WO2021169600A1 - Surface modification method for cell culture apparatus, and cell culture apparatus - Google Patents

Surface modification method for cell culture apparatus, and cell culture apparatus Download PDF

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Publication number
WO2021169600A1
WO2021169600A1 PCT/CN2020/142267 CN2020142267W WO2021169600A1 WO 2021169600 A1 WO2021169600 A1 WO 2021169600A1 CN 2020142267 W CN2020142267 W CN 2020142267W WO 2021169600 A1 WO2021169600 A1 WO 2021169600A1
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Prior art keywords
cell culture
gas
culture device
surface modification
predetermined conditions
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PCT/CN2020/142267
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French (fr)
Chinese (zh)
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李慧伦
方想元
袁晔
袁建华
周志智
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广州洁特生物过滤股份有限公司
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Publication of WO2021169600A1 publication Critical patent/WO2021169600A1/en

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    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J7/00—Chemical treatment or coating of shaped articles made of macromolecular substances
    • C08J7/12—Chemical modification
    • C08J7/123—Treatment by wave energy or particle radiation
    • 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
    • C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
    • C—CHEMISTRY; METALLURGY
    • C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
    • C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
    • C08J2325/00—Characterised by the use of homopolymers or copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and at least one being terminated by an aromatic carbocyclic ring; Derivatives of such polymers
    • C08J2325/02—Homopolymers or copolymers of hydrocarbons
    • C08J2325/04—Homopolymers or copolymers of styrene
    • C08J2325/06—Polystyrene

Definitions

  • the present invention relates to the technical field of cell culture, in particular to a method for surface modification of a cell culture device and a cell culture device.
  • the method of improving the surface properties of plastic products through physical or chemical methods is called surface modification.
  • the main methods of commonly used surface modification of polymer materials include surface mechanical modification, surface coating modification, surface vacuum plating, sputtering, spraying and other physical modification methods; flame modification, solution treatment, discharge, radiation, Chemical modification methods such as ion plating, electroplating, graft polymerization modification, etc.
  • Most of the cells in animals and humans, such as fibroblasts, skeletal tissue (bone and cartilage), myocardial and smooth muscle, liver, lung, kidney, breast skin glial cells, etc., are large when they are cultured in vitro. Most of them grow in an adherent manner, which requires that the cell surface of the in vitro cell culture device has a certain degree of hydrophilicity in order to meet the requirements for in vitro culture of adherent cells.
  • corona discharge modification technology and plasma treatment polymer material surface modification technology are usually used to modify the surface of the material to increase the surface tension of the material and improve the cell adhesion performance.
  • plasma technology has significant advantages in hydrophilic modification of polymer materials, it still has shortcomings, such as insufficient hydrophilicity of the modified material, short hydrophilic residence time, and poor surface morphology, etc. ; These deficiencies will have extremely adverse effects on cell adhesion behavior.
  • the gas source used in plasma surface treatment technology can be divided into two categories: inert gas and reactive gas.
  • the hydrophilic effect of the surface of the polymer material treated by these two gases is obviously different, mainly in the maintenance of hydrophilicity.
  • the length of time The surface of the material treated with reactive plasma gas (such as oxygen) has strong hydrophilicity, but the time for maintaining hydrophilicity is very short; compared with reactive plasma treatment, the surface of the material treated with inert plasma gas (such as argon) is hydrophilic.
  • the performance of water capacity is weak, and the time to maintain hydrophilicity is longer.
  • a method for surface modification of a cell culture device including the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment (1 min) on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface;
  • the second ion treatment using inert gas and reactive plasma gas as initiating substances, plasma etching treatment (3 min) is performed on the substrate under predetermined conditions.
  • the inert initiating gas in step S1 is argon.
  • the volume ratio of the inert gas to the reactive plasma gas in step S2 is 3:1.
  • the reactive plasma gas is itself capable of providing oxygen-containing groups or undergoing chemical bond cleavage with the substrate material and then reacting to generate oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the predetermined conditions for the first ion treatment in step S1 are as follows:
  • the predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-50mT, temperature 30°C-35°C, gas flow rate 20sccm-50sccm, processing power 100w-250w, processing time 300s- 800s.
  • the predetermined conditions for the second ion treatment in step S2 are as follows:
  • the predetermined conditions for the second ion treatment in step S2 are as follows:
  • a cell culture device includes a cell culture scaffold which is modified by the cell culture device surface modification method to obtain a long-term hydrophilic modified surface.
  • the surface modification method of the cell culture device of the present invention can improve the ability of long-term hydrophilic modification on the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and realize serum-free culture to reduce users Low cost and easy to realize industrialization.
  • the hydrophilicity of the substrate surface is enhanced, the contact angle is below 30°, and after 3 months of accelerated aging test, the hydrophilicity of the substrate surface does not decline.
  • the surface of the material is modified with an inert initiating gas, and then a certain proportion of inert gas and reactive plasma gas is used to further modify the material, which will help realize the effect of long-term hydrophilic modification.
  • Cells that are difficult to adhere to culture in vitro can grow well on the basal surface treated by the method of the present invention, and can meet the low serum and serum-free culture of conventional cells and refractory cells.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface, and the plasma etching treatment time is 1 min.
  • the predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 5sccm-80sccm, processing power 60w-350w, and processing time greater than 240s.
  • the inert initiating gas in step S1 is one of inert gases.
  • the second ion treatment using inert gas and reactive plasma gas as initiating substances, plasma etching treatment is performed on the substrate under predetermined conditions, and the plasma etching treatment time is 3 minutes.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 20sccm-180sccm, processing power 60w-350w, and processing time 100s-1200s.
  • the volume ratio of the inert gas to the reactive plasma gas is preferably 3:1.
  • Reactive plasma gas is a compound that can provide oxygen-containing groups by itself or break chemical bonds with the substrate material and then react to generate oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the surface modification method of the cell culture device of the present invention can improve the ability of long-term hydrophilic modification on the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and realize serum-free culture to reduce users Low cost and easy to realize industrialization.
  • the hydrophilicity of the substrate surface is enhanced, the contact angle is below 30°, and after 3 months of accelerated aging test, the hydrophilicity of the substrate surface does not decline.
  • the surface of the material is modified with an inert initiating gas, and then a certain proportion of inert gas and reactive plasma gas is used to further modify the material, which will help to achieve the effect of long-term hydrophilic modification.
  • Cells that are difficult to adhere to culture in vitro can grow well on the basal surface treated by the method of the present invention, and can meet the low serum and serum-free culture of conventional cells and refractory cells.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 5sccm-80sccm, processing power 60w-350w, and processing time greater than 240s.
  • the inert initiating gas in step S1 is preferably argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 20sccm-180sccm, processing power 60w-350w, and processing time 100s-1200s.
  • Reactive plasma gas is a compound that can provide oxygen-containing groups by itself or break chemical bonds with the substrate material and then react to generate oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the predetermined conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 20 mT, a temperature of 30° C., a gas flow rate of 80 sccm, a processing power of 350 w, and a processing time of 300 s.
  • the initiating gas in step S1 is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 20 mT, a temperature of 30° C., a gas flow rate of 180 sccm, a processing power of 350 w, and a processing time of 100 s.
  • the reactive plasma gas is a compound that itself can provide oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the predetermined conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 150 mT, a temperature of 55° C., a gas flow rate of 5 sccm, a processing power of 60 w, and a processing time of 240 s.
  • the inert initiating gas is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 150 mT, a temperature of 55° C., a gas flow rate of 20 sccm, a processing power of 60 w, and a processing time of 1200 s.
  • the reactive plasma gas is a compound that itself can break the chemical bond with the substrate material and then react to generate oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the predetermined conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 30 mT, a temperature of 40° C., a gas flow rate of 60 sccm, a processing power of 250 w, and a processing time of 300 s.
  • the inert initiating gas is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 30 mT, a temperature of 40° C., a gas flow rate of 100 sccm, a processing power of 250 w, and a processing time of 300 s.
  • the reactive plasma gas is a compound that itself can provide oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the process conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT, temperature 30°C, gas flow rate 20sccm, processing power 250w, and processing time 800s.
  • the inert initiating gas is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: vacuum degree 50mT, temperature 40°C, gas flow rate 140sccm, processing power 200w, and processing time 800s.
  • the reactive plasma gas is a compound that itself can provide oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the process conditions for the first ion treatment in step S1 are as follows: vacuum degree 50mT, temperature 35°C, gas flow rate 50sccm, processing power 100w, and processing time 300s.
  • the inert initiating gas is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 100 mT, a temperature of 50° C., a gas flow rate of 160 sccm, a processing power of 100 w, and a processing time of 500 s.
  • the reactive plasma gas is a compound that itself can break the chemical bond with the substrate material and then react to generate oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a method for surface modification of a cell culture device.
  • a method for surface modification of a cell culture device includes the following steps:
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the process conditions for the first ion treatment in step S1 are as follows: vacuum degree 30mT, temperature 30°C, gas flow rate 40sccm, processing power 150w, and processing time 500s.
  • the inert initiating gas is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 700 mT, a temperature of 45° C., a gas flow rate of 150 sccm, a processing power of 150 w, and a processing time of 600 s.
  • the reactive plasma gas is a compound that itself can provide oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • This embodiment provides a cell culture device.
  • a cell culture device includes a cell culture scaffold, the cell culture scaffold is modified by the cell culture device surface modification method described in any one of Embodiments 3-7 to obtain a super-hydrophilic cell growth surface.
  • This embodiment provides a cell culture device.
  • a cell culture device includes a cell culture scaffold, which is modified by the following cell culture device surface modification method to obtain a super-hydrophilic cell growth surface.
  • the first ion treatment the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface.
  • the process conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 30 mT, a temperature of 30° C., a gas flow rate of 40 sccm, a processing power of 150 w, and a processing time of 500 s.
  • the inert initiating gas is argon.
  • the second ion treatment using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions.
  • the predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 700 mT, a temperature of 45° C., a gas flow rate of 150 sccm, a processing power of 150 w, and a processing time of 600 s.
  • Reactive plasma gas is a compound that can provide oxygen-containing groups by itself or break chemical bonds with the substrate material and then react to generate oxygen-containing groups.
  • the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  • the volume ratio of the inert gas to the reactive plasma gas is 3:1.
  • X-polystyrene photoelectron energy is performed on the O and C elements on the surface of the untreated polystyrene substrate, the plasma hydrophilic treatment polystyrene substrate, and the surface modification method of the cell culture device of the present invention. Spectrum scan, the analysis results are shown in Table 1.
  • the surface of the untreated polystyrene substrate, the plasma hydrophilic treated polystyrene substrate, and the surface modification method of the cell culture device of the present invention were used for atomic force microscope analysis.
  • the surface of the untreated polystyrene substrate was Some dust particles adsorbed on the surface during sample preparation are affected, and the surface is extremely smooth and flat. However, the surface becomes rougher after the first step of the conventional hydrophilic treatment, which indicates that the surface modification method of the cell culture device of the present invention can improve the long-term hydrophilic modification ability of the surface of the polystyrene material. Realize the effect of long-term hydrophilic modification.
  • the cell culture device surface modification method of the present invention can improve the ability of long-term hydrophilic modification of the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and realize Serum culture can reduce the cost of users and is easy to realize industrialization.
  • the hydrophilicity of the substrate surface is enhanced, the contact angle is below 30°, and after 3 months of accelerated aging test, the hydrophilicity of the substrate surface does not decline.
  • the surface of the material is modified with an inert initiating gas, and then a certain proportion of inert gas and reactive plasma gas is used to further modify the material, which will help realize the effect of long-term hydrophilic modification.
  • Cells that are difficult to adhere to culture in vitro can grow well on the basal surface treated by the method of the present invention, and can meet the low serum and serum-free culture of conventional cells and refractory cells.

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Abstract

Disclosed in the present invention are a surface modification method for a cell culture apparatus, and a cell culture apparatus. The surface modification method for a cell culture apparatus comprises the following steps: S1, primary ion treatment: an inert initiating gas performing plasma etching treatment on a surface of a substrate under a pre-set condition to cause the surface of the substrate to form a long-lasting hydrophilic modified surface; S2, secondary ion treatment: using an inert gas and a reactive plasma as initiating material to perform plasma etching treatment on the substrate under a pre-set condition. The cell culture apparatus surface modification method can effectively improve surface hydrophilicity and achieve a long-lasting hydrophilicity effect.

Description

细胞培养装置表面改性方法及细胞培养装置Surface modification method of cell culture device and cell culture device 技术领域Technical field
本发明涉及细胞培养技术领域，特别是涉及一种细胞培养装置表面改性方法及细胞培养装置。The present invention relates to the technical field of cell culture, in particular to a method for surface modification of a cell culture device and a cell culture device.
背景技术Background technique
通过物理或化学方法改善塑料制品表面的性能的方法称为表面改性。常用的高分子材料表面改性的主要方法包括表面机械改性、表面涂覆改性、表面真空镀、溅射、喷射等物理改性方法；火焰改性、溶液处理、放电、射线辐照、离子镀、电镀、接枝聚合改性等化学改性方法。而动物和人体内的大部分细胞，比如成纤维细胞，骨骼组织(骨及软骨)，心肌与平滑肌、肝、肺、肾、乳腺皮肤神经胶质细胞等，当离体置于体外培养时大多数均以贴壁方式生长，这就必须要求体外细胞培养装置细胞表面具有一定的亲水性，才能满足贴壁型细胞体外培养要求。The method of improving the surface properties of plastic products through physical or chemical methods is called surface modification. The main methods of commonly used surface modification of polymer materials include surface mechanical modification, surface coating modification, surface vacuum plating, sputtering, spraying and other physical modification methods; flame modification, solution treatment, discharge, radiation, Chemical modification methods such as ion plating, electroplating, graft polymerization modification, etc. Most of the cells in animals and humans, such as fibroblasts, skeletal tissue (bone and cartilage), myocardial and smooth muscle, liver, lung, kidney, breast skin glial cells, etc., are large when they are cultured in vitro. Most of them grow in an adherent manner, which requires that the cell surface of the in vitro cell culture device has a certain degree of hydrophilicity in order to meet the requirements for in vitro culture of adherent cells.
目前在生物实验室耗材领域，通常采用电晕放电改性技术和等离子体处理高分子材料表面改性技术对材料表面进行亲水改性，增大材料表面的张力，提高细胞粘附性能。虽然等离子体技术在高分子材料亲水改性方面具有显著优势，但仍存在不足之处，如改性后材料亲水的能力不够强、亲水驻留时间较短及表面形貌欠佳等；这些方面的不足，都将会对细胞粘附行为产生极不利的影响。At present, in the field of biological laboratory consumables, corona discharge modification technology and plasma treatment polymer material surface modification technology are usually used to modify the surface of the material to increase the surface tension of the material and improve the cell adhesion performance. Although plasma technology has significant advantages in hydrophilic modification of polymer materials, it still has shortcomings, such as insufficient hydrophilicity of the modified material, short hydrophilic residence time, and poor surface morphology, etc. ; These deficiencies will have extremely adverse effects on cell adhesion behavior.
一般地，用于等离子表面处理技术的气源可分为惰性与反应性气体两大类，这两种气体所处理高分子材料表面的亲水效果有较明显的差异，主要表现在亲水维持时间方面的长短。反应性等离子气体(如氧气)处理材料表面后的亲水能力较强，但是维持亲水的时间很短；相比反应性等离子体处理，惰性等离子 气体(如氩气)处理后的材料表面亲水能力表现较弱，而维持亲水的时间较长。有人提出将以上两种等离子气体按照一定的比例混合后处理材料表面，在一定程度上确实弥补了各自的不足，也明显提升了亲水能力和亲水驻留时长，但是，改性后材料的亲水能力仍然会随着时间的推移，而逐渐减弱。Generally, the gas source used in plasma surface treatment technology can be divided into two categories: inert gas and reactive gas. The hydrophilic effect of the surface of the polymer material treated by these two gases is obviously different, mainly in the maintenance of hydrophilicity. The length of time. The surface of the material treated with reactive plasma gas (such as oxygen) has strong hydrophilicity, but the time for maintaining hydrophilicity is very short; compared with reactive plasma treatment, the surface of the material treated with inert plasma gas (such as argon) is hydrophilic. The performance of water capacity is weak, and the time to maintain hydrophilicity is longer. Some people have proposed that the above two plasma gases are mixed in a certain ratio to treat the surface of the material. To a certain extent, it does make up for their respective shortcomings, and also significantly improves the hydrophilic ability and the hydrophilic residence time. However, the modified material The hydrophilic ability will still gradually weaken over time.
发明内容Summary of the invention
基于此，有必要提供一种能够有效提升高分子材料表面长效亲水改性的能力、实现长效亲水改性的效果、拓宽产品的应用范围和领域、降低用户的成本的细胞培养装置表面改性方法及细胞培养装置。Based on this, it is necessary to provide a cell culture device that can effectively improve the ability of long-term hydrophilic modification on the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and reduce user costs. Surface modification method and cell culture device.
一种细胞培养装置表面改性方法，包括如下步骤：A method for surface modification of a cell culture device, including the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理(1min)，使得所述基材的表面形成长效亲水改性表面；S1. The first ion treatment: the inert initiating gas performs plasma etching treatment (1 min) on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface;
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理(3min)。S2. The second ion treatment: using inert gas and reactive plasma gas as initiating substances, plasma etching treatment (3 min) is performed on the substrate under predetermined conditions.
在其中一个实施例中，步骤S1中所述惰性引发气体为氩气。In one of the embodiments, the inert initiating gas in step S1 is argon.
在其中一个实施例中，步骤S2中惰性气体与反应性等离子气体的体积比为3:1。In one of the embodiments, the volume ratio of the inert gas to the reactive plasma gas in step S2 is 3:1.
在其中一个实施例中，所述反应性等离子气体为本身能够提供含氧基团或者与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。In one of the embodiments, the reactive plasma gas is itself capable of providing oxygen-containing groups or undergoing chemical bond cleavage with the substrate material and then reacting to generate oxygen-containing groups.
在其中一个实施例中，所述含氧基团为羰基、羟基或者羧基。In one of the embodiments, the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
在其中一个实施例中，步骤S1中第一次离子处理时的预定条件如下：In one of the embodiments, the predetermined conditions for the first ion treatment in step S1 are as follows:
真空度20mT-150mT、温度30℃-55℃、气体流量5sccm-80sccm、处理功率60w-350w和处理时间大于240s。Vacuum degree 20mT-150mT, temperature 30℃-55℃, gas flow rate 5sccm-80sccm, processing power 60w-350w and processing time greater than 240s.
在其中一个实施例中，步骤S1中第一次离子处理时的预定条件如下：真空 度20mT-50mT、温度30℃-35℃、气体流量20sccm-50sccm、处理功率100w-250w、处理时间300s-800s。In one of the embodiments, the predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-50mT, temperature 30°C-35°C, gas flow rate 20sccm-50sccm, processing power 100w-250w, processing time 300s- 800s.
在其中一个实施例中，步骤S2中第二次离子处理时的预定条件如下：In one of the embodiments, the predetermined conditions for the second ion treatment in step S2 are as follows:
真空度20mT-150mT、温度30℃-55℃、气体流量20sccm-180sccm、处理功率60w-350w、处理时间100s-1200s。Vacuum degree 20mT-150mT, temperature 30℃-55℃, gas flow rate 20sccm-180sccm, processing power 60w-350w, processing time 100s-1200s.
在其中一个实施例中，步骤S2中第二次离子处理时的预定条件如下：In one of the embodiments, the predetermined conditions for the second ion treatment in step S2 are as follows:
真空度50mT-100mT、温度40℃-50℃、气体流量140sccm-160sccm、处理功率100w-200w、处理时间500s-800s。Vacuum degree 50mT-100mT, temperature 40℃-50℃, gas flow rate 140sccm-160sccm, processing power 100w-200w, processing time 500s-800s.
一种细胞培养装置，包括细胞培养支架，所述细胞培养支架通过所述的细胞培养装置表面改性方法进行改性，以获得长效亲水改性表面。A cell culture device includes a cell culture scaffold which is modified by the cell culture device surface modification method to obtain a long-term hydrophilic modified surface.
本发明的细胞培养装置表面改性方法能够提升高分子材料表面长效亲水改性的能力、实现长效亲水改性的效果、拓宽产品的应用范围和领域、实现无血清培养以降低用户的成本、易于实现产业化。采用本发明的细胞培养装置表面改性方法，基材表面亲水性增强，接触角达到30°以下且经过3个月的加速老化试验后，基材表面亲水性不发生衰退。本发明先用惰性引发气体对材料表面进行改性处理后，再用一定比例的惰性气体与反应性等离子气体对材料进一步进行改性，将有利于实现长效亲水改性的效果。难以体外贴壁培养的细胞能够在经本发明的方法处理后的基础表面生长良好，能够满足常规细胞和难养细胞的低血清、无血清培养。The surface modification method of the cell culture device of the present invention can improve the ability of long-term hydrophilic modification on the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and realize serum-free culture to reduce users Low cost and easy to realize industrialization. By adopting the surface modification method of the cell culture device of the present invention, the hydrophilicity of the substrate surface is enhanced, the contact angle is below 30°, and after 3 months of accelerated aging test, the hydrophilicity of the substrate surface does not decline. In the present invention, the surface of the material is modified with an inert initiating gas, and then a certain proportion of inert gas and reactive plasma gas is used to further modify the material, which will help realize the effect of long-term hydrophilic modification. Cells that are difficult to adhere to culture in vitro can grow well on the basal surface treated by the method of the present invention, and can meet the low serum and serum-free culture of conventional cells and refractory cells.
具体实施方式Detailed ways
本发明可以以许多不同的形式来实现，并不限于本文所描述的实施例。相反地，提供这些实施例的目的是使对本发明的公开内容的理解更加透彻全面。The present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the understanding of the disclosure of the present invention more thorough and comprehensive.
除非另有定义，本文所使用的所有的技术和科学术语与属于本发明的技术 领域的技术人员通常理解的含义相同。本文中在本发明的说明书中所使用的术语只是为了描述具体的实施例的目的，不是旨在于限制本发明。本文所使用的术语“和/或”包括一个或多个相关的所列项目的任意的和所有的组合。Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the technical field of the present invention. The terms used in the specification of the present invention herein are only for the purpose of describing specific embodiments, and are not intended to limit the present invention. The term "and/or" as used herein includes any and all combinations of one or more related listed items.
实施例1Example 1
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面,等离子体蚀刻处理时间为1min。步骤S1中第一次离子处理时的预定条件如下：真空度20mT-150mT、温度30℃-55℃、气体流量5sccm-80sccm、处理功率60w-350w和处理时间大于240s。步骤S1中所述惰性引发气体为惰性气体中的一种。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface, and the plasma etching treatment time is 1 min. The predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 5sccm-80sccm, processing power 60w-350w, and processing time greater than 240s. The inert initiating gas in step S1 is one of inert gases.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理，等离子体蚀刻处理时间为3min。步骤S2中第二次离子处理时的预定条件如下：真空度20mT-150mT、温度30℃-55℃、气体流量20sccm-180sccm、处理功率60w-350w、处理时间100s-1200s。步骤S2中惰性气体与反应性等离子气体的体积比优选为3:1。反应性等离子气体为本身能够提供含氧基团或者与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。S2. The second ion treatment: using inert gas and reactive plasma gas as initiating substances, plasma etching treatment is performed on the substrate under predetermined conditions, and the plasma etching treatment time is 3 minutes. The predetermined conditions for the second ion treatment in step S2 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 20sccm-180sccm, processing power 60w-350w, and processing time 100s-1200s. In step S2, the volume ratio of the inert gas to the reactive plasma gas is preferably 3:1. Reactive plasma gas is a compound that can provide oxygen-containing groups by itself or break chemical bonds with the substrate material and then react to generate oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
本发明的细胞培养装置表面改性方法能够提升高分子材料表面长效亲水改性的能力、实现长效亲水改性的效果、拓宽产品的应用范围和领域、实现无血清培养以降低用户的成本、易于实现产业化。采用本发明的细胞培养装置表面改性方法，基材表面亲水性增强，接触角达到30°以下且经过3个月的加速老化试验后，基材表面亲水性不发生衰退。本发明先用惰性引发气体对材料表面进行改性处理后，再用一定比例的惰性气体与反应性等离子气体对材料进一步 进行改性，将有利于实现长效亲水改性的效果。难以体外贴壁培养的细胞能够在经本发明的方法处理后的基础表面生长良好，能够满足常规细胞和难养细胞的低血清、无血清培养。The surface modification method of the cell culture device of the present invention can improve the ability of long-term hydrophilic modification on the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and realize serum-free culture to reduce users Low cost and easy to realize industrialization. By adopting the surface modification method of the cell culture device of the present invention, the hydrophilicity of the substrate surface is enhanced, the contact angle is below 30°, and after 3 months of accelerated aging test, the hydrophilicity of the substrate surface does not decline. In the present invention, the surface of the material is modified with an inert initiating gas, and then a certain proportion of inert gas and reactive plasma gas is used to further modify the material, which will help to achieve the effect of long-term hydrophilic modification. Cells that are difficult to adhere to culture in vitro can grow well on the basal surface treated by the method of the present invention, and can meet the low serum and serum-free culture of conventional cells and refractory cells.
实施例2Example 2
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的预定条件如下：真空度20mT-150mT、温度30℃-55℃、气体流量5sccm-80sccm、处理功率60w-350w和处理时间大于240s。步骤S1中所述惰性引发气体优选为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 5sccm-80sccm, processing power 60w-350w, and processing time greater than 240s. The inert initiating gas in step S1 is preferably argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度20mT-150mT、温度30℃-55℃、气体流量20sccm-180sccm、处理功率60w-350w、处理时间100s-1200s。反应性等离子气体为本身能够提供含氧基团或者与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: vacuum degree 20mT-150mT, temperature 30°C-55°C, gas flow rate 20sccm-180sccm, processing power 60w-350w, and processing time 100s-1200s. Reactive plasma gas is a compound that can provide oxygen-containing groups by itself or break chemical bonds with the substrate material and then react to generate oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例3Example 3
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离 子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的预定条件如下：真空度20mT、温度30℃、气体流量80sccm、处理功率350w和处理时间300s。步骤S1中所述引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The predetermined conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 20 mT, a temperature of 30° C., a gas flow rate of 80 sccm, a processing power of 350 w, and a processing time of 300 s. The initiating gas in step S1 is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度20mT、温度30℃、气体流量180sccm、处理功率350w、处理时间100s。反应性等离子气体为本身能够提供含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 20 mT, a temperature of 30° C., a gas flow rate of 180 sccm, a processing power of 350 w, and a processing time of 100 s. The reactive plasma gas is a compound that itself can provide oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例4Example 4
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的预定条件如下：真空度150mT、温度55℃、气体流量5sccm、处理功率60w和处理时间240s。步骤S1中所述惰性引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The predetermined conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 150 mT, a temperature of 55° C., a gas flow rate of 5 sccm, a processing power of 60 w, and a processing time of 240 s. In step S1, the inert initiating gas is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度150mT、温度55℃、气体流量20sccm、处理功率60w、处理时间1200s。反应性等离子气体为本身能够与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 150 mT, a temperature of 55° C., a gas flow rate of 20 sccm, a processing power of 60 w, and a processing time of 1200 s. The reactive plasma gas is a compound that itself can break the chemical bond with the substrate material and then react to generate oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例5Example 5
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的预定条件如下：真空度30mT、温度40℃、气体流量60sccm、处理功率250w和处理时间300s。步骤S1中所述惰性引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The predetermined conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 30 mT, a temperature of 40° C., a gas flow rate of 60 sccm, a processing power of 250 w, and a processing time of 300 s. In step S1, the inert initiating gas is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度30mT、温度40℃、气体流量100sccm、处理功率250w、处理时间300s。反应性等离子气体为本身能够提供含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 30 mT, a temperature of 40° C., a gas flow rate of 100 sccm, a processing power of 250 w, and a processing time of 300 s. The reactive plasma gas is a compound that itself can provide oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例6Example 6
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的工艺条件如下：真空度20mT、温度30℃、气体流量20sccm、处理功率250w、处理时间800s。步骤S1中所述惰性引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The process conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT, temperature 30°C, gas flow rate 20sccm, processing power 250w, and processing time 800s. In step S1, the inert initiating gas is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次 离子处理时的预定条件如下：真空度50mT、温度40℃、气体流量140sccm、处理功率200w、处理时间800s。反应性等离子气体为本身能够提供含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: vacuum degree 50mT, temperature 40°C, gas flow rate 140sccm, processing power 200w, and processing time 800s. The reactive plasma gas is a compound that itself can provide oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例7Example 7
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的工艺条件如下：真空度50mT、温度35℃、气体流量50sccm、处理功率100w、处理时间300s。步骤S1中所述惰性引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The process conditions for the first ion treatment in step S1 are as follows: vacuum degree 50mT, temperature 35°C, gas flow rate 50sccm, processing power 100w, and processing time 300s. In step S1, the inert initiating gas is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度100mT、温度50℃、气体流量160sccm、处理功率100w、处理时间500s。反应性等离子气体为本身能够与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 100 mT, a temperature of 50° C., a gas flow rate of 160 sccm, a processing power of 100 w, and a processing time of 500 s. The reactive plasma gas is a compound that itself can break the chemical bond with the substrate material and then react to generate oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例8Example 8
本实施例提供了一种细胞培养装置表面改性方法。This embodiment provides a method for surface modification of a cell culture device.
一种细胞培养装置表面改性方法包括如下步骤：A method for surface modification of a cell culture device includes the following steps:
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次 离子处理时的工艺条件如下：真空度30mT、温度30℃、气体流量40sccm、处理功率150w、处理时间500s。步骤S1中所述惰性引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The process conditions for the first ion treatment in step S1 are as follows: vacuum degree 30mT, temperature 30°C, gas flow rate 40sccm, processing power 150w, and processing time 500s. In step S1, the inert initiating gas is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度700mT、温度45℃、气体流量150sccm、处理功率150w、处理时间600s。反应性等离子气体为本身能够提供含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 700 mT, a temperature of 45° C., a gas flow rate of 150 sccm, a processing power of 150 w, and a processing time of 600 s. The reactive plasma gas is a compound that itself can provide oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
实施例8Example 8
本实施例提供了一种细胞培养装置。This embodiment provides a cell culture device.
一种细胞培养装置，包括细胞培养支架，所述细胞培养支架通过实施例3-7任一实施例所述的细胞培养装置表面改性方法进行改性，以获得超亲水细胞生长表面。A cell culture device includes a cell culture scaffold, the cell culture scaffold is modified by the cell culture device surface modification method described in any one of Embodiments 3-7 to obtain a super-hydrophilic cell growth surface.
实施例9Example 9
本实施例提供了一种细胞培养装置。This embodiment provides a cell culture device.
一种细胞培养装置，包括细胞培养支架，所述细胞培养支架通过下述细胞培养装置表面改性方法进行改性，以获得超亲水细胞生长表面。A cell culture device includes a cell culture scaffold, which is modified by the following cell culture device surface modification method to obtain a super-hydrophilic cell growth surface.
S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面。步骤S1中第一次离子处理时的工艺条件如下：真空度30mT、温度30℃、气体流量40sccm、处理功率150w、处理时间500s。步骤S1中所述惰性引发气体为氩气。S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface. The process conditions for the first ion treatment in step S1 are as follows: a vacuum degree of 30 mT, a temperature of 30° C., a gas flow rate of 40 sccm, a processing power of 150 w, and a processing time of 500 s. In step S1, the inert initiating gas is argon.
S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。步骤S2中第二次离子处理时的预定条件如下：真空度700mT、温度45℃、气体流量150sccm、处理功率150w、处理时间600s。反应性等离子气体为本身能够提供含氧基团或者与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。所述含氧基团为羰基、羟基或者羧基。步骤S2中惰性气体与反应性等离子气体的体积比为3:1。S2. The second ion treatment: using inert gas and reactive plasma gas as the initiating substance as the initiating substance, and performing plasma etching treatment on the substrate under predetermined conditions. The predetermined conditions for the second ion treatment in step S2 are as follows: a vacuum degree of 700 mT, a temperature of 45° C., a gas flow rate of 150 sccm, a processing power of 150 w, and a processing time of 600 s. Reactive plasma gas is a compound that can provide oxygen-containing groups by itself or break chemical bonds with the substrate material and then react to generate oxygen-containing groups. The oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group. In step S2, the volume ratio of the inert gas to the reactive plasma gas is 3:1.
通过对未处理聚苯乙烯基材、等离子亲水处理聚苯乙烯基材和本发明细胞培养装置表面改性方法处理的聚苯乙烯基材表面的O元素和C元素进行X聚苯乙烯光电子能谱扫描，分析结果如表1所示。X-polystyrene photoelectron energy is performed on the O and C elements on the surface of the untreated polystyrene substrate, the plasma hydrophilic treatment polystyrene substrate, and the surface modification method of the cell culture device of the present invention. Spectrum scan, the analysis results are shown in Table 1.
表1聚苯乙烯基材表面不同处理方式的光电子能谱分析结果Table 1 Photoelectron spectroscopy analysis results of different treatment methods on the surface of polystyrene substrate
处理方式Processing method C1s(atomic％)C1s(atomic%) O1s(atomic％)O1s(atomic%)
普通未处理Normal untreated 98.9798.97 1.031.03
常规亲水处理Conventional hydrophilic treatment 94.8294.82 5.185.18
超亲水处理Super hydrophilic treatment 73.5873.58 26.4226.42
由表1可以看出未处理聚苯乙烯基材表面几乎全是碳元素，只有吸附的或者发生表面氧化的少量的氧元素存在；经过常规亲水处理后的聚苯乙烯基材表面的氧元素含量增加到了5.18％，这说明第一次离子处理后表面发生了部分氧化，引入了羟基、羰基等含氧基团；表面经过丙烯酸等离子接枝处理后，聚苯乙烯基材表面的氧元素含量出现了急剧的增加，达到了26.42％，不仅有羟基、羰基，还有大量的羧基官能团的存在，这些分析结果和前面的接触角测量、红外光谱分析以及X聚苯乙烯表面元素分析结果都是一致的。It can be seen from Table 1 that the surface of the untreated polystyrene substrate is almost entirely carbon, and only a small amount of oxygen is adsorbed or oxidized on the surface; oxygen on the surface of the polystyrene substrate after conventional hydrophilic treatment The content increased to 5.18%, which indicates that the surface was partially oxidized after the first ion treatment, introducing hydroxyl groups, carbonyl groups and other oxygen-containing groups; after the surface was treated with acrylic acid plasma grafting, the oxygen content on the surface of the polystyrene substrate There has been a sharp increase, reaching 26.42%. Not only are hydroxyl groups and carbonyl groups, but also a large number of carboxyl functional groups. These analysis results are the same as the previous contact angle measurement, infrared spectroscopy and X-polystyrene surface elemental analysis. Consistent.
通过对未处理聚苯乙烯基材、等离子亲水处理聚苯乙烯基材和本发明细胞培养装置表面改性方法处理聚苯乙烯基材表面进行原子力显微镜分析，未处理聚苯乙烯基材表面除了一些制样过程中吸附在表面的尘埃颗粒影响外，表面极其光滑平整。而通过第一步的常规亲水处理后表面变得比较粗糙，这说明聚苯 乙烯材料表面通过本发明的细胞培养装置表面改性方法能够提升高分子材料表面长效亲水改性的能力、实现长效亲水改性的效果。The surface of the untreated polystyrene substrate, the plasma hydrophilic treated polystyrene substrate, and the surface modification method of the cell culture device of the present invention were used for atomic force microscope analysis. The surface of the untreated polystyrene substrate was Some dust particles adsorbed on the surface during sample preparation are affected, and the surface is extremely smooth and flat. However, the surface becomes rougher after the first step of the conventional hydrophilic treatment, which indicates that the surface modification method of the cell culture device of the present invention can improve the long-term hydrophilic modification ability of the surface of the polystyrene material. Realize the effect of long-term hydrophilic modification.
综上所述，本发明的细胞培养装置表面改性方法能够提升高分子材料表面长效亲水改性的能力、实现长效亲水改性的效果、拓宽产品的应用范围和领域、实现无血清培养以降低用户的成本、易于实现产业化。采用本发明的细胞培养装置表面改性方法，基材表面亲水性增强，接触角达到30°以下且经过3个月的加速老化试验后，基材表面亲水性不发生衰退。本发明先用惰性引发气体对材料表面进行改性处理后，再用一定比例的惰性气体与反应性等离子气体对材料进一步进行改性，将有利于实现长效亲水改性的效果。难以体外贴壁培养的细胞能够在经本发明的方法处理后的基础表面生长良好，能够满足常规细胞和难养细胞的低血清、无血清培养。In summary, the cell culture device surface modification method of the present invention can improve the ability of long-term hydrophilic modification of the surface of polymer materials, realize the effect of long-term hydrophilic modification, broaden the application range and fields of products, and realize Serum culture can reduce the cost of users and is easy to realize industrialization. By adopting the surface modification method of the cell culture device of the present invention, the hydrophilicity of the substrate surface is enhanced, the contact angle is below 30°, and after 3 months of accelerated aging test, the hydrophilicity of the substrate surface does not decline. In the present invention, the surface of the material is modified with an inert initiating gas, and then a certain proportion of inert gas and reactive plasma gas is used to further modify the material, which will help realize the effect of long-term hydrophilic modification. Cells that are difficult to adhere to culture in vitro can grow well on the basal surface treated by the method of the present invention, and can meet the low serum and serum-free culture of conventional cells and refractory cells.
以上所述实施例的各技术特征可以进行任意的组合，为使描述简洁，未对上述实施例中的各个技术特征所有可能的组合都进行描述，然而，只要这些技术特征的组合不存在矛盾，都应当认为是本说明书记载的范围。The technical features of the above-mentioned embodiments can be combined arbitrarily. In order to make the description concise, all possible combinations of the various technical features in the above-mentioned embodiments are not described. However, as long as there is no contradiction in the combination of these technical features, All should be considered as the scope of this specification.
以上所述实施例仅表达了本发明的几种实施方式，其描述较为具体和详细，但并不能因此而理解为对本发明专利范围的限制。应当指出的是，对于本领域的普通技术人员来说，在不脱离本发明构思的前提下，还可以做出若干变形和改进，这些都属于本发明的保护范围。因此，本发明专利的保护范围应以所附权利要求为准。The above-mentioned embodiments only express several implementation modes of the present invention, and the description is relatively specific and detailed, but it should not be understood as a limitation to the patent scope of the present invention. It should be pointed out that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can be made, and these all fall within the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention should be subject to the appended claims.

Claims (10)

  1. 一种细胞培养装置表面改性方法，其特征在于，包括如下步骤：A method for surface modification of a cell culture device, which is characterized in that it comprises the following steps:
    S1、第一次离子处理：惰性引发气体在预定条件下对基材的表面进行等离子体蚀刻处理，使得所述基材的表面形成长效亲水改性表面；S1. The first ion treatment: the inert initiating gas performs plasma etching treatment on the surface of the substrate under predetermined conditions, so that the surface of the substrate forms a long-lasting hydrophilic modified surface;
    S2、第二次离子处理：以惰性气体与反应性等离子气体作为引发物质，在预定条件下对所述基材进行等离子体蚀刻处理。S2. The second ion treatment: using inert gas and reactive plasma gas as initiating substances, plasma etching treatment is performed on the substrate under predetermined conditions.
  2. 根据权利要求1所述的细胞培养装置表面改性方法，其特征在于，步骤S1中所述惰性引发气体为氩气。The method for surface modification of a cell culture device according to claim 1, wherein the inert initiating gas in step S1 is argon.
  3. 根据权利要求2所述的细胞培养装置表面改性方法，其特征在于，步骤S2中惰性气体与反应性等离子气体的体积比为3:1。The surface modification method of a cell culture device according to claim 2, wherein the volume ratio of the inert gas to the reactive plasma gas in step S2 is 3:1.
  4. 根据权利要求1所述的细胞培养装置表面改性方法，其特征在于，所述反应性等离子气体为能够提供含氧基团或者与基材材料发生化学键断裂再反应可以生成含氧基团的化合物。The method for surface modification of a cell culture device according to claim 1, wherein the reactive plasma gas is a compound capable of providing oxygen-containing groups or breaking a chemical bond with the substrate material and then reacting to generate oxygen-containing groups .
  5. 根据权利要求4所述的细胞培养装置表面改性方法，其特征在于，所述含氧基团为羰基、羟基或者羧基。The method for surface modification of a cell culture device according to claim 4, wherein the oxygen-containing group is a carbonyl group, a hydroxyl group or a carboxyl group.
  6. 根据权利要求1-5任意一项所述的细胞培养装置表面改性方法，其特征在于，步骤S1中第一次离子处理时的预定条件如下：The method for modifying the surface of a cell culture device according to any one of claims 1-5, wherein the predetermined conditions for the first ion treatment in step S1 are as follows:
    真空度20mT-150mT、温度30℃-55℃、气体流量5sccm-80sccm、处理功率60w-350w和处理时间大于240s。Vacuum degree 20mT-150mT, temperature 30℃-55℃, gas flow rate 5sccm-80sccm, processing power 60w-350w and processing time greater than 240s.
  7. 根据权利要求6所述的细胞培养装置表面改性方法，其特征在于，步骤S1中第一次离子处理时的预定条件如下：真空度20mT-50mT、温度30℃-35℃、气体流量20sccm-50sccm、处理功率100w-250w、处理时间300s-800s。The cell culture device surface modification method according to claim 6, wherein the predetermined conditions for the first ion treatment in step S1 are as follows: vacuum degree 20mT-50mT, temperature 30°C-35°C, gas flow rate 20sccm- 50sccm, processing power 100w-250w, processing time 300s-800s.
  8. 根据权利要求1-5任意一项所述的细胞培养装置表面改性方法，其特征在于，步骤S2中第二次离子处理时的预定条件如下：The method for modifying the surface of a cell culture device according to any one of claims 1-5, wherein the predetermined conditions during the second ion treatment in step S2 are as follows:
    真空度20mT-150mT、温度30℃-55℃、气体流量20sccm-180sccm、处理功率60w-350w、处理时间100s-1200s。Vacuum degree 20mT-150mT, temperature 30℃-55℃, gas flow rate 20sccm-180sccm, processing power 60w-350w, processing time 100s-1200s.
  9. 根据权利要求8所述的细胞培养装置表面改性方法，其特征在于，步骤S2中第二次离子处理时的预定条件如下：The method for surface modification of a cell culture device according to claim 8, wherein the predetermined conditions for the second ion treatment in step S2 are as follows:
    真空度50mT-100mT、温度40℃-50℃、气体流量140sccm-160sccm、处理功率100w-200w、处理时间500s-800s。Vacuum degree 50mT-100mT, temperature 40℃-50℃, gas flow rate 140sccm-160sccm, processing power 100w-200w, processing time 500s-800s.
  10. 一种细胞培养装置，其特征在于，包括细胞培养支架，所述细胞培养支架通过权利要求1-9任意一项所述的细胞培养装置表面改性方法进行改性，以获得长效亲水改性表面。A cell culture device, characterized by comprising a cell culture scaffold, the cell culture scaffold is modified by the cell culture device surface modification method of any one of claims 1-9 to obtain a long-term hydrophilic modification Sexual surface.
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