A low-shear chaotic flow field bioreactor and a method for culturing adherent cells.

Abstract

This invention provides a low-shear chaotic flow field bioreactor and a method for culturing adherent cells. The low-shear chaotic flow field bioreactor includes a tank, a stirring device, and a control device. The stirring device includes a drive mechanism and a stirring mechanism, which is disposed inside the tank. The stirring mechanism includes an Oloid-shaped impeller. The drive mechanism is kinetically connected to the Oloid-shaped impeller and can drive the Oloid-shaped impeller to perform a composite three-dimensional motion including revolution around the central axis of the tank and rotation around its own axis. The control device is electrically connected to the drive mechanism and is used to control the drive mechanism. This invention not only constructs a low-shear chaotic flow field, effectively protecting mechanically sensitive cells, but also achieves uniform mixing throughout the entire process, eliminating sedimentation "dead zones." Simultaneously, it significantly broadens the process window for cell culture, adapting to the needs of large-scale cell culture.

Description

Technical Field

[0001] This invention relates to the field of bioengineering equipment technology, specifically to a low-shear chaotic flow field bioreactor and a method for culturing adherent cells. Background Technology

[0002] In the field of cell therapy, the use of bioreactors for three-dimensional microcarrier culture of adherent cells (such as mesenchymal stem cells) is a key technology for achieving large-scale production. Its core requirement is to achieve efficient cell expansion and standardized culture while ensuring high cell activity and purity, thus laying the foundation for the industrial application of cell therapy products.

[0003] Currently, the most commonly used bioreactors in the industry employ traditional paddle agitators, such as three-bladed elephant ear paddles. These agitators work by using rotating blades to cut the fluid, generating convection and turbulence to mix the materials inside the tank, thereby suspending three-dimensional microcarriers and providing a uniform growth environment for adherent cells. However, this traditional agitation method has inherent and insurmountable drawbacks: the resulting flow field is essentially two-dimensional and periodic, with a simple and predictable structure, leading to significant differences in velocity gradients and highly uneven shear force distribution within the tank.

[0004] Specifically, a significant high shear force zone is formed in areas such as the tip of the blades of a traditional paddle mixer. The peak shear force in this zone is too high. Since most adherent cells (such as mesenchymal stem cells) are mechanically sensitive, excessive shear force can easily cause mechanical damage to the cells, disrupting cell morphology and cell membrane integrity, and even leading to apoptosis or undesirable differentiation, seriously affecting the quality of cell culture and subsequent application effects.

[0005] Meanwhile, in order to keep the three-dimensional microcarriers in a stable suspended state throughout the tank and avoid the formation of sedimentation "dead zones" at the bottom and walls of the tank, and to ensure full contact between cells and nutrients, existing technologies often require increasing the stirring speed. However, increasing the stirring speed will further exacerbate the velocity gradient of the flow field, further increasing the shear force value in the high shear force zone, further aggravating cell damage, and creating a "dilemma".

[0006] In summary, due to the limitations of flow field characteristics, the existing stirring systems of bioreactors have always faced a fundamental contradiction between "uniform mixing throughout the entire process" and "ultra-low shear throughout the entire process". This results in an extremely narrow process window for cell culture, making it difficult to balance the suspension effect of microcarriers, the uniformity of material mixing and the protection of cell viability. As a result, it cannot meet the actual needs of large-scale, high-quality culture of adherent cells, thus restricting the industrialization of cell therapy technology. Summary of the Invention

[0007] The purpose of this invention is to provide a low-shear chaotic flow field bioreactor and a method for culturing adherent cells, so as to solve the above-mentioned technical problems existing in the prior art; the preferred technical solutions among the many technical solutions provided by this invention can produce many technical effects, as detailed below.

[0008] To achieve the above objectives, the present invention provides the following technical solution:

[0009] This invention provides a low-shear chaotic flow field bioreactor, comprising a tank, a stirring device, and a control device, wherein: the stirring device includes a drive mechanism and a stirring mechanism, the stirring mechanism being disposed inside the tank, the stirring mechanism including an Oloid-shaped stirring blade, the drive mechanism being tractively connected to the Oloid-shaped stirring blade, the drive mechanism being capable of driving the Oloid-shaped stirring blade to perform a composite three-dimensional motion including revolution around the central axis of the tank and rotation around its own axis; the control device is electrically connected to the drive mechanism and is used to control the drive mechanism.

[0010] Preferably, the stirring mechanism includes a support frame, and the Oloid-shaped stirring blade is disposed on the support frame; the driving mechanism includes a revolution driving component, which is disposed on the tank body and connected to the support frame, and the revolution driving component can drive the support frame and the Oloid-shaped stirring blade to revolve around the central axis of the tank body.

[0011] Preferably, the driving mechanism includes a rotation drive assembly, which includes a rotation power assembly and a double eccentric fork linkage assembly, wherein: the rotation power assembly is fixedly mounted on the support frame, and the rotation power assembly is connected to the input end of the double eccentric fork linkage assembly; the double eccentric fork linkage assembly is provided with a first eccentric output end and a second eccentric output end, the first eccentric output end being connected to the Oloid-shaped stirring blade, and the second eccentric output end being connected to the Oloid-shaped stirring blade.

[0012] Preferably, the Oloid-shaped stirring blades are made of a biocompatible material.

[0013] Preferably, the ratio of the rotational speed to the revolution speed of the Oloid-shaped stirring blade is a non-integer ratio.

[0014] Preferably, the non-integer ratio is 1.2 to 1.8.

[0015] Preferably, the tank is a transparent tank; Preferably, the control device includes a human-machine interface screen.

[0016] This invention provides a method for culturing adherent cells using any of the aforementioned low-shear chaotic flow field bioreactors, comprising at least the following steps: Step 1: Add adherent cells and cell base into the container; Step two, during the cell culture stage, the ratio of the rotation speed and revolution speed of the Oloid-shaped stirring blade is set to a non-integer ratio by a control device.

[0017] Preferably, in step two, the cell culture stage includes a cell seeding stage and a cell proliferation stage, wherein: During the cell seeding stage, the first set of revolution speed and rotation speed parameters were used for continuous operation. During the cell proliferation phase, the second set of revolution speed and rotation speed parameters were used for continuous operation.

[0018] Preferably, the first set of revolution speed and rotation speed parameters are as follows: The revolution speed of the first set of parameters is 5~10 rpm; The rotational speed of the first set of parameters is 8~15 rpm.

[0019] Preferably, the second set of revolution speed and rotation speed parameters are as follows: The second set of parameters has a revolution speed of 10~20 rpm; The rotation speed of the second set of parameters is 15~30 rpm.

[0020] The low-shear chaotic flow field bioreactor and the method for culturing adherent cells provided by this invention have at least the following beneficial effects: I. Constructing a low-shear chaotic flow field to effectively protect mechanically sensitive cells: The stirring mechanism of this invention employs an Oloid-shaped impeller, and the drive mechanism enables the impeller to perform a composite three-dimensional motion of revolving around the central axis of the tank and rotating around itself. This breaks through the limitations of the two-dimensional, periodic flow field generated by traditional paddle stirrers. The Oloid-shaped structure itself has the characteristics of being without sharp edges and streamlined, which can avoid the formation of local high shear force zones at the impeller tips. At the same time, the composite three-dimensional motion makes the movement trajectory of the impeller within the tank unpredictable, forming a chaotic flow field. This significantly reduces the velocity gradient within the tank, ensuring that the shear force is evenly distributed throughout the entire range and maintained at an ultra-low level. This fundamentally avoids the mechanical damage of high shear force to adherent cells (such as mesenchymal stem cells), effectively protecting the integrity of cell morphology and cell viability, reducing the occurrence of apoptosis and undesirable differentiation, and ensuring the quality of cell culture.

[0021] II. Achieve uniform mixing across the entire area and eliminate settlement "dead zones": The compound three-dimensional motion of the Oloid-shaped stirring blades drives the fluid inside the tank to flow in a multi-directional, irregular, chaotic manner. The fluid movement covers the entire tank area, eliminating dead zones. Compared to traditional paddle stirrers, this invention achieves stable suspension of three-dimensional microcarriers without increasing the stirring speed. This effectively avoids the formation of sedimentation "dead zones" on the bottom and walls of the tank, ensuring uniform dispersion of the microcarriers throughout the culture system. It also guarantees sufficient contact between nutrients, oxygen, and cells, improving cell proliferation efficiency and culture uniformity. This solves the dilemma in existing technologies where "increasing the speed protects suspension, but increasing the speed damages cells."

[0022] III. Expanding the process window to meet the needs of large-scale cell culture: This invention achieves a synergistic unity of "globally uniform mixing" and "globally ultra-low shear" through the construction of a chaotic flow field, significantly broadening the process window for cell culture. The control device can precisely control the operating parameters of the drive mechanism according to different stages of cell culture (such as cell adhesion and proliferation), adjusting the revolution and rotation speeds of the stirring blades to adapt to the needs of different growth stages of adherent cells, providing a stable and suitable growth environment for the cells. Simultaneously, this structure is adaptable to tanks of different sizes, meeting the industrialization needs of large-scale, standardized culture of adherent cells, and promoting the industrial application of cell therapy technology.

[0023] IV. Simple structure and convenient control reduce cultivation costs: This invention achieves its core technical effects simply by optimizing the structure and movement of the stirring mechanism. The overall structure is simple, requiring no additional complex auxiliary devices, thus reducing equipment manufacturing and maintenance costs. The control device is electrically connected to the drive mechanism, enabling precise control of stirring motion parameters. It is easy to operate, reduces human error, improves the reproducibility and stability of cell culture, and further reduces the overall cost of large-scale culture. Attached Figure Description

[0024] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0025] Figure 1 This is a schematic diagram of the structure of the low-shear chaotic flow field bioreactor of the present invention; Figure 2 This is a schematic diagram of the structure of the tank body of the present invention; Figure 3 This is a schematic diagram of the stirring device of the present invention; Figure 4 This is a schematic diagram of the structure of the double eccentric fork connecting rod assembly and the Oloid-shaped stirring blade of the present invention; Figure 5 This is a control block diagram of the drive mechanism of the present invention; Figure 6 This is a flowchart of the method for culturing adherent cells according to the present invention.

[0026] Figure Labels 1. Tank body; 2. Agitator; 21. Drive mechanism; 211. Revolution drive assembly; 212. Rotation drive assembly; 2121. Rotation power assembly; 2122. Double eccentric fork linkage assembly; 21221. Drive shaft; 21222. Driven shaft; 21223. Driven fork linkage; 21224. Driven fork linkage; 22. Agitator; 221. Oloid-shaped agitator blades; 222. Support frame; 3. Control device; 31. Human-machine interface screen. Detailed Implementation

[0027] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be described in detail below. Obviously, the described embodiments are merely some embodiments of this invention, and not all embodiments. Based on the embodiments of this invention, all other implementation methods obtained by those skilled in the art without creative effort are within the scope of protection of this invention.

[0028] Example 1: This invention provides a low-shear chaotic flow field bioreactor, reference Figures 1 to 5 As shown, the low-shear chaotic flow field bioreactor includes a tank 1, a stirring device 2, and a control device 3.

[0029] The stirring device 2 includes a drive mechanism 21 and a stirring mechanism 22. The stirring mechanism 22 is located inside the tank 1 and includes an Oloid-shaped stirring blade 221. The output end of the drive mechanism 21 is inserted into the tank 1 and is connected to the Oloid-shaped stirring blade 221 for transmission.

[0030] Oloid-shaped stirring blade 221 refers to a geometric body composed of a convex hull surface formed by two circles with the same radius, a center distance equal to the radius, and perpendicular mid-planes. In this invention, it gently propels the fluid through an overall flipping motion to achieve mixing, which differs from the cutting action of traditional impellers.

[0031] The control device 3 is electrically connected to the drive mechanism 21 and is used to control the drive mechanism 21.

[0032] During operation, the control device 3 controls the drive mechanism 21 to move, thereby driving the Oloid-shaped stirring blades 221 to perform compound three-dimensional motion within the tank 1.

[0033] In the above process, the composite three-dimensional motion includes the revolution of the Oloid-shaped stirring blade 221 around the central axis of the tank 1, and the rotation of the blade around itself.

[0034] The stirring mechanism 22 of this invention adopts an Oloid-shaped stirring blade 221, and the stirring action includes revolution around the central axis of the tank 1 and rotation around itself, a three-dimensional composite motion. This not only creates a low-shear chaotic flow field to effectively protect mechanically sensitive cells, but also achieves uniform mixing throughout the entire area, eliminates sedimentation "dead zones", and significantly broadens the process window for cell culture to meet the needs of large-scale cell culture.

[0035] Example 2 Example 2 is based on Example 1: like Figures 1 to 5 As shown, the stirring mechanism 22 includes a support frame 222, and an Oloid-shaped stirring blade 221 is mounted on the support frame 222; the driving mechanism 21 includes a revolution driving assembly 211, which is mounted on the tank 1 and connected to the support frame 222. The revolution driving assembly 211 can drive the support frame 222 and the Oloid-shaped stirring blade 221 to revolve around the central axis of the tank.

[0036] Specifically, the revolution drive assembly 211 adopts an electric power assembly, including a motor and a reducer, which is fixedly installed on the top cover of the tank body 1, and its output shaft is inserted into the tank body 1 along the axis of the tank body 1 and connected to the support frame 222.

[0037] The orbital drive component 211 can stably drive the support frame 222 to rotate, ensuring the orbital effect of the Oloid-shaped stirring blades 221.

[0038] As an optional implementation, the drive mechanism 21 includes a rotation drive assembly 212, which includes a rotation power assembly 2121 and a double eccentric fork linkage assembly 2122. The rotation power assembly 2121 is fixedly mounted on the support frame 222 and is connected to the input end of the double eccentric fork linkage assembly 2122. The double eccentric fork linkage assembly 2122 is provided with a first eccentric output end and a second eccentric output end. The first eccentric output end is connected to the Oloid-shaped stirring blade 221, and the second eccentric output end is connected to the Oloid-shaped stirring blade 221.

[0039] This configuration ensures that the Oloid-shaped stirring blades 221 are subjected to uniform force, guaranteeing their stable and uniform rotation. It further optimizes the formation of chaotic flow fields, making the shear force distribution in the flow field more uniform and stable, and adapting to the flow field requirements of different cell culture scenarios.

[0040] Specifically, the self-rotation power assembly 2121 adopts an electric power assembly, including a motor and a reducer. The double eccentric shift fork linkage assembly 2122 includes a drive shaft 21221, a driven shaft 21222, a drive shift fork linkage 21223, and a driven shift fork linkage 21224. The first end of the drive shaft 21221 is connected to the output end of the self-rotation power assembly 2121, and the second end of the drive shaft 21221 is rotatably connected to the drive shift fork linkage 21223. The shift fork of the drive shift fork linkage 21223 is open. Both ends of the shaft are rotatably connected to the Oloid-shaped stirring blade 221. The first end of the driven shaft 21222 is rotatably mounted on the support frame 222. The second section of the driven shaft 21222 is rotatably connected to the driven fork link 21224. Both ends of the fork opening of the driven fork link 21224 are rotatably connected to the Oloid-shaped stirring blade 221. The driving fork link 21223 and the driven fork link 21224 respectively form the first eccentric output end and the second eccentric output end.

[0041] As an optional implementation, the Oloid-shaped stirring blades 221 are made of a biocompatible material.

[0042] Adherent cells (such as mesenchymal stem cells) come into indirect contact with the stirring blades during culture. Using biocompatible materials can prevent the release of harmful substances from the blades or the initiation of cellular immune responses, effectively protecting cell viability and morphological integrity, and preventing cell damage, apoptosis, or abnormal differentiation due to contact with non-biocompatible materials. At the same time, biocompatible materials usually have good corrosion resistance and biostability, can adapt to the culture medium environment during cell culture, are not prone to material degradation or shedding, avoid contamination of the culture system, ensure the purity and safety of cell culture, meet the biosafety requirements for culture equipment in the field of cell therapy, and provide material-level assurance for cell culture quality.

[0043] Specifically, the Oloid-shaped impeller 221 is made of medical-grade 316L stainless steel or TC4 titanium alloy through precision machining.

[0044] As an optional implementation, the ratio of the rotational speed to the revolution speed of the Oloid-shaped stirring blade 221 is a non-integer ratio.

[0045] The non-integer ratio refers to the ratio of the rotational speed to the revolution speed of the Oloid-shaped stirring blade 221 being a non-integer value. The purpose of this setting is to break the periodicity of the stirring motion, thereby generating a non-periodic and unpredictable chaotic flow field in the reactor.

[0046] The chaotic flow field refers to a non-periodic fluid motion state in which streamlines are fully stretched and folded in space. It is generated by composite motions with non-integer ratios, which can achieve global uniform mixing at low speeds while maintaining fluid shear stress at a low level, thus providing a suitable mechanical microenvironment for cells.

[0047] Specifically, the non-integer ratio is 1.2 to 1.8, which is an optimized range obtained through fluid dynamics simulation and experimental verification. Within this range, the generation efficiency of the chaotic flow field is relatively high, enabling uniform mixing of the entire tank in a shorter time. At the same time, the energy input (i.e., rotational speed) required by the agitator can be maintained at a low level, thereby synergistically ensuring a low-shear environment.

[0048] Preferably, the non-integer ratio is set to 1.618.

[0049] As an optional implementation, the tank 1 is set as a transparent tank; this allows operators to easily observe the cell culture status, microcarrier suspension and flow field changes inside the tank 1 in real time, and complete visual monitoring without opening the tank 1, avoiding contamination of the culture system caused by opening the lid. At the same time, it facilitates the timely detection of abnormalities in the culture process (such as microcarrier sedimentation, cell aggregation, etc.), and timely adjustment of culture parameters, thereby improving the controllability of cell culture.

[0050] Specifically, tank 1 is made of polycarbonate (PC) material.

[0051] PC material is characterized by high strength, impact resistance, high temperature resistance, and chemical corrosion resistance, making it suitable for the sterilization and temperature control requirements in cell culture processes. It is also non-toxic, odorless, and will not contaminate the culture system.

[0052] As an optional implementation, the control device 3 includes a human-machine interface screen 31, which is disposed on the tank wall of the tank body 1.

[0053] The human-machine interface screen 31 enables the visualization, real-time display, and precise control of parameters such as stirring speed (rotation and revolution). It is convenient and intuitive to operate, reduces human error, and allows operators to quickly adjust parameters according to different stages of cell culture, improving operational efficiency, adapting to the standardized operation requirements in large-scale culture, and reducing the workload of operators.

[0054] Example 2: Example 2 is based on Example 1: like Figure 6As shown, the present invention provides a method for culturing adherent cells using the aforementioned low-shear chaotic flow field bioreactor, aiming to apply the hydrodynamic characteristics of the reactor to actual biological culture and solve the problems of cell damage and low growth efficiency caused by stirring in existing culture methods.

[0055] The method for culturing adherent cells includes at least the following steps: Step 1: Add adherent cells and cell base into container 1; Step 2: During the cell culture stage, the ratio of the rotation speed and revolution speed of the Oloid-shaped stirring blade 221 is adjusted to a non-integer ratio by the control device 3, thereby generating and maintaining a low-shear chaotic flow field in the tank 1, providing a suitable mechanical microenvironment for cell attachment and proliferation.

[0056] As an optional implementation, in step two, the cell culture stage includes a cell seeding stage and a cell proliferation stage, wherein: During the cell seeding stage, the first set of revolution speed and rotation speed parameters were used for continuous operation. During the cell proliferation phase, the second set of revolution speed and rotation speed parameters were used for continuous operation.

[0057] The purpose of this design is that during the inoculation phase, cells are relatively fragile and require a mild environment to facilitate stable attachment to the microcarriers; while during the proliferation phase, the increased cell number and accelerated metabolism necessitate stronger mixing to ensure adequate nutrient supply and timely removal of metabolic waste. By setting different parameters for different phases, precise control of the culture process is achieved, which helps improve cell attachment efficiency, survival rate, and final yield.

[0058] As an optional implementation, the first set of revolution speed and rotation speed parameters are specifically as follows: The revolution speed of the first set of parameters is 5~10 rpm; The rotational speed of the first set of parameters is 8~15 rpm.

[0059] The continuous and gentle stirring method during the cell seeding stage avoids periodic shear shocks and local accumulation during the stationary phase, thus improving the uniformity of adhesion and the early survival rate of cells.

[0060] As an optional implementation, the second set of revolution speed and rotation speed parameters are specifically as follows: The second set of parameters has a revolution speed of 10~20 rpm; The rotation speed of the second set of parameters is 15~30 rpm.

[0061] Increasing the rotation speed appropriately during the cell proliferation phase can enhance macroscopic mixing and mass transfer efficiency, meeting the needs of high-density culture. Thanks to the inherent low-shear characteristics of chaotic flow fields, even with increased rotation speed, the shear stress in the entire flow field can still be maintained at a low level that is safe for cells.

[0062] In the description of this application, it should be understood that the terms "upper", "lower", "inner", "outer", "top", "bottom", etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are only for the convenience of describing this application and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on this application.

[0063] Furthermore, the terms "first" and "second" are used for descriptive purposes only and should not be construed as indicating or implying relative importance or implicitly specifying the number of technical features indicated. Thus, a feature defined as "first" or "second" may explicitly or implicitly include at least one of that feature. In the description of this application, "multiple" or "several" means at least two, such as two, three, etc., unless otherwise explicitly specified.

[0064] In this application, unless otherwise expressly specified and limited, the terms "installation," "connection," "linking," "fixing," etc., should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral part; they can refer to a direct connection or an indirect connection through an intermediate medium; they can refer to the internal communication of two components or the interaction between two components, unless otherwise expressly limited. Those skilled in the art can understand the specific meaning of the above terms in this application according to the specific circumstances.

[0065] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A low-shear chaotic flow field bioreactor, characterized in that, Includes a tank, a stirring device, and a control device, wherein: The stirring device includes a drive mechanism and a stirring mechanism. The stirring mechanism is disposed inside the tank and includes an Oloid-shaped stirring blade. The drive mechanism is connected to the Oloid-shaped stirring blade and can drive the Oloid-shaped stirring blade to perform a composite three-dimensional motion including revolution around the central axis of the tank and rotation around itself. The control device is electrically connected to the drive mechanism and is used to control the drive mechanism.

2. The low-shear chaotic flow field bioreactor according to claim 1, characterized in that, The stirring mechanism includes a support frame, and the Oloid-shaped stirring blades are disposed on the support frame; The driving mechanism includes a revolution driving component, which is disposed on the tank and connected to the support frame. The revolution driving component can drive the support frame and the Oloid-shaped stirring blade to revolve around the central axis of the tank.

3. The low-shear chaotic flow field bioreactor according to claim 2, characterized in that, The drive mechanism includes a rotation drive assembly, which comprises a rotation power assembly and a double eccentric shift fork linkage assembly, wherein: The self-rotation power component is fixedly mounted on the support frame, and the self-rotation power component is connected to the input end of the double eccentric shift fork linkage assembly; The double eccentric fork linkage assembly is provided with a first eccentric output end and a second eccentric output end. The first eccentric output end is connected to the Oloid-shaped stirring blade, and the second eccentric output end is connected to the Oloid-shaped stirring blade.

4. The low-shear chaotic flow field bioreactor according to claim 1, characterized in that, The Oloid-shaped agitator blades are made of biocompatible materials.

5. The low-shear chaotic flow field bioreactor according to claim 1, characterized in that, The ratio of the rotational speed to the revolution speed of the Oloid-shaped stirring blade is a non-integer ratio; The non-integer ratio is 1.2 to 1.

8.

6. The low-shear chaotic flow field bioreactor according to claim 1, characterized in that, The tank is designed to be transparent. The control device includes a human-machine interface screen.

7. A method for culturing adherent cells using a low-shear chaotic flow field bioreactor as described in any one of claims 1 to 6, characterized in that, At least the following steps are included: Step 1: Add adherent cells and cell base into the container; Step two, during the cell culture stage, the ratio of the rotation speed and revolution speed of the Oloid-shaped stirring blade is set to a non-integer ratio by a control device.

8. The method for culturing adherent cells according to claim 7, characterized in that, In step two, the cell culture stage includes a cell seeding stage and a cell proliferation stage, wherein: During the cell seeding stage, the first set of revolution speed and rotation speed parameters were used for continuous operation. During the cell proliferation phase, the second set of revolution speed and rotation speed parameters were used for continuous operation.

9. The method for culturing adherent cells according to claim 8, characterized in that, The specific parameters for the first set of revolution speed and rotation speed are as follows: The revolution speed of the first set of parameters is 5~10 rpm; The rotational speed of the first set of parameters is 8~15 rpm.

10. The method for culturing adherent cells according to claim 8, characterized in that, The specific parameters for the second set of revolution speed and rotation speed are as follows: The second set of parameters has a revolution speed of 10~20 rpm; The rotation speed of the second set of parameters is 15~30 rpm.

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