A polypeptide bionano surface for serum-free cell culture and its preparation method

A cell culture and bio-nano technology, which is applied in biochemical equipment and methods, microorganisms, tissue culture, etc., can solve the problems of increased risk of cell culture experiment research and clinical application, cells that cannot meet clinical applications, and unsatisfactory cell adhesion effects, etc. problems, to achieve the effect of reducing the risk of clinical application, good adhesion and proliferation, and improving biocompatibility

Active Publication Date: 2014-10-22
BEAVERNANO TECH
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In the past research and application, the most used method for cell adherence culture is to add serum components to the medium to enhance cell adhesion ability, such as mesenchymal stem cells (MSC), liver cancer cells (HepG2), etc.; however, this The cells obtained by this method cannot meet the requirements of clinical application. This is mainly because the serum used for cell culture has problems such as unclear components, complex sources, and large differences between each batch due to individual differences. The experimental research and clinical application increase the risk
In addition, the methods for enhancing cell adhesion ability in serum-free culture system mainly include: coating the culture surface with biological protein, such as coating the culture surface with laminin in neural stem cell culture; or by some Physical and chemical methods are used to modify the culture surface, such as using plasma to treat the cell culture surface, introducing groups containing elements such as oxygen and nitrogen to improve the hydrophilicity of the surface, and improving the hydrophilicity of cells on the surface. and these traditional methods also have some objective deficiencies: first, the various coating proteins (such as Laminin) used as surface substrate coating materials are expensive, and limited in any relevant subject Funding is a severe test, and these proteins are generally animal-derived proteins, which will bring certain risks to human clinical treatment; secondly, the modified treatment using plasma technology alone, under the condition of serum-free culture, cells Adhesive effect is not ideal
However, in the process of modifying the surface of the culture plate in this way, some organic reagents are more or less introduced to assist in the completion of the grafting of these specific polypeptide sequences, and these organic reagents will also bring unresolved effects to the subsequent cell culture. known risk

Method used

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  • A polypeptide bionano surface for serum-free cell culture and its preparation method
  • A polypeptide bionano surface for serum-free cell culture and its preparation method
  • A polypeptide bionano surface for serum-free cell culture and its preparation method

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Embodiment 1

[0043] A method for preparing a polypeptide bionano surface for serum-free cell culture, specifically comprising the following steps:

[0044] (1) Put the cell culture surface made of polystyrene into the plasma reaction chamber for pretreatment for 5 minutes; the working parameters of the plasma reaction chamber are: power 35-600W, pressure 1-80Pa, gas flow rate 5-500cc / min, Temperature 25~45℃;

[0045] (2) Expose the pretreated culture surface to the air for 5 minutes;

[0046] (3) Polypeptide CH was synthesized according to the literature (Hu Chunling, Tang Yinping, Shi Jingni, etc. Solid-phase synthesis of anti-hepatic fibrosis active polypeptide of turtle shell. Medicine Herald. 2011, 30(5): 561-562.) 2 =CH-CONH-R-(RADA) 4 -GGIKVAV (where R is y=0);

[0047] (4) Dissolving the synthesized polypeptide in water to form a solution A with a mass volume concentration of 0.5%; spreading the solution A evenly to the culture surface obtained in step (2) to form a liquid surf...

Embodiment 2

[0052] A method for preparing a polypeptide bionano surface for serum-free cell culture, specifically comprising the following steps:

[0053] (1) Put the cell culture surface made of polyethylene into the plasma reaction chamber for pretreatment for 10 minutes; the working parameters of the plasma reaction chamber are: power 35-600W, pressure 1-80Pa, gas flow rate 5-500cc / min, temperature 25~45℃;

[0054] (2) Expose the pretreated culture surface to oxygen for 5 minutes;

[0055] (3) Polypeptide CH was synthesized according to the literature (Hu Chunling, Tang Yinping, Shi Jingni, etc. Solid-phase synthesis of anti-hepatic fibrosis active polypeptide of turtle shell. Medicine Herald. 2011, 30(5): 561-562.) 2 =CH-CONH-R-(RADA) 4 -GGYIGSR (where R is y=14); and dissolving the synthesized polypeptide in water to form a solution A with a mass volume concentration of 0.05%; spreading the solution A evenly to the culture surface obtained in step (2) to form a liquid surface lay...

Embodiment 3

[0060] A method for preparing a polypeptide bionano surface for serum-free cell culture, specifically comprising the following steps:

[0061] (1) Put the cell culture surface made of polyethylene into the plasma reaction chamber for pretreatment for 10 minutes; the working parameters of the plasma reaction chamber are: power 35-600W, pressure 1-80Pa, gas flow rate 5-500cc / min, temperature 25~45℃;

[0062] (2) Expose the pretreated culture surface to oxygen for 30 seconds;

[0063] (3) Polypeptide CH was synthesized according to the literature (Hu Chunling, Tang Yinping, Shi Jingni, etc. Solid-phase synthesis of anti-hepatic fibrosis active polypeptide of turtle shell. Medicine Herald. 2011, 30(5): 561-562.) 2 =CH-CONH-R-(RADA) 4 -GGAASIKVAVSADR (where R is y=0); and dissolving the synthesized polypeptide in water to form a solution A with a mass volume concentration of 0.5%; spreading the solution A evenly to the culture surface obtained in step (2) to form a liquid surfa...

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Abstract

The invention discloses a polypeptide biological nano surface for serum-free cell culture and a preparation method thereof. The polypeptide biological nano surface is prepared from the following steps of: pre-treating cell culture surface by plasma, then exposing the cell culture surface in air or oxygen, then spreading a polypeptide solution on the cell culture surface, and irradiating under ultraviolet light to obtain the polypeptide biological nano surface for serum-free cell culture. The obtained polypeptide biological nano surface is stable, can be stored for a long time at normal temperature, is definite in grafting ingredients on the grafted surface, and can substitute for routine various protein-coated surfaces in the market, so that cells realize excellent wall adhesion and proliferation under the serum-free condition; and as compared with various animal-derived protein-coated surface, short peptide has the characteristics of no heterology or weak heterology, therefore, the chemically synthesized polypeptide grafted surface greatly reduces clinical application risk, thereby laying a solid foundation for subsequent clinical application.

Description

technical field [0001] The invention belongs to the technical field of cell culture surface treatment, and in particular relates to a polypeptide bionano surface for serum-free cell culture and a preparation method thereof. Background technique [0002] With the continuous development of modern science, especially the rapid progress of biomedicine, cell culture has become a very important basic work in many research and application fields. [0003] For a long time, people have generally used polystyrene culture plates or culture bottles as tools for cell culture. Polystyrene is a colorless and transparent thermoplastic, usually non-toxic, odorless, colorless and transparent particles, glass-like brittle material, the general formula is [(CH 2 CHC 6 h 5 ) n ], the English name is Polystyrene, referred to as PS. Its products have excellent water resistance and moisture resistance. In people's production and life, polystyrene has a wide range of uses, and being a cell cul...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/00
Inventor 任辉董友玉秦炜韩蓝青张曙光
Owner BEAVERNANO TECH
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