CN108148759B - A multipurpose gas permeable container for adherent cell culture/co-culture - Google Patents
A multipurpose gas permeable container for adherent cell culture/co-culture Download PDFInfo
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- 238000003501 co-culture Methods 0.000 title claims abstract description 36
- 238000004115 adherent culture Methods 0.000 title claims abstract description 10
- 229920001296 polysiloxane Polymers 0.000 claims abstract description 24
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 claims abstract description 22
- 229910002092 carbon dioxide Inorganic materials 0.000 claims abstract description 11
- 239000001569 carbon dioxide Substances 0.000 claims abstract description 11
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical group O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims abstract description 6
- 239000012528 membrane Substances 0.000 claims description 24
- 239000000463 material Substances 0.000 claims description 10
- 239000007788 liquid Substances 0.000 claims description 7
- 229920000515 polycarbonate Polymers 0.000 claims description 5
- 239000004417 polycarbonate Substances 0.000 claims description 5
- -1 polytetrafluoroethylene Polymers 0.000 claims description 3
- 229920001343 polytetrafluoroethylene Polymers 0.000 claims description 3
- 239000004810 polytetrafluoroethylene Substances 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 2
- 239000006059 cover glass Substances 0.000 claims 1
- 230000035699 permeability Effects 0.000 abstract description 8
- 238000007789 sealing Methods 0.000 abstract description 4
- 239000001963 growth medium Substances 0.000 abstract description 3
- 210000004027 cell Anatomy 0.000 description 23
- 238000004113 cell culture Methods 0.000 description 11
- 239000002609 medium Substances 0.000 description 10
- 238000000034 method Methods 0.000 description 5
- 238000011109 contamination Methods 0.000 description 4
- 238000002474 experimental method Methods 0.000 description 3
- 229920002529 medical grade silicone Polymers 0.000 description 3
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000001954 sterilising effect Effects 0.000 description 2
- 238000004659 sterilization and disinfection Methods 0.000 description 2
- 230000001464 adherent effect Effects 0.000 description 1
- 210000004102 animal cell Anatomy 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
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Abstract
本发明公开了一种用于贴壁细胞培养/共培养的多用途透气容器，其包括：容器盒、容器盖、共培养内框，硅胶圈和透气不透水薄膜；容器盖是由带有十字支撑架的盖体、上下两个硅胶圈、透气不透水薄膜组成；容器盒底部有四个底部支撑架；将一个硅胶圈压入容器盖的内侧四周，同时将透气不透水薄膜嵌入容器盖内，然后在薄膜四周再压入一个硅胶圈，将透气薄膜固定在容器盖内，容器盖上十字支撑架用于支撑透气薄膜。本容器的特点是即可以保证容器的密封，防止培养基的渗漏、污染，又可以保证容器二氧化碳的透气率；同时该容器还可以实现细胞的远距共培养。
The invention discloses a multipurpose gas permeable container for adherent cell culture/co-culture, which comprises: a container box, a container cover, a co-cultivation inner frame, a silica gel ring and a gas-permeable and water-permeable film; The cover of the support frame, the upper and lower silicone rings, and the breathable and watertight film are composed; there are four bottom support frames at the bottom of the container box; a silicone ring is pressed into the inner circumference of the container cover, and the breathable and watertight film is embedded in the container cover at the same time. , and then press a silicone ring around the film to fix the breathable film in the container cover, and a cross support frame is placed on the container cover to support the breathable film. The characteristics of the container are that it can not only ensure the sealing of the container, prevent the leakage and pollution of the culture medium, but also ensure the air permeability of carbon dioxide in the container; at the same time, the container can also realize the long-distance co-culture of cells.
Description
技术领域technical field
本发明涉及细胞培养技术领域，具体为一种用于贴壁细胞培养/共培养的多用途透气容器。The invention relates to the technical field of cell culture, in particular to a multipurpose gas permeable container for adherent cell culture/co-culture.
背景技术Background technique
目前在开展细胞生物学方面研究过程中大多采用的是培养皿、培养板或者培养瓶细胞培养容器。但这些细胞培养容器在使用过程中都存在一定的缺陷：首先这些培养容器加入培养基后，在培养箱中孵育的过程中，为了保证容器的透气性(动物细胞培养要求一定浓度的二氧化碳气体)，容器盖不旋紧，这就导致在取放容器的过程中，容易造成培养基的泄露，从而造成培养基的污染，但如果旋紧容器盖并采用封口膜密封则会影响容器的透气率，造成培养基酸碱度变化，从而造成细胞死亡；其次，这些容器均无法实现细胞的共培养，而专用的细胞共培养容器价格均较为昂贵；再次这些培养容器大多是一次性的细胞培养容器，不能够高压灭菌，价格昂贵，因此长期实验的费用较高。At present, in the process of carrying out cell biology research, culture dishes, culture plates or culture flasks are mostly used for cell culture vessels. However, these cell culture containers have certain defects in the use process: first, after adding the medium to these culture containers, in the process of incubating in the incubator, in order to ensure the permeability of the container (animal cell culture requires a certain concentration of carbon dioxide gas) , the container cover is not tightened, which leads to the leakage of the medium during the process of taking and placing the container, resulting in contamination of the medium, but if the container cover is tightened and sealed with a parafilm, the air permeability of the container will be affected. , causing the pH of the medium to change, resulting in cell death; secondly, these containers cannot realize co-culture of cells, and the price of dedicated cell co-culture containers is relatively expensive; again, these culture containers are mostly disposable cell culture containers, not Can be autoclaved, expensive, and therefore more expensive for long-term experiments.
发明内容SUMMARY OF THE INVENTION
本发明的目的在于提供一种用于贴壁细胞培养/共培养的多用途透气容器。本容器的特点是即可以保证容器的密封，防止培养基的渗漏、污染，又可以保证容器二氧化碳的透气率；同时该容器还可以实现细胞的远距共培养。The object of the present invention is to provide a multipurpose gas permeable container for adherent cell culture/co-culture. The characteristics of the container are that it can not only ensure the sealing of the container, prevent the leakage and pollution of the culture medium, but also ensure the air permeability of carbon dioxide in the container; at the same time, the container can also realize the long-distance co-culture of cells.
为实现上述目的，本发明提供如下技术方案：To achieve the above object, the present invention provides the following technical solutions:
一种用于贴壁细胞培养/共培养的多用途透气容器，其包括：容器盒、容器盖、硅胶圈和透气不透水薄膜；容器盖是由带有十字支撑架的盖体、上下两个硅胶圈、透气不透水薄膜组成；容器盒底部有四个底部支撑架；将一个硅胶圈压入容器盖的内侧四周，同时将透气不透水薄膜嵌入容器盖内，然后在薄膜四周再压入一个硅胶圈，将透气薄膜固定在容器盖内，容器盖上十字支撑架用于支撑透气薄膜；当进行单独培养时，将接种有细胞的盖玻片通过镊子放入容器盒内，盖玻片受到四个支撑架支撑，或者直接将细胞接种在容器盒底部，然后容器盖和容器盒通过内外螺纹旋紧。A multipurpose gas permeable container for adherent cell culture/co-culture, comprising: a container box, a container cover, a silica gel ring and a gas-permeable and watertight film; Silicone ring, breathable and impermeable film; there are four bottom support frames at the bottom of the container box; a silicone ring is pressed into the inner circumference of the container cover, and the breathable and impermeable film is embedded in the container cover, and then another one is pressed around the film. The silicone ring fixes the breathable film in the container cover, and the container is covered with a cross support frame to support the breathable film; when culturing alone, put the cell-seeded coverslip into the container box through tweezers, and the coverslip is subjected to Four support frames support, or the cells are directly seeded on the bottom of the container box, and then the container cover and the container box are screwed on by internal and external threads.
硅胶圈和透气不透水薄膜，两个硅胶圈固定透气膜后嵌入容器盖内部，然后容器盖和容器盒通过内外螺纹旋紧，硅胶圈即起到固定透气膜的作用，同时也起到密封容器的作用，防止培养基液体渗漏，透气不透水薄膜即可保证一定浓度二氧化碳气体的通过率，又可防止培养基液体的渗漏，十字支撑架用于支撑透气不透水薄膜，Silicone ring and breathable impermeable film, two silicone rings fix the breathable film and then embed inside the container cover, then the container cover and the container box are screwed tightly through the inner and outer threads, the silicone ring not only plays the role of fixing the breathable film, but also sealing the container It can prevent the leakage of the medium liquid. The breathable and impermeable film can ensure the passage rate of a certain concentration of carbon dioxide gas and prevent the leakage of the medium liquid. The cross support frame is used to support the breathable and impermeable film.
共培养内框为中空圆柱体，内框四周为一凹槽，用于固定共培养膜，内框顶部和底部四周边缘为弧形设计，当进行共培养时，将共培养膜用一个或两个硅胶圈，通过共培养框上的凹槽固定在共培养框上，使共培养框的一端为共培养膜，另一端通透，将共培养细胞接种在共培养膜上，然后放将接种有共培养细胞的共培养框放入底部接种有细胞的容器盒内，然后容器盖和容器盒通过内外螺纹旋紧。The inner frame of the co-cultivation is a hollow cylinder with a groove around the inner frame for fixing the co-cultivation membrane. The edges around the top and bottom of the inner frame are arc-shaped. A silica gel ring is fixed on the co-cultivation frame through the groove on the co-cultivation frame, so that one end of the co-cultivation frame is a co-cultivation membrane and the other end is permeable, and the co-cultivation cells are inoculated on the co-cultivation membrane. The co-culture frame with the co-cultured cells is put into the container box with the cells seeded at the bottom, and then the container cover and the container box are screwed together by internal and external threads.
容器盒上部分四周为外螺纹，容器内径32.6毫米，外径35毫米，容器盒底部有四个平均分配的支撑架，支撑架长4毫米、宽2毫米、高1毫米(细胞远距共培养距离要求)，便于取放盖玻片和用于细胞远距共培养实验，培养盒的材料为聚碳酸酯，具有较好的生物相容性，可以耐受湿热灭菌条件，可以重复使用，减少了白色污染。The upper part of the container box is surrounded by external threads, the inner diameter of the container is 32.6 mm, and the outer diameter is 35 mm. There are four evenly distributed support frames at the bottom of the container box. Distance requirements), easy to pick and place coverslips and used for long-distance co-culture experiments of cells, the material of the culture box is polycarbonate, which has good biocompatibility, can withstand moist heat sterilization conditions, and can be reused. Reduced white pollution.
容器盖为内螺纹，盖的上表面为中空，便于放置透气不透水薄膜，用于透过一定浓度的二氧化碳气体，但有一十字支撑架，防止透气薄膜变形，容器盖由聚碳酸酯材料制作。The container cover is internally threaded, and the upper surface of the cover is hollow, which is convenient for placing a breathable and watertight film to pass through a certain concentration of carbon dioxide gas, but there is a cross support frame to prevent the deformation of the breathable film. The container cover is made of polycarbonate material.
共培养内框为中空圆柱体，内框高度为8mm，内框直径29mm，内框四周为一凹槽，用于固定共培养膜，内框凹槽固定共培养膜后带硅胶圈总直径为31mm，内框顶部和底部四周边缘为弧形设计，防止膜损害。The inner frame of the co-cultivation is a hollow cylinder, the height of the inner frame is 8 mm, the diameter of the inner frame is 29 mm, and the inner frame is surrounded by a groove for fixing the co-cultivation membrane. 31mm, the edges around the top and bottom of the inner frame are curved to prevent membrane damage.
硅胶圈由医用硅胶制作，共计四个硅胶圈，两个放于透气不透水薄膜的上下两面，用于固定透气不透水薄膜，同时密封容器口；两个放置在共培养内框凹槽内，用于固定细胞共培养膜。The silicone ring is made of medical silicone. There are four silicone rings in total, two of which are placed on the upper and lower sides of the breathable and impermeable film to fix the breathable and impermeable film and seal the container mouth at the same time; two are placed in the groove of the co-cultivation inner frame, Used to fix cell co-culture membranes.
透气不透水薄膜材料为聚四氟乙烯，通过上下两个硅胶圈固定后放入容器盖的内部，即可保证二氧化碳气体的透气率，又可保证培养基不渗漏，防止细胞污染。The material of the breathable and water-impermeable film is polytetrafluoroethylene, which is fixed by the upper and lower two silicone rings and then placed inside the container cover, which can ensure the permeability of carbon dioxide gas, and also ensure that the medium does not leak and prevent cell contamination.
透气不透水薄膜和细胞共培养膜可采用商业化共培养膜，根据容器盖内径尺寸和共培养内框大小适当裁剪。Commercially available co-culture membranes can be used for gas-permeable and water-impermeable membranes and cell co-culture membranes, which are appropriately tailored according to the inner diameter of the container lid and the inner frame of the co-culture.
与现有技术相比，本发明的有益效果如下：Compared with the prior art, the beneficial effects of the present invention are as follows:
1.本细胞培养容器可以保证对培养基液体的密封性，防止液体渗漏；1. The cell culture container can ensure the tightness of the medium liquid and prevent liquid leakage;
2.本细胞培养容器可以保证一定浓度的二氧化碳的透气率，保证细胞的正常生长；2. The cell culture container can ensure the permeability of carbon dioxide with a certain concentration and ensure the normal growth of cells;
3.本细胞培养容器的所有材料均可以耐受湿热高压灭菌条件，减少了微生物污染的可能性，同时还可以重复使用，降低了实验费用；3. All the materials of this cell culture container can withstand moist heat autoclaving conditions, reducing the possibility of microbial contamination, and can also be reused, reducing experimental costs;
4.本细胞培养容器可实现细胞的远距共培养。4. The cell culture vessel can realize the long-distance co-culture of cells.
附图说明Description of drawings
图1是本发明的容器盒立体图。Fig. 1 is a perspective view of a container box of the present invention.
图2是本发明的容器盒的俯视图。Fig. 2 is a plan view of the container box of the present invention.
图3是本发明的容器盒的剖面图。Fig. 3 is a cross-sectional view of the container box of the present invention.
图4是本发明的容器盖的立体图。Fig. 4 is a perspective view of the container lid of the present invention.
图5是本发明的容器盖的俯视图。Fig. 5 is a plan view of the container lid of the present invention.
图6是本发明的容器盖的剖面图。Fig. 6 is a cross-sectional view of the container lid of the present invention.
图7是本发明的共培养内框立体图Figure 7 is a perspective view of the inner frame of the co-culture of the present invention
图8是本发明的共培养内框俯视图Figure 8 is a top view of the inner frame of the co-culture of the present invention
图9是本发明的共培养内框的剖面图。Fig. 9 is a cross-sectional view of the inner frame of the co-culture of the present invention.
在图1-9中：1底部支撑架，2底部支撑架，3底部支撑架，4底部支撑架，5外螺纹，6十字支撑架，7十字支撑架，8凹槽,In Figure 1-9: 1 bottom support frame, 2 bottom support frame, 3 bottom support frame, 4 bottom support frame, 5 external thread, 6 cross support frame, 7 cross support frame, 8 groove,
具体实施方式Detailed ways
下面将结合本发明实施例中的附图，对本发明实施例中的技术方案进行清楚、完整地描述，显然，所描述的实施例仅仅是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有做出创造性劳动前提下所获得的所有其他实施例，都属于本发明保护的范围。The technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, but not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.
如图1-9，一种用于贴壁细胞培养的多用途透气容器，其包括：容器盒图1、容器盖图4、共培养内框图7、硅胶圈、透气不透水薄膜和共培养膜；As shown in Figures 1-9, a multi-purpose gas permeable container for adherent cell culture, which includes: container box Figure 1, container cover Figure 4, co-culture
贴壁细胞培养具体实施方式：The specific embodiment of adherent cell culture:
首先将医用硅胶圈压入图4容器盖的内侧四周，同时将透气不透水薄膜嵌入容器盖内，然后在薄膜四周再压入一个硅胶圈，将透气薄膜固定在容器盖内，容器盖上十字支撑架6、7用于支撑透气薄膜，保证透气薄膜的平整性，同时防止透气薄膜损坏。First, press the medical silicone ring into the inner circumference of the container cover as shown in Figure 4, and at the same time embed the breathable and impermeable film into the container cover, and then press a silicone ring around the film to fix the breathable film in the container cover. The
其次，图1容器盒内添加一定量的培养基后，将接种有细胞的盖玻片通过镊子放入容器盒内，盖玻片受到四个支撑架1、2、3、4支撑，便于盖玻片的取放；也可将细胞直接接种在容器底部。Next, after adding a certain amount of culture medium to the container box shown in Figure 1, put the cell-seeded coverslip into the container box through tweezers. Pick and place slides; cells can also be seeded directly on the bottom of the container.
最后将固定有透气薄膜的容器盖通过螺纹和容器盒旋紧。硅胶圈可保证容器的密封，不渗漏培养基液体，透气不透水薄膜可保证一定浓度的二氧化碳气体的透气率。Finally, screw the container cover with the breathable film fixed to the container box through the screw thread. The silicone ring can ensure the sealing of the container, and the medium liquid does not leak, and the breathable and watertight film can ensure the air permeability of a certain concentration of carbon dioxide gas.
贴壁细胞共培养具体实施方式：Adherent cell co-culture specific embodiment:
首先将图7共培养内框底部放置共培养膜，通过硅胶圈和凹槽固定共培养膜；First, place the co-culture membrane at the bottom of the co-culture inner frame in Figure 7, and fix the co-culture membrane through the silica gel ring and groove;
其次按照细胞培养具体实施实例，将一种细胞接种在容器底部。同时在另一容器内将另外一种细胞接种在共培养膜上表面；Next, according to the specific example of cell culture, a kind of cell is seeded on the bottom of the container. At the same time, another cell is seeded on the surface of the co-culture membrane in another container;
最后将接种有细胞的共培养内框放置在容器内部的四个支撑架上，旋紧固定有透气薄膜的容器盖，放置在培养箱中孵育，实现两种细胞的远距共培养。Finally, the co-culture inner frame inoculated with cells is placed on the four support frames inside the container, the container cover fixed with the gas permeable film is tightened, and the container is placed in the incubator for incubation to realize the long-distance co-culture of the two cells.
本发明细胞培养/共培养多用途透气容器所使用材料均可以高压灭菌，重复使用。The materials used in the cell culture/co-culture multipurpose gas permeable container of the present invention can be autoclaved and reused.
容器盒上部分四周为外螺纹，容器内径32.6毫米，外径35毫米，容器盒底部有四个平均分配的支撑架，支撑架长4毫米、宽2毫米、高1毫米(细胞远距共培养距离要求)，便于取放盖玻片和用于细胞远距共培养实验，培养盒的材料为聚碳酸酯，具有较好的生物相容性，可以耐受湿热灭菌条件，可以重复使用，减少了白色污染。The upper part of the container box is surrounded by external threads, the inner diameter of the container is 32.6 mm, and the outer diameter is 35 mm. There are four evenly distributed support frames at the bottom of the container box. Distance requirements), easy to pick and place coverslips and used for long-distance co-culture experiments of cells, the material of the culture box is polycarbonate, which has good biocompatibility, can withstand moist heat sterilization conditions, and can be reused. Reduced white pollution.
容器盖为内螺纹，盖的上表面为中空，便于放置透气薄膜，用于透过一定浓度的二氧化碳气体，但有一十字支撑架，防止透气薄膜变形，容器盖由聚碳酸酯材料制作。The container cover is internally threaded, and the upper surface of the cover is hollow, which is convenient for placing a breathable film to pass through a certain concentration of carbon dioxide gas, but there is a cross support frame to prevent the deformation of the breathable film. The container cover is made of polycarbonate material.
共培养内框为中空圆柱体，内框高度为8mm，内框直径29mm，内框四周为一凹槽，用于固定共培养膜，内框凹槽固定共培养膜后带硅胶圈总直径为31mm，内框顶部和底部四周边缘为弧形设计，防止膜损害。The inner frame of the co-cultivation is a hollow cylinder, the height of the inner frame is 8 mm, the diameter of the inner frame is 29 mm, and the inner frame is surrounded by a groove for fixing the co-cultivation membrane. 31mm, the edges around the top and bottom of the inner frame are curved to prevent membrane damage.
硅胶圈由医用硅胶制作，共计四个硅胶圈，两个放于透气膜的上下两面，用于固定透气膜，同时密封容器口；两个放置在共培养内框凹槽内，用于固定细胞共培养膜。The silicone ring is made of medical silicone, with a total of four silicone rings, two of which are placed on the upper and lower sides of the breathable membrane to fix the breathable membrane and seal the mouth of the container; two are placed in the groove of the co-culture inner frame to fix cells Co-culture membrane.
透气不透水薄膜材料为聚四氟乙烯，通过上下两个硅胶圈固定后放入容器盖的内部，即可保证二氧化碳气体的透气率，又可保证培养基不渗漏，防止细胞污染。The material of the breathable and water-impermeable film is polytetrafluoroethylene, which is fixed by the upper and lower two silicone rings and then placed inside the container cover, which can ensure the permeability of carbon dioxide gas, and also ensure that the medium does not leak and prevent cell contamination.
细胞共培养膜可采用商业化共培养膜，根据共培养内框大小适当裁剪。The cell co-culture membrane can be a commercial co-culture membrane, which is appropriately tailored according to the size of the inner frame of the co-culture.
尽管已经示出和描述了本发明的实施例，对于本领域的普通技术人员而言，可以理解在不脱离本发明的原理和精神的情况下可以对这些实施例进行多种变化、修改、替换和变型，本发明的范围由所附权利要求及其等同物限定。Although embodiments of the present invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, and substitutions can be made in these embodiments without departing from the principle and spirit of the invention and modifications, the scope of the present invention is defined by the appended claims and their equivalents.
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