WO2023024335A1 - 一种促进细胞贴壁和生长的装置及其使用方法 - Google Patents

一种促进细胞贴壁和生长的装置及其使用方法 Download PDF

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Publication number
WO2023024335A1
WO2023024335A1 PCT/CN2021/137730 CN2021137730W WO2023024335A1 WO 2023024335 A1 WO2023024335 A1 WO 2023024335A1 CN 2021137730 W CN2021137730 W CN 2021137730W WO 2023024335 A1 WO2023024335 A1 WO 2023024335A1
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cell culture
electret
culture equipment
cell
tray
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PCT/CN2021/137730
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English (en)
French (fr)
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方鹏
曹江浪
李光林
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中国科学院深圳先进技术研究院
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Publication of WO2023024335A1 publication Critical patent/WO2023024335A1/zh

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/08—Flask, bottle or test tube
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/10—Petri dish
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/12—Well or multiwell plates
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/20—Material Coatings
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/48—Holding appliances; Racks; Supports
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
    • C12M35/02—Electrical or electromagnetic means, e.g. for electroporation or for cell fusion
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M37/00—Means for sterilizing, maintaining sterile conditions or avoiding chemical or biological contamination
    • C12M37/04—Seals

Definitions

  • the invention belongs to the technical field of cell culture, and in particular relates to a device for promoting cell attachment and growth and an application method thereof.
  • Adherent cells need to adhere to the surface of the culture device in order to migrate, proliferate and differentiate.
  • the adhesion-promoting matrix is generally coated on the surface of the culture device to enhance the interaction between the integrin (receptor) on the cell membrane surface and the adhesion protein (ligand) in the adhesion-promoting matrix function, improve the cell adhesion rate, or culture in a culture device containing a microstructure that absorbs the cell function.
  • CN112458048A discloses a primary culture of dental pulp stem cells that promotes adherence matrix and a culture method for primary dental pulp stem cells.
  • the invention provides a material that can promote the growth of cell adherence, which is used to coat culture dishes for dental pulp
  • the primary culture of stem cells the establishment of a mature in vitro culture system, and the provision of high-quality and abundant cell sources for subsequent experimental research.
  • polyphenol protein is used as an adhesion-promoting agent to pre-coat the culture dish, which increases the crawling rate of tissue block cells and the number of primary cells, shortens the primary preparation time, and improves the The success rate of primary culture.
  • adding polyphenol protein will change the composition of the medium and affect the normal physiological metabolism of cells.
  • this method only promotes the cultivation of dental pulp stem cells, and its applicability is poor.
  • the present invention provides a device for promoting cell attachment and growth and a method for using the device.
  • the device promotes cell attachment through the action of an electric field, and does not need to use an adhesion-promoting matrix for coating and culturing. equipment, and there is no need to use special microstructures to adsorb cells, which has little effect on the physiological state of cells and has high application value.
  • the present invention provides a device for promoting cell attachment and growth, the device comprising:
  • the thickness of the bottom wall of the cell culture equipment is 0.1-0.5 mm, for example, it can be 0.1 mm, 0.2 mm, 0.3 mm, 0.4 mm or 0.5 mm, etc. Other specific points within this value range can be selected. I won't repeat them one by one.
  • a stable electric field can be generated, thereby promoting the adhesion of the adherent cells, and further promoting the proliferation of the cells, the growth rate of the cells is faster, and the culture effect is better.
  • This process does not add an adhesion-promoting matrix to the culture medium or use a corresponding matrix to coat the cell culture device, nor does it rely on a device with a special microstructure, so it is convenient to use and easy to produce.
  • the thickness of the bottom wall of the cell culture equipment is between 0.1 and 0.5 In the range of mm, it can achieve a better effect of promoting adhesion and cell proliferation. If the thickness of the bottom wall of the cell culture equipment is less than 0.1 mm, the normal physiological activities of the cells will be affected due to the excessive electric field intensity. If the cell culture equipment If the thickness of the bottom wall of the device is greater than 0.5 mm, the effect of promoting cell adhesion and proliferation will not be produced because the electric field intensity received is too small.
  • the cell culture equipment includes any one or a combination of at least two of cell culture flasks, cell culture dishes or cell culture plates.
  • the tray includes support arms and a support base.
  • the height of the bracket arm is 3-5 mm, for example, it can be 3 mm, 3.2 mm, 3.4 mm, 3.6 mm, 3.8 mm, 4 mm, 4.2 mm, 4.4 mm, 4.6 mm, 4.8 mm or 5 mm etc.
  • the thickness of the support arm is 0.5 ⁇ 2 mm, such as 0.5 mm, 0.6 mm, 0.7 mm, 0.8 mm, 0.9 mm, 1 mm, 1.1 mm, 1.2 mm, 1.3 mm, 1.4 mm, 1.5 mm, 1.6 mm, 1.7 mm, 1.8 mm, 1.9 mm or 2 mm, etc., other specific point values within this value range can be selected, and will not be repeated here.
  • the thickness of the bottom is 0.8-1.5 mm, such as 0.8 mm, 0.9 mm, 1 mm, 1.1 mm, 1.2 mm, 1.3 mm, 1.4 mm or 1.5 mm, etc.
  • Other specific values within this range Point values can be selected, so I won’t go into details one by one here.
  • the thickness of the electret is 5-500 ⁇ m, for example, 5 ⁇ m, 10 ⁇ m, 50 ⁇ m, 100 ⁇ m, 150 ⁇ m, 200 ⁇ m, 250 ⁇ m, 300 ⁇ m, 350 ⁇ m, 400 ⁇ m, 450 ⁇ m ⁇ m or 500 ⁇ m, etc., other specific point values within this value range can be selected, and will not be repeated here.
  • the thickness of the electret can be determined according to the properties of the electret material used, the cell culture environment and other conditions.
  • the raw materials for the preparation of the cell culture equipment include polypropylene (PP), polystyrene (PS), polytetrafluoroethylene (PTFE), fluorinated ethylene propylene copolymer (FEP) or polychlorotrifluoroethylene (PCTFE ) in any one or a combination of at least two.
  • PP polypropylene
  • PS polystyrene
  • PTFE polytetrafluoroethylene
  • FEP fluorinated ethylene propylene copolymer
  • PCTFE polychlorotrifluoroethylene
  • the cell culture equipment is made of a polymer material with good biocompatibility, and its size and size can be determined according to specific actual conditions.
  • the raw material for preparing the electret includes polypropylene and/or fluorinated ethylene-propylene copolymer.
  • an electret is a dielectric material that exhibits a "quasi-permanent" charge.
  • the charges in an electret can be "real" charges, such as surface charges stored on the surface of the material and space charges stored in the bulk of the material; they can also be dipolar charges, that is, orientation dipoles (or displacement charges); or both Those who share it.
  • an electret with good biocompatibility and good charge storage capacity is selected.
  • the size of the electret can be determined according to specific actual conditions.
  • the raw materials for the sealing ring include biological glue and/or medical adhesive tape.
  • the sealing ring is selected from a material with good sealing performance, water resistance and viscosity to ensure that the electret sealed between the cell culture equipment and the tray is not affected by environmental factors such as moisture, and its size can be determined according to the actual situation. The situation is determined.
  • the device for promoting cell attachment and growth of the present invention includes:
  • the cell culture equipment includes any one or a combination of at least two of cell culture flasks, cell culture dishes or cell culture plates, and the thickness of the bottom wall of the cell culture equipment is 0.1-0.5 mm;
  • the pallet includes a supporting arm and a supporting base, and the height of the supporting arm is 3 to 5 mm, the thickness of the support arm is 0.5 ⁇ 2 mm, the thickness of the bottom is 0.8 ⁇ 1.5 mm;
  • the thickness of the electret is 5 ⁇ 500 ⁇ m
  • the raw materials for the preparation of the cell culture equipment include any one or a combination of at least two of polypropylene, polystyrene, polytetrafluoroethylene, fluorinated ethylene-propylene copolymer or polychlorotrifluoroethylene;
  • the preparation raw material of described electret comprises polypropylene and/or fluorinated ethylene propylene copolymer
  • the raw materials for the preparation of the sealing ring include biological glue and/or medical adhesive plaster.
  • the present invention provides a method of using the device for promoting cell attachment and growth described in the first aspect, the method of using comprising:
  • the device is placed in a culture environment and the cells are cultured.
  • the device for promoting cell attachment and growth is convenient to use, simple to operate, high in success rate and good in fault tolerance, which promotes the popularization and use of related devices.
  • the step of cleaning the electret is also included before the polarization.
  • said cleaning comprises ultrasonic cleaning.
  • the step of drying the electret is also included after the cleaning.
  • the polarization method includes a corona charging method and/or an electrical breakdown charging method.
  • the standing time is not less than 24 h, for example can be 24 h, 25 h, 26 h, 27 h, 28 h, 29 h or 30 h, etc., other specific point values within this value range can be selected, and will not be repeated here.
  • the function of standing still is to ensure the stability of the electric charge stored in the electret, thereby providing a stable electric field environment for the cells.
  • the using method further includes the step of replacing the culture medium and/or the electret.
  • replacing the medium and/or electret is not a necessary operation, it can be determined according to the specific cell growth state (such as cell shape, cell density, etc.), if the requirements are met, there is no need to replace the medium and/or electret; if the requirements are not met, the medium and/or electret need to be replaced.
  • the specific cell growth state such as cell shape, cell density, etc.
  • the method for using the device for promoting cell attachment and growth of the present invention comprises the following steps:
  • Ultrasonic cleaning is carried out on the electret, and after drying, the electret is polarized by the corona charging method and/or the electric breakdown charging method, and it is left to stand for not less than 24 h;
  • Inoculating cells into the sterilized cell culture equipment for example, adding a culture medium containing cells to be cultured into a cell culture dish;
  • the device for promoting cell attachment and growth described in the present invention is simple in structure and strong in applicability.
  • the electric field generated by the electret is used to promote cell attachment and proliferation without resorting to complicated culture equipment or adding an attachment-promoting matrix.
  • the physiological state of the device is less affected, does not affect the subsequent experimental operation, and the experimental results obtained are more accurate; by optimizing the preparation materials of the device, the thickness of the bottom wall and other related parameters, the cell adhesion rate is further improved, and the cell adhesion rate is improved.
  • the rate is between 40% and 50%; the preparation of the device is simple and efficient, convenient and flexible to use, has the value of being applied to actual scientific research work, and has broad application prospects.
  • Fig. 1 is the operation flowchart of the device for promoting cell attachment and growth of the present invention
  • Figure 2 is a schematic structural view of the device for promoting cell attachment and growth in Example 1 of the present invention (in the figure, 1-cell culture dish, 2-tray, 3-electret, 4-sealing ring);
  • Figure 3 is a schematic structural view of the device for promoting cell attachment and growth in Example 2 of the present invention (in the figure, 1-cell culture flask, 2-tray, 3-electret, 4-sealing ring);
  • Fig. 4 is a schematic structural diagram of the device for promoting cell attachment and growth in Example 3 of the present invention (in the figure, 1-cell culture plate, 2-tray, 3-electret, 4-sealing ring).
  • Cell culture dishes, cell culture flasks and cell culture plates were purchased from Taizhou Xinlian Chengrun Biotechnology Co., Ltd., and all were uncoated products.
  • the electret was purchased from Shanghai Electret New Material Co., Ltd.
  • Bioglue was purchased from Shenzhen Kanglibang Technology Co., Ltd.
  • HT1080 human fibrosarcoma cells were purchased from Wuhan Punuosai Life Technology Co., Ltd.
  • DMEM medium and FBS were purchased from Thermo Fisher Scientific (China) Co., Ltd.
  • the device for promoting cell attachment and growth includes a cell culture dish 1, a tray 2 supporting the cell culture dish 1, an electret 3 between the cell culture dish 1 and the tray 2, and the electret
  • the body 3 is sealed with a sealing ring 4 between the cell culture dish 1 and the tray 2;
  • the diameter of the bottom wall of the cell culture dish 1 is 35 mm, and the thickness of the bottom wall is 0.3 mm;
  • the tray 2 includes a support arm and a bottom, the height of the support arm is 4 mm, the thickness of the support arm is 1 mm, and the thickness of the support bottom is 1 mm;
  • the thickness of the electret 3 is 30 ⁇ m
  • the preparation raw material of described cell culture dish 1 is polystyrene
  • the preparation raw material of described electret 3 is fluorinated ethylene propylene copolymer
  • the raw material for the preparation of the sealing ring 4 is medical adhesive tape.
  • the device for promoting cell attachment and growth includes a cell culture bottle 1, a tray 2 supporting the cell culture bottle 1, an electret 3 between the cell culture bottle 1 and the tray 2, and the electret
  • the body 3 is sealed in the sealing ring 4 between the cell culture bottle 1 and the tray 2;
  • the bottom wall area diameter of the cell culture bottle 1 is 50 mm, and the bottom wall thickness is 0.5 mm;
  • the tray 2 comprises a support arm and a bottom, the height of the support arm is 5 mm, the thickness of the support arm is 2 mm, and the thickness of the support bottom is 1.5 mm;
  • the thickness of the electret 3 is 500 ⁇ m
  • the preparation raw material of described cell culture bottle 1 is polypropylene
  • the preparation raw material of described electret 3 is polypropylene
  • the raw material for the preparation of the sealing ring 4 is biological glue.
  • the device for promoting cell attachment and growth includes a cell culture plate 1, a tray 2 supporting the cell culture plate 1, an electret 3 between the cell culture plate 1 and the tray 2, and the electret
  • the body 3 is sealed with a sealing ring 4 between the cell culture plate 1 and the tray 2;
  • the diameter of the bottom wall of the cell culture plate 1 is 20 mm, and the thickness of the bottom wall is 0.1 mm;
  • the tray 2 includes a support arm and a bottom, the height of the support arm is 3 mm, the thickness of the support arm is 0.5 mm, and the thickness of the support bottom is 0.8 mm;
  • the thickness of the electret 3 is 5 ⁇ m
  • the raw material for the preparation of the cell culture plate 1 is a fluorinated ethylene propylene copolymer
  • the preparation raw material of described electret 3 is fluorinated ethylene propylene copolymer
  • the raw material for the preparation of the sealing ring 4 is medical adhesive tape.
  • Embodiment 1 The only difference from Embodiment 1 is that the thickness of the electret 3 in this embodiment is 1 ⁇ m, and other materials are the same as in Embodiment 1.
  • Example 1 The only difference from Example 1 is that the raw material for the cell culture dish 1 in this example is polyethylene, and the rest of the materials are the same as in Example 1.
  • Example 1 The only difference from Example 1 is that the raw material for preparing the electret 3 in this example is polytetrafluoroethylene, and the rest of the materials are the same as in Example 1.
  • Embodiment 1 The only difference from Embodiment 1 is that the raw material for the sealing ring 4 in this embodiment is silica gel, and the rest of the materials are the same as in Embodiment 1.
  • Example 1 The only difference from Example 1 is that the thickness of the bottom wall of the cell culture dish 1 in this comparative example is 0.05 mm, and other materials are the same as in Example 1.
  • Example 1 The only difference from Example 1 is that the thickness of the bottom wall of the cell culture dish 1 in this comparative example is 1 mm, and other materials are the same as in Example 1.
  • Example 1 The only difference from Example 1 is that no electret 3 is placed between the cell culture dish 1 and the tray 2 in this comparative example, and the rest of the materials are the same as in Example 1.
  • HT1080 human fibrosarcoma cells were cultured, and the culture steps were as follows:
  • Example 1 Adjusts the grid voltage of the polarization device to the corresponding voltage value, specifically: Example 1, Examples 3-7 and Comparative Examples 1-3 are 1 kV, and Example 2 is 2 kV;
  • the electret 3 is placed between the cell culture equipment and the tray 2 and sealed with a sealing ring 4 .
  • Adherence rate (%) number of adherent cells/number of inoculated cells ⁇ 100%.
  • the bottom wall of the culture dish in comparative example 2 was thicker, which affected the effect of the electret on the cells, and the cell adhesion rate was significantly affected, which was only 32%; the bottom wall of the culture dish in comparative example 1 was thin, but the cells The adherence rate of the culture dish did not increase significantly, indicating that when the thickness of the bottom wall was less than 0.8 mm, the effect of the electret on promoting cell adhesion was relatively stable, but too thin a bottom wall affected the firmness of the culture dish.
  • the cell culture in Comparative Example 3 was not affected by the electric field of the electret, and the cell adhesion rate was less than 10%.
  • the present invention provides a device for promoting cell attachment and growth.
  • the electric field promotes cell attachment and proliferation.
  • the device has a simple structure, high production efficiency, strong applicability, convenient use, and broad application prospects.
  • the present invention illustrates the detailed methods of the present invention through the above-mentioned examples, but the present invention is not limited to the above-mentioned detailed methods, that is, it does not mean that the present invention can only be implemented depending on the above-mentioned detailed methods.
  • Those skilled in the art should understand that any improvement of the present invention, the equivalent replacement of each raw material of the product of the present invention, the addition of auxiliary components, the selection of specific methods, etc., all fall within the scope of protection and disclosure of the present invention.

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Abstract

一种促进细胞贴壁和生长的装置及其使用方法，所述装置包括：细胞培养器材（1）、支撑所述细胞培养器材的托盘（2）、位于所述细胞培养器材（1）和托盘（2）之间的驻极体（3）以及将所述驻极体密封在细胞培养器材（1）和托盘（2）之间的密封圈（4）；所述细胞培养器材（1）的底壁厚度为0.1～0.5mm。所述促进细胞贴壁和生长的装置结构简单，生产效率高，使用方便，不改变培养基的组分，对细胞的生理状态影响较小，应用前景广阔。

Description

一种促进细胞贴壁和生长的装置及其使用方法 技术领域
本发明属于细胞培养技术领域，尤其涉及一种促进细胞贴壁和生长的装置及其使用方法。
 背景技术
贴壁型细胞需要粘附在培养装置表面才能进行迁移、增殖和分化等。培养贴壁型细胞时一般通过在培养装置表面包被促贴壁基质的方式，增强其细胞膜表面的整联蛋白（受体）与促贴壁基质中的粘附蛋白（配体）间的相互作用，提高细胞的贴壁率，或者在含有吸附细胞功能的微结构的培养装置中进行培养。
CN112458048A公开了一种牙髓干细胞原代培养促贴壁基质及原代牙髓干细胞的培养方法，该发明提供了一种可促进细胞贴壁生长的材料，用其包被培养皿，为牙髓干细胞原代培养、建立成熟的体外培养体系并且为后续实验研究提供了优质及丰富的细胞来源。该发明在牙髓干细胞原代培养时，使用多酚蛋白作为促贴壁试剂预包被培养皿，增加了组织块细胞爬出率及原代细胞的数量，缩短了原代制备时间，提升了原代培养的成功率。但添加多酚蛋白会改变培养基的组分，影响细胞正常的生理代谢。此外，该方法仅对牙髓干细胞的培养具有促进作用，适用性较差。
目前，通过添加促贴壁基质或使用具有细胞吸附微结构的培养装置来促进贴壁细胞的贴壁与生长往往存在改变培养基成分、适应性较差以及制造难度大的问题。因此，如何提供一种可以促进细胞贴壁及增殖的产品及相应的方法，可以有效解决上述问题，且对不同种类的细胞具有良好的适用性，已成为亟待解决的问题。
 技术问题
针对现有技术的不足和实际需求，本发明提供一种促进细胞贴壁和生长的装置及其使用方法，该装置通过电场作用来促进细胞的贴壁，不需要使用促贴壁基质包被培养器材，也无需使用特殊的微结构来吸附细胞，对细胞的生理状态影响较小，应用价值较高。
 技术解决方案
为达此目的，本发明采用如下技术方案：
第一方面，本发明提供了一种促进细胞贴壁和生长的装置，所述装置包括：
细胞培养器材、支撑所述细胞培养器材的托盘、位于所述细胞培养器材和托盘之间的驻极体以及将所述驻极体密封在细胞培养器材和托盘之间的密封圈；
所述细胞培养器材的底壁厚度为0.1~0.5 mm，例如可以是0.1 mm、0.2 mm、0.3 mm、0.4 mm或0.5 mm等，该数值范围内的其他具体点值均可选择，在此便不再一一赘述。
本发明中，通过设置预先极化好的驻极体，可以产生稳定的电场，从而促进贴壁细胞的贴壁，进而促进细胞的增殖，细胞的增长速率更快，培养效果更好。这一过程并未向培养基中添加促贴壁基质或使用相应的基质包被细胞培养装置，也未借助于具有特殊微结构的装置，使用方便，容易生产。
本发明中，细胞培养器材的底壁厚度在0.1~0.5 mm的范围内可以达到较好的促贴壁以及促进细胞增殖的效果，若细胞培养器材的底壁厚度小于0.1 mm，会因受到的电场强度过大从而影响细胞正常的生理活动，若细胞培养器材的底壁厚度大于0.5 mm，则会因受到的电场强度过小无法产生促进细胞贴壁、增殖的效果。
优选地，所述细胞培养器材包括细胞培养瓶、细胞培养皿或细胞培养板中的任意一种或至少两种的组合。
优选地，所述托盘包括托臂和托底。
优选地，所述托臂的高度为3~5 mm，例如可以是3 mm、3.2 mm、3.4 mm、3.6 mm、3.8 mm、4 mm、4.2 mm、4.4 mm、4.6 mm、4.8 mm或5 mm等，所述托臂的厚度为0.5~2 mm，例如可以是0.5 mm、0.6 mm、0.7 mm、0.8 mm、0.9 mm、1 mm、1.1 mm、1.2 mm、1.3 mm、1.4 mm、1.5 mm、1.6 mm、1.7 mm、1.8 mm、1.9 mm或2 mm等，该数值范围内的其他具体点值均可选择，在此便不再一一赘述。
优选地，所述托底的厚度为0.8~1.5 mm，例如可以是0.8 mm、0.9 mm、1 mm、1.1 mm、1.2 mm、1.3 mm、1.4 mm或1.5 mm等，该数值范围内的其他具体点值均可选择，在此便不再一一赘述。
优选地，所述驻极体的厚度为5~500 μm，例如可以是5 μm、10 μm、50 μm、100 μm、150 μm、200 μm、250 μm、300 μm、350 μm、400 μm、450 μm或500 μm等，该数值范围内的其他具体点值均可选择，在此便不再一一赘述。
本发明中，驻极体的厚度可根据所用驻极体材料性能、细胞培养环境等条件进行确定。
优选地，所述细胞培养器材的制备原料包括聚丙烯（PP）、聚苯乙烯（PS）、聚四氟乙烯（PTFE）、氟化乙丙烯共聚物（FEP）或聚三氟氯乙烯（PCTFE）中的任意一种或至少两种的组合。
本发明中，所述细胞培养器材由生物相容性良好的聚合材料制成，其大小和尺寸可根据具体的实际情况进行确定。
优选地，所述驻极体的制备原料包括聚丙烯和/或氟化乙丙烯共聚物。
本发明中，驻极体是一种呈现“准永久”电荷的电介质材料。驻极体中的电荷可以是“真实”电荷，如储存在材料表面的表面电荷和储存在材料体内的空间电荷；也可以是偶极电荷，即取向偶极子（或位移电荷）；或两者共有之。本发明中选用生物相容性好、电荷储存能力较好的驻极体。
本发明中，所述驻极体的大小可根据具体的实际情况进行确定。
优选地，所述密封圈的制备原料包括生物胶和/或医用胶布。
本发明中，密封圈选用密封性、防水性以及粘性均较好的材料，保证密封在细胞培养器材和托盘之间的驻极体不受潮湿等环境因素的影响，其大小可根据具体的实际情况进行确定。
作为优选技术方案，本发明所述促进细胞贴壁和生长的装置，包括：
细胞培养器材、支撑所述细胞培养器材的托盘、位于所述细胞培养器材和托盘之间的驻极体以及密封所述驻极体的密封圈；
所述细胞培养器材包括细胞培养瓶、细胞培养皿或细胞培养板中的任意一种或至少两种的组合，所述细胞培养器材的底壁厚度为0.1~0.5 mm；
所述托盘包括托臂和托底，所述托臂的高度为3~5 mm，所述托臂的厚度为0.5~2 mm，所述托底的厚度为0.8~1.5 mm；
所述驻极体的厚度为5~500 μm；
所述细胞培养器材的制备原料包括聚丙烯、聚苯乙烯、聚四氟乙烯、氟化乙丙烯共聚物或聚三氟氯乙烯中的任意一种或至少两种的组合；
所述驻极体的制备原料包括聚丙烯和/或氟化乙丙烯共聚物；
所述密封圈的制备原料包括生物胶和/或医用胶布。
第二方面，本发明提供了一种第一方面所述的促进细胞贴壁和生长的装置的使用方法，所述使用方法包括：
对驻极体进行极化，静置；
向灭菌后的细胞培养器材中接种细胞；
将驻极体放置在细胞培养器材与托盘之间，使用密封圈密封；
将装置放置在培养环境中，培养细胞。
本发明中，所述促进细胞贴壁和生长的装置使用方便，操作简单，成功率高，容错性好，促进了相关装置的推广与使用。
优选地，所述极化前还包括对驻极体进行清洁的步骤。
优选地，所述清洁包括超声清洗。
优选地，所述清洁后还包括干燥驻极体的步骤。
优选地，所述极化的方法包括电晕充电法和/或电击穿充电法。
优选地，所述静置的时间为不少于24 h，例如可以是24 h、25 h、26 h、27 h、28 h、29 h或30 h等，该数值范围内的其他具体点值均可选择，在此便不再一一赘述。
本发明中，静置的作用在于保证驻极体所储存的电荷稳定，从而给细胞提供一个稳定的电场环境。
优选地，所述使用方法还包括更换培养基和/或驻极体的步骤。
本发明中，更换培养基和/或驻极体并非是必须要进行的操作，可以根据具体的细胞生长状态（如细胞形态、细胞密度等）来确定，如达到要求，则不需要更换培养基和/或驻极体；如没达到要求，则需要更换培养基和/或驻极体。通常而言，当细胞传代周期远小于驻极体对细胞的作用时间时，需要定期更换培养基，以保证细胞生长所需的养分；当细胞传代周期远大于驻极体对细胞的作用时间时，需要定期更换驻极体，以保证驻极体对细胞生长的持续作用。
作为优选技术方案，本发明所述促进细胞贴壁和生长的装置的使用方法，包括以下步骤：
对驻极体进行超声清洗，干燥后，通过电晕充电法和/或电击穿充电法对驻极体进行极化，静置不少于24 h；
向灭菌后的细胞培养器材中接种细胞；
将驻极体放置在细胞培养器材与托盘之间，使用密封圈密封；
将装置放置在培养环境中，培养细胞；
根据细胞的生长状态，更换培养基和/或驻极体，继续培养。
具体的使用操作流程图如图1所示：
裁剪大小、厚度合适的驻极体并进行清洁；
通过电晕充电法和/或电击穿充电法对驻极体进行极化，放置至少24 h；
向消毒灭菌后的细胞培养器材中接种细胞，例如可以是向细胞培养皿中加入含有待培养细胞的培养基；
在细胞培养器材和托盘之间放置驻极体，并使用密封圈密封；
将整个装置放置在适合细胞生长的温度、湿度的环境中，培养细胞；
细胞培养8~48小时后，观察细胞的生长状态是否达到了要求，若细胞状态满足相关的要求，则可以收集细胞，细胞培养结束；若细胞状态未满足相关的要求，则更换培养基和驻极体，密封后继续培养。
 有益效果
本发明所述促进细胞贴壁和生长的装置结构简单，适用性强，通过驻极体产生的电场来促进细胞的贴壁与增殖，无需借助复杂的培养设备或添加促贴壁基质，对细胞的生理状态影响较小，不影响后续的实验操作，获得的实验结果也更加准确；通过对装置的制备原料、底壁厚度等相关参数进行优化，进一步提高了细胞的贴壁率，细胞贴壁率在40%~50%之间；所述装置制备简单高效，使用方便灵活，具有应用于实际科研工作中的价值，应用前景广阔。
 附图说明
为了更清楚地说明本申请实施例中的技术方案，下面将对实施例或现有技术描述中所需要使用的附图作简单地介绍，显而易见地，下面描述中的附图仅仅是本申请的一些实施例，对于本领域普通技术人员来讲，在不付出创造性劳动的前提下，还可以根据这些附图获得其他的附图。
图1是本发明所述促进细胞贴壁和生长的装置的使用操作流程图；
图2为本发明实施例1中的促进细胞贴壁和生长的装置的结构示意图（图中，1-细胞培养皿，2-托盘，3-驻极体，4-密封圈）；
图3为本发明实施例2中的促进细胞贴壁和生长的装置的结构示意图（图中，1-细胞培养瓶，2-托盘，3-驻极体，4-密封圈）；
图4为本发明实施例3中的促进细胞贴壁和生长的装置的结构示意图（图中，1-细胞培养板，2-托盘，3-驻极体，4-密封圈）。
 本发明的实施方式
为进一步阐述本发明所采取的技术手段及其效果，以下结合实施例和附图对本发明作进一步地说明。可以理解的是，此处所描述的具体实施方式仅仅用于解释本发明，而非对本发明的限定。
实施例中未注明具体技术或条件者，按照本领域内的文献所描述的技术或条件，或者按照产品说明书进行。所用试剂或仪器未注明生产厂商者，均为可通过正规渠道商购获得的常规产品。
材料：
细胞培养皿、细胞培养瓶和细胞培养板购自泰州鑫联诚润生物技术有限公司，均为未经包被处理的产品。
驻极体购自上海驻极新材料有限公司。
生物胶购自深圳市康利邦科技有限公司。
医用胶带购自深圳市大森生物科技有限公司。
HT1080人纤维肉瘤细胞来自购自武汉普诺赛生命科技有限公司。
DMEM培养基和FBS购自赛默飞世尔科技（中国）有限公司。
实施例1
本实施例提供一种促进细胞贴壁和生长的装置，其结构图如图2所示。所述促进细胞贴壁和生长的装置包括细胞培养皿1、支撑所述细胞培养皿1的托盘2、位于所述细胞培养皿1和托盘2之间的驻极体3以及将所述驻极体3密封在细胞培养皿1和托盘2之间的密封圈4；
所述细胞培养皿1的底壁直径为35 mm，底壁厚度为0.3 mm；
所述托盘2包括托臂和托底，所述托臂的高度为4 mm，所述托臂的厚度为1 mm，所述托底的厚度为1 mm；
所述驻极体3的厚度为30 μm；
所述细胞培养皿1的制备原料为聚苯乙烯；
所述驻极体3的制备原料为氟化乙丙烯共聚物；
所述密封圈4的制备原料为医用胶布。
实施例2
本实施例提供一种促进细胞贴壁和生长的装置，其结构图如图3所示。所述促进细胞贴壁和生长的装置包括细胞培养瓶1、支撑所述细胞培养瓶1的托盘2、位于所述细胞培养瓶1和托盘2之间的驻极体3以及将所述驻极体3密封在细胞培养瓶1和托盘2之间的密封圈4；
所述细胞培养瓶1的底壁面积直径为50 mm，底壁厚度为0.5 mm；
所述托盘2包括托臂和托底，所述托臂的高度为5 mm，所述托臂的厚度为2 mm，所述托底的厚度为1.5 mm；
所述驻极体3的厚度为500 μm；
所述细胞培养瓶1的制备原料为聚丙烯；
所述驻极体3的制备原料为聚丙烯；
所述密封圈4的制备原料为生物胶。
实施例3
本实施例提供一种促进细胞贴壁和生长的装置，其结构图如图4所示。所述促进细胞贴壁和生长的装置包括细胞培养板1、支撑所述细胞培养板1的托盘2、位于所述细胞培养板1和托盘2之间的驻极体3以及将所述驻极体3密封在细胞培养板1和托盘2之间的密封圈4；
所述细胞培养板1的底壁直径为20 mm，底壁厚度为0.1 mm；
所述托盘2包括托臂和托底，所述托臂的高度为3 mm，所述托臂的厚度为0.5 mm，所述托底的厚度为0.8 mm；
所述驻极体3的厚度为5 μm；
所述细胞培养板1的制备原料为氟化乙丙烯共聚物；
所述驻极体3的制备原料为氟化乙丙烯共聚物；
所述密封圈4的制备原料为医用胶布。
实施例4
与实施例1的区别仅在于，本实施例中驻极体3的厚度为1μm，其余材料与实施例1相同。
实施例5
与实施例1的区别仅在于，本实施例中细胞培养皿1的制备原料为聚乙烯，其余材料与实施例1相同。
实施例6
与实施例1的区别仅在于，本实施例中驻极体3的制备原料为聚四氟乙烯，其余材料与实施例1相同。
实施例7
与实施例1的区别仅在于，本实施例中密封圈4的制备原料为硅胶，其余材料与实施例1相同。
对比例1
与实施例1的区别仅在于，本对比例中细胞培养皿1的底壁厚度为0.05 mm，其余材料与实施例1相同。
对比例2
与实施例1的区别仅在于，本对比例中细胞培养皿1的底壁厚度为1 mm，其余材料与实施例1相同。
对比例3
与实施例1的区别仅在于，本对比例中细胞培养皿1和托盘2之间不放置驻极体3，其余材料与实施例1相同。
贴壁率检测
使用实施例1~7以及对比例1~3制备的促进细胞贴壁和生长的装置，对HT1080人纤维肉瘤细胞进行培养，培养步骤如下：
（1）对驻极体3进行超声清洗，干燥后，通过电晕充电法对驻极体3进行极化，静置24 h；
所述电晕充电法的具体步骤如下：
①清洁后的驻极体3薄膜放置在极化装置底座上；
②将极化装置的栅网电压调节到相应的电压值，具体为：实施例1、实施例3~7以及对比例1~3为1 kV，实施例2为2 kV；
③将极化装置的极化针电压调节到相应的电压值，具体为：实施例1、实施例3~7以及对比例1~3为25 kV，实施例2为20 kV；
④秒表计时30 s，结束后，关掉极化针电压、栅网电压，取出所极化的驻极体3。
（2）向灭菌后的细胞培养器材中接种HT1080人纤维肉瘤细胞，使用的培养基为体积分数为1%双抗和10%FBS的DMEM培养基。
（3）将驻极体3放置在细胞培养器材与托盘2之间，使用密封圈4密封。
（4）将装置放置在37℃、CO2浓度为5%的培养环境中，培养细胞。
贴壁率计算
培养细胞48 h后，去除培养液上的清液，加入含0.1%结晶紫染液（PBS溶液配制）至总体积为2 mL，置于37℃恒温孵育30 min，消化，吹打让细胞从培养皿底表面脱落，于血球计数板上进行细胞计数，计算各组贴壁率。
贴壁率(%)=贴壁细胞数/接种细胞数×100%。
结果显示，实施例1~3中的细胞贴壁率在40%~50%之间，比普通培养皿有明显提高，细胞生长状态良好；
与实施例1~3进行比较，实施例4中驻极体的厚度偏小，产生的电场强度较小，贴壁率略有下降；实施例6中选用了其他的驻极体材料，对贴壁率的影响较为明显，仅为27%；实施例5中选用了聚乙烯作为培养皿的材料，细胞贴壁率无明显影响，但细胞的生长状态稍差，表明聚乙烯的生物相容性较差；实施例7中选用硅胶作为密封圈材料，对细胞贴壁无明显影响，但硅胶的价格较高，相应产品的生产成本有所提升，不利于产品的推广与使用；
对比例2中培养皿的底壁较厚，影响了驻极体对细胞的作用，细胞贴壁率受到了明显的影响，仅为32%；对比例1中培养皿底壁偏薄，但细胞的贴壁率无显著提高，说明当底壁厚度小于0.8 mm后，驻极体对细胞的促贴壁作用相对稳定，但底壁过薄影响了培养皿的结实程度，在后续的实验操作过程中易发生破裂；对比例3中的细胞培养不受驻极体的电场作用，其细胞贴壁率不到10%。
综上所述，本发明提供了一种促进细胞贴壁和生长的装置，通过在托盘和细胞培养器材之间填入预先极化好的驻极体，通过电场促进细胞的贴壁及增殖，无需添加促贴壁基质或借助具有特殊结构的培养装置，对细胞的生理状态影响较小，实验结果更加准确；装置结构简单，生产效率高，适用性强，使用方便，应用前景广阔。
申请人声明，本发明通过上述实施例来说明本发明的详细方法，但本发明并不局限于上述详细方法，即不意味着本发明必须依赖上述详细方法才能实施。所属技术领域的技术人员应该明了，对本发明的任何改进，对本发明产品各原料的等效替换及辅助成分的添加、具体方式的选择等，均落在本发明的保护范围和公开范围之内。

Claims (10)

  1. 一种促进细胞贴壁和生长的装置，其特征在于，所述装置包括：
    细胞培养器材、支撑所述细胞培养器材的托盘、位于所述细胞培养器材和托盘之间的驻极体以及将所述驻极体密封在细胞培养器材和托盘之间的密封圈；
    所述细胞培养器材的底壁厚度为0.1~0.5 mm。
  2. 根据权利要求1所述的促进细胞贴壁和生长的装置，其特征在于，所述细胞培养器材包括细胞培养瓶、细胞培养皿或细胞培养板中的任意一种或至少两种的组合；
    优选地，所述托盘包括托臂和托底；
    优选地，所述托臂的高度为3~5 mm，所述托臂的厚度为0.5~2 mm；
    优选地，所述托底的厚度为0.8~1.5 mm。
  3. 根据权利要求1或2所述的促进细胞贴壁和生长的装置，其特征在于，所述驻极体的厚度为5~500 μm。
  4. 根据权利要求1~3任一项所述的促进细胞贴壁和生长的装置，其特征在于，所述细胞培养器材的制备原料包括聚丙烯、聚苯乙烯、聚四氟乙烯、氟化乙丙烯共聚物或聚三氟氯乙烯中的任意一种或至少两种的组合；
    优选地，所述驻极体的制备原料包括聚丙烯和/或氟化乙丙烯共聚物；
    优选地，所述密封圈的制备原料包括生物胶和/或医用胶布。
  5. 根据权利要求1~4任一项所述的促进细胞贴壁和生长的装置，其特征在于，所述装置包括：
    细胞培养器材、支撑所述细胞培养器材的托盘、位于所述细胞培养器材和托盘之间的驻极体以及密封所述驻极体的密封圈；
    所述细胞培养器材包括细胞培养瓶、细胞培养皿或细胞培养板中的任意一种或至少两种的组合，所述细胞培养器材的底壁厚度为0.1~0.5 mm；
    所述托盘包括托臂和托底，所述托臂的高度为3~5 mm，所述托臂的厚度为0.5~2 mm，所述托底的厚度为0.8~1.5 mm；
    所述驻极体的厚度为5~500 μm；
    所述细胞培养器材的制备原料包括聚丙烯、聚苯乙烯、聚四氟乙烯、氟化乙丙烯共聚物或聚三氟氯乙烯中的任意一种或至少两种的组合；
    所述驻极体的制备原料包括聚丙烯和/或氟化乙丙烯共聚物；
    所述密封圈的制备原料包括生物胶和/或医用胶布。
  6. 一种权利要求1~5任一项所述的促进细胞贴壁和生长的装置的使用方法，其特征在于，所述使用方法包括：
    对驻极体进行极化，静置；
    向灭菌后的细胞培养器材中接种细胞；
    将驻极体放置在细胞培养器材与托盘之间，使用密封圈密封；
    将装置放置在培养环境中，培养细胞。
  7. 根据权利要求6所述的促进细胞贴壁和生长的装置的使用方法，其特征在于，所述极化前还包括对驻极体进行清洁的步骤；
    优选地，所述清洁包括超声清洗；
    优选地，所述清洁后还包括干燥驻极体的步骤。
  8. 根据权利要求6或7所述的促进细胞贴壁和生长的装置的使用方法，其特征在于，所述极化的方法包括电晕充电法和/或电击穿充电法；
    优选地，所述静置的时间为不少于24 h。
  9. 根据权利要求6~8任一项所述的促进细胞贴壁和生长的装置的使用方法，其特征在于，所述使用方法还包括更换培养基和/或驻极体的步骤。
  10. 根据权利要求6~9任一项所述的促进细胞贴壁和生长的装置的使用方法，其特征在于，所述使用方法包括：
    对驻极体进行超声清洗，干燥后，通过电晕充电法和/或电击穿充电法对驻极体进行极化，静置不少于24 h；
    向灭菌后的细胞培养器材中接种细胞；
    将驻极体放置在细胞培养器材与托盘之间，使用密封圈密封；
    将装置放置在培养环境中，培养细胞；
    根据细胞的生长状态，更换培养基和/或驻极体，继续培养。
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Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH02154685A (ja) * 1988-12-06 1990-06-14 Toray Ind Inc 細胞および微生物培養担体
CN111511891A (zh) * 2017-10-19 2020-08-07 皇家飞利浦有限公司 细胞保存或培养装置
CN111933452A (zh) * 2020-07-28 2020-11-13 深圳先进技术研究院 一种驻极体的封装方法以及驻极体封装组件
CN112522100A (zh) * 2020-12-04 2021-03-19 中国科学院深圳先进技术研究院 一种细胞体外培养装置
CN214193294U (zh) * 2020-12-04 2021-09-14 中国科学院深圳先进技术研究院 一种体外细胞培养装置
CN113699045A (zh) * 2021-08-27 2021-11-26 中国科学院深圳先进技术研究院 一种促进细胞贴壁和生长的装置及其使用方法

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
AU1684595A (en) * 1994-01-21 1995-08-08 Brown University Research Foundation Biocompatible implants
CN1634606A (zh) * 2003-12-29 2005-07-06 唐耀明 外敷型医用止痛速愈驻极体膜及其制造方法
CN203833942U (zh) * 2014-04-18 2014-09-17 广东省医学实验动物中心 细胞培养器皿

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPH02154685A (ja) * 1988-12-06 1990-06-14 Toray Ind Inc 細胞および微生物培養担体
CN111511891A (zh) * 2017-10-19 2020-08-07 皇家飞利浦有限公司 细胞保存或培养装置
CN111933452A (zh) * 2020-07-28 2020-11-13 深圳先进技术研究院 一种驻极体的封装方法以及驻极体封装组件
CN112522100A (zh) * 2020-12-04 2021-03-19 中国科学院深圳先进技术研究院 一种细胞体外培养装置
CN214193294U (zh) * 2020-12-04 2021-09-14 中国科学院深圳先进技术研究院 一种体外细胞培养装置
CN113699045A (zh) * 2021-08-27 2021-11-26 中国科学院深圳先进技术研究院 一种促进细胞贴壁和生长的装置及其使用方法

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
"Doctoral Dissertation", 1 May 2018, CHINESE PEOPLE'S LIBERATION ARMY NAVAL MILITARY MEDICAL UNIVERSITY, CN, article YUAN, WANG: "Effect of Electret 5-fluorouracil Patch on Inhibition of Hypertrophic Scar in Rat Skin and Its Mechanism", pages: 1 - 142, XP009544017 *
HAN, YIHUA: "Design and Application of Permanent Electroacupuncture", TEACHING APPARATUS AND EXPERIMENT, vol. 1, no. 3, 31 December 1984 (1984-12-31), pages 118 - 120, XP009543830, ISSN: 1003-3416 *

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