Cell culture system and bioreactor

Abstract

The utility model aims to provide a cell culture system and a bioreactor. Wherein the culture equipment comprises a mixer used for mixing substances required by cell culture, the mixer comprises a mixing container and a mixing driving part, the mixing container provides a sealed mixing space and is communicated with the outside through a first interface and a second interface, and the mixing driving part can drive the substances to be mixed to move and mix; the bioreactor comprises a reaction container and a plurality of culture trays, the reaction container provides a closed space, and the plurality of culture trays are arranged in the closed space; the closed space is communicated with the outside through a third interface and a fourth interface, the third interface can be communicated with the second interface of the mixer, the fourth interface can be communicated with the first interface of the mixer, and perfusion culture in the bioreactor is provided.

Description

Technical Field

[0001] The present application relates to a cell culture system and a bioreactor. Background Art

[0002] In the prior art, the in vitro culture of adherent cells mostly adopts two-dimensional static culture in "open or semi-open containers", such as culture dishes, culture bottles, multi-layer culture bottles, cell factories and other containers. Usually, the bottom surface of such culture containers is treated with TC (tissue culture) to facilitate the culture of adherent cells. The operator spots the cell suspension in such culture containers, adds sufficient culture medium, and uses the "cross or 8-shaped" shaking method to evenly disperse the cells in the container, and then puts it in a cell culture incubator for static adhesion culture. The cells are tightly attached to the bottom of the container and exchange substances with the outside world through the free surface. The operator needs to regularly replace the culture medium in the container to ensure a stable growth environment for the cells.

[0003] The inventors found that the cultivation method of the prior art introduced above has many disadvantages. On the one hand, there are many steps of personnel intervention in the cultivation process, the operation is cumbersome and causes a greater risk of contamination. The scale of cell culture is limited and is not suitable for large-scale cultivation. On the other hand, the cell culture mold is relatively simple, and can only be used for cells to adhere to the bottom surface of the cell culture device for cultivation, resulting in a large waste of culture space, and such "semi-open containers" are "co-existing in the same room" in the incubator, which is prone to cause large-scale cell contamination. In addition, the material exchange efficiency of cell adherence static culture is low, the growth rate is slow, and a large amount of nutrients are wasted to maintain a better cell growth environment. At the same time, this static culture method causes the cells to gradually lose their original properties in the body, which is far from the state of natural growth in the body in terms of morphology, structure and function. For example, the cells gradually become flatter, divide abnormally and lose their differentiation phenotype.

[0004] Therefore, the art needs a new cell culture system and bioreactor to solve the problems existing in the prior art. Utility Model Content

[0005] A cell culture system according to the first aspect of the present application comprises: a mixer for mixing substances required for cell culture, the mixer comprising a mixing container and a mixing drive, the mixing container providing a sealed mixing space, communicating with the outside world through a first interface and a second interface, the mixing drive being able to drive the substances to be mixed to move and mix; a bioreactor comprising a reaction container and a plurality of culture plates, the reaction container providing a closed space, the plurality of culture plates being arranged in the closed space; the closed space being communicated with the outside world through a third interface and a fourth interface, the third interface being able to communicate with the second interface of the mixer, the fourth interface being able to communicate with the first interface of the mixer, providing perfusion culture in the bioreactor; the direction from the third interface to the fourth interface is defined as a first direction, a plurality of the culture plates being adjacently distributed in the first direction, at least some of the culture plates having through holes, the through holes penetrating the thickness of the culture plates in the first direction; preferably, a plurality of circles of the through holes are distributed in the direction from the center to the periphery of the culture plates, each circle having a plurality of the through holes; more preferably, the bioreactor is a disposable consumable.

[0006] The beneficial effects of the cell culture system described above include, but are not limited to, the culture system provides perfusion culture in the bioreactor through a structure connected between the mixer and the bioreactor, especially the third interface can be connected to the second interface of the mixer, and the fourth interface can be connected to the first interface of the mixer. In addition, a structure of culture plates with through holes is stacked in the bioreactor. The culture plates not only provide more locations for cell growth to attach, but also their arrangement direction and the structure of the through holes further optimize the flow effect of the culture medium, further enhance the effect of cell perfusion culture in the bioreactor, realize sufficient material exchange between the culture material and the cells in the bioreactor, improve the culture effect, make the cell growth rate faster, save the culture material, and also make the cells maintain a normal natural growth form.

[0007] In one or more embodiments of the cell culture system, the interval between adjacent culture plates is 0.5 mm to 10 mm; preferably, the interval between adjacent culture plates is 0.5 mm to 2 mm.

[0008] In one or more embodiments of the cell culture system, the plurality of adjacent culture plates are separated by a support member; or, a mounting shaft is provided in the closed space, and the plurality of adjacent culture plates are fixedly disposed adjacent to each other on the mounting shaft.

[0009] In one or more embodiments of the cell culture system, the reaction container further comprises a pressure interface, and the pressure interface is used to connect to a pressure source; preferably, the pressure source is a filtering mechanism or a pressurizing mechanism.

[0010] In one or more embodiments of the cell culture system, an observation window and an observation device are further included, wherein the observation window provides a visible surface to at least a portion of the enclosed space, and the observation device is optically connected to the visible surface; preferably, the observation device also includes a moving mechanism.

[0011] In one or more embodiments of the cell culture system, the bioreactor further comprises a heat exchange structure disposed at the periphery of the reaction container.

[0012] In one or more embodiments of the cell culture system, the reaction container is separately connected to the multiple culture plates, or is integrally connected to the multiple culture plates.

[0013] In one or more embodiments of the cell culture system, a rotation mechanism is further included, which can drive the bioreactor to rotate around an axis.

[0014] In one or more embodiments of the cell culture system, the mixing container includes a mixing tank body, the mixing drive is arranged inside the mixing tank body, and includes a stirring paddle or a stirring rod; or, the mixing container includes a mixing bag, the mixing drive is arranged outside the mixing bag and connected thereto, and the mixing drive includes an oscillating shaker.

[0015] A bioreactor according to the second aspect of the present application comprises a reaction container and a plurality of culture trays, wherein the reaction container provides a closed space, and the plurality of culture trays are arranged in the closed space; the closed space is connected to the outside through a third interface and a fourth interface, the third interface is arranged below the reaction container, and the fourth interface is arranged above the reaction container; the direction in which the third interface points to the fourth interface is defined as a first direction, and a plurality of the culture trays are adjacently distributed in the first direction, and at least some of the culture trays have through holes, and the through holes penetrate the thickness of the culture trays in the first direction; preferably, the bioreactor is used in the cell culture system as described in the first aspect; more preferably, the bioreactor is a disposable consumable; and still more preferably, the reaction container and the culture trays are manufactured integrally.

[0016] The beneficial effects of using the bioreactor introduced above include, but are not limited to, a structure in which culture plates with through holes are stacked in the bioreactor. The culture plates not only provide more locations for cell growth to attach, but their arrangement direction and the structure of the through holes also further optimize the flow effect of the culture medium, further enhance the effect of cell perfusion culture in the bioreactor, achieve sufficient material exchange between the culture material and the cells in the bioreactor, improve the culture effect, make the cell growth rate faster, save the culture material, and also allow the cells to maintain a normal natural growth form.

[0017] According to a biological culture method of the third aspect of the present application, using the cell culture system as described in the first aspect, the culture method includes: after the culture medium is mixed in the mixer, it flows into the reaction container through the second interface and the third interface, flows along the first direction and through at least part of the through holes of the culture plate, flows to the fourth interface, and flows back to the mixer through the first interface.

[0018] The beneficial effects of the above-described biological culture method include, but are not limited to, achieving sufficient material exchange between culture materials and cells, improving culture effects, making cells grow faster, saving culture materials, and allowing cells to maintain a normal natural growth morphology.

[0019] In one or more embodiments of the biological culture method, the method further includes cutting off the connection between the bioreactor and the mixer, closing one of the third and fourth interfaces of the bioreactor, and discharging the culture medium from the other interface; then the other interface is closed, the one interface is opened, and an enzymatic solution is injected; after treatment, the one interface is closed, the other interface is opened, and the cell suspension is discharged. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The above and other features, properties and advantages of the present invention will become more apparent through the following description in conjunction with the accompanying drawings and embodiments, in which:

[0021] Figure 1 It is a schematic structural diagram of a cell culture system according to an embodiment.

[0022] Figure 2 It is a schematic structural diagram of a mixer of a cell culture system according to an embodiment.

[0023] Figure 3 It is a schematic diagram of the internal structure of a bioreactor according to an embodiment.

[0024] Figure 4 It is a schematic diagram of the internal structure of a reaction container and a culture plate of a bioreactor in one embodiment.

[0025] Figure 5 It is a schematic diagram of the structure of a culture dish of a bioreactor according to an embodiment.

[0026] Figure 6 It is a schematic diagram of the structure of a bioreactor and an observation device of a cell culture system in one embodiment. DETAILED DESCRIPTION

[0027] The following discloses a variety of different implementation methods or embodiments of the subject technical solution. To simplify the disclosure, specific examples of each element and arrangement are described below. Of course, these are only examples and are not intended to limit the scope of protection of the present invention. For example, the first feature subsequently recorded in the specification is formed above or on the second feature, which may include an implementation method in which the first and second features are formed by direct connection, and may also include an implementation method in which an additional feature is formed between the first and second features, so that the first and second features may not be directly connected. In addition, the figure marks and / or letters may be repeated in different examples in these disclosures. This repetition is for simplicity and clarity, and does not itself represent the relationship between the various implementation methods and / or structures to be discussed. Further, when the first element is described in a manner connected or combined with the second element, the description includes an implementation method in which the first and second elements are directly connected or combined with each other, and also includes the use of one or more other intervening elements to indirectly connect or combine the first and second elements.

[0028] It should be understood that, according to the actual situation, the preceding or following steps are not necessarily performed precisely in order. Other steps may also be added to these processes, or one or more steps may be removed from these processes.

[0029] like Figure 1 As shown, the cell culture system 100 may include a mixer 1 and a bioreactor 2, which are separately arranged and connected through a pipeline. In addition, the cell culture system 100 may also include a control unit to control the mixer 1 and the bioreactor 2 to perform relevant steps of cell culture.

[0030] refer to Figure 2As shown, the mixer 1 here means a device for mixing substances required for cell culture, for example, it can be gas-liquid mixing, or liquid-liquid mixing. The mixer 1 includes a mixing container 11 and a mixing drive 12. The mixing container 11 provides a sealed mixing space, which can be communicated with the outside world through a first interface 111 and a second interface 112. The first interface 111 and the second interface 112 can be used for liquid inlet and liquid outlet, respectively. For example, the first interface 111 can be used to add culture medium, cleaning fluid, etc. or liquid flowing out of a bioreactor, and the second interface 112 can be used to flow out the mixed culture medium, cleaning fluid, etc. The mixing drive 12 can drive the substances to be mixed to move and mix. In certain embodiments, the mixing container can be a tank or bag structure, and its material can be metal, glass or plastic. The specific form of the mixing drive 12 can be a stirring paddle, a magnetic stirrer, or a shaking table arranged outside the mixing container, which can be arranged inside the mixing container.

[0031] Preferably, if the culture medium in the mixer 1 is a mixture of liquid and gas, the mixing container 11 is generally further provided with a fifth interface and a sixth interface for gas inlet and gas outlet. The gas entering the fifth interface (gas inlet) may be, for example, CO 2 , O 2 、N 2 Mixed gas, but not limited to this. The sixth interface (gas outlet) can be connected to a filter or set with a waterproof breathable membrane to balance the internal and external air pressure. When the internal pressure of the mixing container is greater than the atmospheric pressure, the gas in the mixing container releases gas to the outside through the gas outlet to achieve the pressure in the mixing container is equal to the atmospheric pressure. At the same time, due to the filtering effect of the filter or the waterproof breathable membrane, external particles, pathogens, etc. will not be introduced into the cell culture system.

[0032] The mixing container 11 of the mixer 1 is used to mix gases (such as air, oxygen and carbon dioxide) and / or reaction liquids (such as culture medium, cleaning liquid or enzymatic solution) required for cell culture. 2 , O 2 、N 2 Gas mixing, or mixing culture medium with glucose, acid-base buffer, growth factors, etc.

[0033] refer to Figure 2As shown, in some embodiments, the specific structure of the mixer 1 can be that the mixing container 11 includes a mixing tank body 113 and a mixing tank cover 114, and a first interface 111 is provided on the mixing tank cover 114, and the first interface 111 is connected to the liquid inlet pipe 118 and extends into the mixing tank body 113 to achieve liquid inlet. A second interface 112 is provided below the mixing tank body 113, and a sealing gasket 115 is also provided at the connection position between the mixing tank body 113 and the mixing tank cover 114, and can be fixedly connected by bolts. A stirring paddle 116 and a corresponding motor 117 are integrated and installed on the mixing tank cover 114. The number of stirring paddles 116 is not limited to one group shown in the figure, and can also be multiple groups, for example, two groups of stirring paddles are provided along the direction of the installation axis to promote gas-liquid mixing in the mixing container, and the upper paddle can also prevent the generation of bubbles. The stirring paddle 116 is driven to rotate by the motor 117, and its rotation speed can be adjusted as needed, and the rotation speed is preferably below 200rpm. It can be understood that in some embodiments, the mixing drive 12 can also be a stirrer or a stirring paddle corresponding to a magnetic stirrer, which generates a rotating magnetic field through a motor, drives the stirring paddle or a stirring paddle inside the mixing tank to rotate, and stirs. The rotating speed can be adjusted as needed, and the rotating speed is preferably below 200rpm. In addition, in some embodiments, the mixing container can also be a mixing bag, such as a disposable mixing bag, the material of which is EVA (ethylene-vinyl acetate copolymer). The mixing bag can also have the above-mentioned first interface and second interface for liquid inlet and outlet, as well as the fifth interface and sixth interface for air inlet and outlet, and the specific role is similar to that described above, and no further description is given here. If a mixing bag is used as the mixing container 11, then the corresponding mixing drive 12 can be a shaking table, which refers to a biological shaking table here, and the specific meaning is similar to that of the biological field, and no further introduction is given here.

[0034] In addition, a temperature sensor, and / or a dissolved oxygen sensor, and / or a pH sensor may be provided inside the mixer 1 to detect various parameters of the culture medium in the mixer 1, and corresponding adjustments may be made based thereon.

[0035] refer to Figures 3 to 6 As shown, the bioreactor 2 is used to provide nutrients and environment required for the growth of adherent cells so as to carry out various biological reactions of cell culture.

[0036] The bioreactor 2 may include a reaction vessel 21 and a plurality of culture trays 22. The reaction vessel 21 provides a closed space, and the plurality of culture trays 22 are arranged in the closed space. The reaction vessel 21 may include a reaction tank body 213 and a reaction tank cover 214. The material of the reaction vessel 21 may be stainless steel or plastic, for example, polystyrene, polycarbonate, polyethylene, polypropylene, polyethylene terephthalate, polyvinyl chloride, polyvinyl pyrrolidone, polybutadiene, polyethylene oxide, polypyrrole and polypropylene oxide, etc., and may be disposable.

[0037] The closed space can be connected to the outside world through the third interface 211 and the fourth interface 212. For example, the third interface 211 is arranged at the lower part of the reaction tank body 213, and can be connected to the second interface 112 (i.e., the liquid outlet) of the mixer 1. The fourth interface 212 is arranged on the reaction tank cover 214, and can be connected to the first interface 111 (i.e., the liquid inlet) of the mixer 1, so as to provide perfusion culture of cells in the bioreactor 2. The meaning of perfusion culture here is similar to that in the art, that is, after the cells and the culture medium are added to the reactor, part of the culture medium is continuously taken out during the cell growth and product formation process, and new culture medium is continuously perfused. It can be understood that the role of the third interface 211 is not limited to connecting the second interface 112 to realize the input of the culture medium flowing out of the mixer, but can also be used to harvest the product of cell culture, and this function will be described in detail later. The function of the fourth interface 212 located at the upper part is also not limited to outputting the culture medium to the first interface 111 of the mixer 1, for example, it can also be an interface for inoculating cells, adding cleaning solution, enzymolysis solution, etc.

[0038] Define the direction from the third interface 211 to the fourth interface 212 as the first direction. Figure 3 , Figure 4 In the direction of the arrows from bottom to top shown in FIG, a plurality of culture trays 22 are adjacently distributed in a first direction, and the culture trays 22 may be disc-shaped, but are not limited thereto. Figure 5 As shown, at least part of the culture tray 22 has a through hole 221, and the through hole 221 runs through the thickness of the culture tray 22 in the first direction. Preferably, all the culture trays 22 have a through hole 221. The through hole 221 allows cells or liquid in the reaction tank to pass through, which is conducive to the uniform distribution of cells or culture medium among the culture trays 22. Preferably, reference Figure 5 As shown, the distribution of the through holes 221 on the culture plate 22 may be that there are multiple circles of through holes 221 distributed from the center to the periphery of the culture plate 22, and each circle has multiple through holes. For example, the culture plate 22 may be circular, and then there are multiple circles of through holes 221 distributed from the center to the circumference, and the size of each circle of through holes is substantially the same, and the outer circle has more through holes due to its longer length.

[0039] The material of the cell culture dish 22 can be selected from materials with good biocompatibility such as polystyrene, polycarbonate, polyethylene, polypropylene, polyethylene terephthalate, polyvinyl chloride, polyvinyl pyrrolidone, polybutadiene, polyethylene oxide, polypyrrole and polypropylene oxide, glass, etc., and can be a disposable consumable. Preferably, one or both sides of the cell culture dish 22 can be treated with surface TC, so as to be more conducive to the attachment and growth of adherent cells.

[0040] Through the above structure, especially the structure in which the culture system 100 is connected through the mixer 1 and the bioreactor 2, especially the third interface 211 can be connected to the second interface 112 of the mixer 1, and the fourth interface 212 can be connected to the first interface 111 of the mixer 1, perfusion culture is provided in the bioreactor 2, and the structure of the culture tray 22 with the through hole 221 is stacked in the bioreactor 2. The culture tray 22 not only provides more locations for cell growth to attach, but also its arrangement direction (for example, the up and down direction) and the structure of the through hole 221 make the culture medium dispersed in the reaction vessel better, and further optimize the flow effect of the culture medium, further enhance the effect of perfusion culture of cells in the bioreactor, realize sufficient material exchange between the culture material and the cells in the bioreactor, improve the culture effect, make the cell growth rate faster, and save the culture material, and also make the cells maintain a normal natural growth form.

[0041] The distribution structure of the plurality of culture trays 22 may be that the cell culture trays 22 are separated by supporting members, such as legs or gaskets. Figure 3 as well as Figure 4 As shown, a central axis 215 may be provided inside the reaction tank 213, and the central hole of the disc-shaped culture plate 22 passes through the central axis 215 and is fixed on the central axis to limit and separate the cell culture plates, for example, a slot structure may be provided on the central axis for fixing. Generally speaking, the interval between adjacent culture plates 22 is 0.5 mm to 10 mm, preferably 0.5 mm to 2 mm, to achieve high-density attachment and proliferation of adherent cells.

[0042] It can be understood that the reaction vessel 21 and the culture plate 22 can be split structures, which are processed separately and then assembled into the bioreactor 2. In this case, the reaction vessel 21 can generally be reused after being fully washed and disinfected, while the culture plate is a disposable consumable. However, this is not limited to the above. For example, the two can be an integrally formed structure. During manufacturing, the reaction vessel 21 and the culture plate 22 are directly manufactured as one body. In this case, there is no need to assemble the two. In this case, the entire bioreactor 2 is a disposable consumable.

[0043] refer to Figure 3As shown, in some embodiments, the bioreactor 2 further includes a heat exchange structure 23, which is disposed on the periphery of the reaction vessel 21. The heat exchange structure 23 generally functions to heat, and may be in the form of a water bath thermostatic barrel, and may include a temperature sensor and a heating device to achieve constant temperature heating of the reaction vessel 21. However, this is not intended to be limiting, and for example, the possibility of the heat exchange structure 23 cooling the reaction vessel 21 is not excluded. For example, when the ambient temperature is too high, the reaction vessel 21 may need to be cooled to keep the biological reaction at a suitable temperature.

[0044] refer to Figure 4 As shown, in some embodiments, the reaction vessel 21 further includes a pressure interface 217 for connecting to a pressure source, such as a filter or a waterproof breathable membrane, and connecting to the outside atmospheric pressure. Its function can be similar to that of the gas outlet of the mixer 1. When the internal pressure of the reaction vessel 21 is greater than the atmospheric pressure, the gas in the tank releases gas to the outside through the filter or the waterproof breathable membrane, so that the pressure inside the reaction vessel 21 is equal to the atmospheric pressure. At the same time, due to the filtering effect of the filter or the waterproof breathable membrane, external particles, bacteria, etc. will not be introduced into the reaction vessel 21. In addition, the pressure interface 217 can also be connected to other pressure sources, such as a pressurizing mechanism, to provide periodic pressure fluctuations for the internal liquid culture medium of the reaction vessel 21. The pressure fluctuations can be, for example, 110 kPa to 115 kPa, which is used to simulate the culture of cells under human blood pressure conditions. The pressurizing mechanism can be a common piston cylinder, peristaltic pump, etc., but is not limited thereto.

[0045] refer to Figure 6 As shown, in some embodiments, the reaction vessel 21 may also have an observation window 218, for example, a glass or other transparent material observation window (such as PMMA) is set on the reaction tank cover 214, and the observation window 218 provides a visible surface for at least part of the enclosed space. In addition, the culture system 100 may also include an observation device 4, which is optically connected to the visible surface provided by the above-mentioned observation window 218. For example, the observation device may be a microscope observation system for online real-time observation of cell growth status, such as cell implantation, cell division growth, cell plane distribution density, etc. The microscopic observation system is arranged above the reaction vessel 21, and the reaction vessel 21 has an observation window, and the observation window is one or more. The microscopic observation system may include a coaxial light source microscope 41, which can automatically focus and shoot the culture state of cells on the culture dish 22 through the observation window. Preferably, there are multiple observation windows. Preferably, the observation device 4 may also include a moving mechanism to drive the coaxial light source microscope 41 to move and observe the cell culture state in different observation windows. It can be understood that the observation device 4 may also be in other forms, as long as the cell production state inside the reaction vessel 21 is obtained through the observation window 218. The observation results may be static images or dynamic images, which are not limited thereto.

[0046] Continue to refer Figure 5 As shown, in some embodiments, the culture system 100 may further include a rotating mechanism 5, which can drive the bioreactor 2 to rotate around an axis. For example, it can be a one-dimensional rotary motor, providing a certain flip angle range, such as ±30°. Its function can be to make the cells uniformly inoculated by flipping and shaking during the cell inoculation stage. In the cell harvesting stage after the end of the culture stage, the adherent cells are peeled off from the culture plate 22 by flipping and disturbing the bioreactor.

[0047] It can be understood that the present application also provides a bioreactor, whose structure is the same as the bioreactor 2 of the cell culture system 100 introduced in the above embodiment, and will not be repeatedly introduced here.

[0048] According to the cell culture system 100 introduced above, the specific cell culture process can be.

[0049] Pre-cleaning: Inject the culture medium into the mixing container 11 through the pipeline, start the first peristaltic pump to rotate forward, and pump the culture medium into the reaction container 21. After the reaction container 21 is full, the culture medium flows back to the mixing container 11 from the fourth interface 212 at the top. Repeat this process for 1 hour, collect the culture medium and test it to ensure that the collected culture medium is not contaminated by bacteria or viruses.

[0050] Cell inoculation: The cell suspension is injected into the input bag, the third interface 211 of the reaction container 21 is closed, and the cell suspension is injected into the reaction container 21 from the fourth interface 212 through the input bag using the second peristaltic pump. The reaction container 21 can be driven to flip and shake for about 10 minutes by the rotating mechanism 5 to ensure uniform cell inoculation. For example, for human cells, they can be left to stand in a water bath at a constant temperature of 37°C, and the cell production can be observed by the observation device 4 to ensure that the cells are completely attached to the wall.

[0051] Cell culture: The culture medium is injected into the mixing container 11 from the first interface 111 through the pipeline, the third interface 211 of the reaction container 21 is connected with the second interface 112 of the mixing container 11, and the first peristaltic pump is started to rotate forward. The culture medium is mixed with nitrogen, air and carbon dioxide in the mixing container 11 to provide the environment and nutrients required for cell growth. The mixed culture medium is transported to the interior of the reaction container 21 through the third interface 211 at the bottom of the reaction container 21 by the first peristaltic pump. The reacted culture medium flows back to the mixing container 11 from the fourth interface 212 at the top of the reaction container 21 and is mixed with nitrogen, air and carbon dioxide again. During the culture process, the cell growth can be observed under a microscope.

[0052] During cell culture, for example, a control mechanism can be used to control the pH value in the mixer 1 and the bioreactor 2 to 7.5±0.02, the DO value to 40%±10%, and the temperature to 37±0.2°C. The flow rate of the mixer 1 can be 50 mL / min.

[0053] Harvesting cells: close the fourth interface 212 at the top of the reaction container 21, switch the third interface 211 at the bottom of the reaction container 21 to connect with the waste liquid bag, reverse the first peristaltic pump, and recover the culture medium supernatant into the waste liquid bag. Close the third interface 211 at the bottom of the reaction container 21, open the fourth interface 212 at the top of the reaction container 21, use the second peristaltic pump to inject the enzymatic solution into the reaction container 21, and drive the reaction container 21 to flip and shake by the rotating mechanism 5. After standing for a period of time (for example, about 1 minute to 5 minutes), open the third interface 211 at the bottom of the reaction container 21, and reverse the first peristaltic pump to harvest all the cell suspension into the harvest bag.

[0054] As described above, the present application further provides a cultivation method, using the cultivation system 100 described in the above embodiment, the cultivation method comprises the following steps:

[0055] After being mixed in the mixer 1 , the culture medium flows into the reaction container 21 through the second interface 112 and the third interface 211 , flows along the first direction and through at least part of the through hole 221 of the culture plate 22 , flows to the fourth interface 212 , and flows back to the mixer 1 through the first interface 11 .

[0056] In this way, sufficient material exchange between culture materials and cells is achieved, the culture effect is improved, the cell growth rate is faster, the culture materials are saved, and the cells can maintain their normal natural growth form.

[0057] In one or more embodiments of the culture method, as described above, the cell harvesting step may be to cut off the connection between the bioreactor 2 and the mixer 1, close one of the third interface 211 and the fourth interface 212 of the bioreactor (e.g., the fourth interface 212), and discharge the culture medium from the other interface (e.g., the third interface 211); then, the other interface (e.g., the third interface 211) is closed, the one interface (e.g., the fourth interface 212) is opened, and the enzymatic solution is injected; after treatment, the one interface (e.g., the fourth interface 212) is closed, the other interface is opened (e.g., the third interface 211), and the cell suspension is discharged. In this way, a relatively simple and easy-to-operate cell harvesting step is achieved, further improving the production efficiency of cells.

[0058] In summary, the beneficial effects of the cell culture system, bioreactor, and culture method introduced above include, but are not limited to, the structure of the cell culture system connecting the mixer and the bioreactor, especially the third interface can be connected to the second interface of the mixer, and the fourth interface can be connected to the first interface of the mixer, providing perfusion culture in the bioreactor. In addition, the structure of the culture trays with through holes is stacked in the bioreactor. The culture trays not only provide more locations for cell growth to attach, but also their arrangement direction and the structure of the through holes further optimize the flow effect of the culture medium, further enhance the effect of the perfusion culture of the cells in the bioreactor, realize the sufficient material exchange between the culture material and the cells in the bioreactor, improve the culture effect, make the cell growth rate faster, and save the culture material, and also make the cells maintain the normal natural growth form.

[0059] Although the utility model is disclosed as above with preferred embodiments, it is not intended to limit the utility model. Any person skilled in the art can make possible changes and modifications without departing from the spirit and scope of the utility model. Therefore, any modification, equivalent change and modification made to the above embodiments based on the technical essence of the utility model without departing from the content of the technical solution of the utility model shall fall within the protection scope defined by the claims of the utility model.

Claims

1. A cell culture system, characterized in that: include: A mixer, used for mixing substances required for cell culture, the mixer comprising a mixing container and a mixing drive, the mixing container providing a sealed mixing space, communicating with the outside through a first interface and a second interface, the mixing drive being able to drive the substances to be mixed to move and mix; A bioreactor comprises a reaction container and a plurality of culture plates, wherein the reaction container provides a closed space, and the plurality of culture plates are arranged in the closed space; the closed space is connected to the outside through a third interface and a fourth interface, the third interface can be connected to the second interface of the mixer, and the fourth interface can be connected to the first interface of the mixer, so as to provide perfusion culture in the bioreactor; the direction from the third interface to the fourth interface is defined as a first direction, and a plurality of the culture plates are adjacently distributed in the first direction, and at least some of the culture plates have through holes, and the through holes penetrate the thickness of the culture plates in the first direction.

2. The cell culture system according to claim 1, characterized in that There are multiple circles of through holes distributed from the center to the periphery of the culture dish, and each circle has multiple through holes.

3. The cell culture system according to claim 1, characterized in that The bioreactor is a disposable consumable.

4. The cell culture system according to claim 1, characterized in that The interval between adjacent culture plates is 0.5 mm to 10 mm.

5. The cell culture system according to claim 4, characterized in that The interval between adjacent culture plates is 0.5 mm to 2 mm.

6. The cell culture system according to claim 1, wherein The adjacent multiple culture trays are separated by a supporting member; or, a mounting shaft is provided in the closed space, and the adjacent multiple culture trays are fixedly disposed adjacently on the mounting shaft.

7. The cell culture system according to claim 1, wherein The reaction container also includes a pressure interface, and the pressure interface is used to connect to a pressure source.

8. The cell culture system according to claim 7, characterized in that The pressure source is a filtering mechanism or a pressurizing mechanism.

9. The cell culture system according to claim 1, wherein: It also includes a viewing window and a viewing device, wherein the viewing window provides a viewing surface for at least a portion of the enclosed space, and the viewing device is optically connected to the viewing surface.

10. The cell culture system according to claim 9, characterized in that The observation device also includes a movement mechanism.

11. The cell culture system according to claim 1, wherein The bioreactor also includes a heat exchange structure, which is arranged on the periphery of the reaction container.

12. The cell culture system according to claim 1, wherein The reaction container is connected to the plurality of culture plates separately or integrally.

13. The cell culture system according to claim 1, wherein It also includes a rotating mechanism capable of driving the bioreactor to rotate around an axis.

14. The cell culture system according to claim 1, wherein The mixing container includes a mixing tank body, the mixing drive is arranged inside the mixing tank body, and includes a stirring paddle or a stirring rod; or, the mixing container includes a mixing bag, the mixing drive is arranged outside the mixing bag and connected thereto, and the mixing drive includes an oscillating shaker.

15. A bioreactor, characterized in that: include: A reaction container and a plurality of culture plates, wherein the reaction container provides a closed space, and the plurality of culture plates are arranged in the closed space; the closed space is connected to the outside through a third interface and a fourth interface, the third interface is arranged below the reaction container, and the fourth interface is arranged above the reaction container; the direction from the third interface to the fourth interface is defined as a first direction, and a plurality of the culture plates are adjacently distributed in the first direction, and at least some of the culture plates have through holes, and the through holes penetrate the thickness of the culture plates in the first direction.

16. The bioreactor according to claim 15, characterized in that The bioreactor is used in the cell culture system according to any one of claims 1 to 14.

17. The bioreactor according to claim 15, characterized in that The bioreactor is a disposable consumable.

18. The bioreactor according to claim 15, characterized in that The reaction container is manufactured integrally with the culture dish.

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