host-guest complex, a crosslinking agent N,N'-methylene bisacrylamide and an initiating agent so as to achieve the surface imprinting of the bovine serum albumin molecules; and 4, repeatedly flushing the imprinted microspheres with sodium chloride and then flushing the imprinted microspheres with distilled water so as to obtain the bovine serum albumin molecular surface imprinted polymer microspheres with a large quantity of template molecular imprinted cavities retained on the surfaces. According to the method, the protein imprinting is carried out on the basis of the electrostatic interactions between the template protein and each functional monomer; and a new molecular design thought for developing a protein molecular imprinting technology is provided. Thus, the method provided by the invention has a positive reference value in the protein separation and purification field.

Description

technical field [0001] The invention belongs to the technical field of protein engineering and functional macromolecules, and relates to the preparation of molecular surface imprinted polymer microspheres with the characteristics of identifying and selecting biological macromolecules, in particular to the preparation of bovine serum albumin molecular surface imprinted polymer microspheres method. Background technique [0002] Molecularly imprinted polymers (MIPs) are well-tailored functional polymer materials, in which a large number of imprinted holes of template molecules are distributed inside. and other aspects are highly matched with template molecules, so that molecularly imprinted polymers can specifically recognize template molecules at the molecular level, which is similar to the specificity of antibody-antigen recognition and binding, so it is called artificial Antibody (Artificial antibody) or artificial acceptor (Artificial acceptor). Molecular imprinting techn...

Claims

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Application Information

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IPC IPC(8): C08F261/04C08F220/34C08F216/36C08F222/38C08J9/26B01J13/14
Inventor 史楠高保娇陈涛乔宗文李继文
Owner 山西铁居环保科技有限公司
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