Three-dimensional cell culture device with automatic liquid exchange function

Abstract

The present application relates to the technical field of biological cell culture, and particularly relates to a three-dimensional cell culture device with automatic liquid replacement function, comprising: a box body; a rotating assembly, a liquid replacement assembly and a control display assembly are integrally installed in the box body. The three-dimensional cell culture device with automatic liquid replacement function provided by the present application integrates microgravity simulation and liquid replacement function in a relatively closed culture box body space, ensures that the culture environment in the cell culture process has high stability by not excessively affecting the air flow, temperature and humidity environment, etc. in the culture box body during liquid replacement; the device for replacing culture solution does not need to rotate, effectively releases the culture space while reducing the energy consumption required in the rotation process, can load multiple culture bottles for three-dimensional culture and automatic liquid replacement at the same time, and has high potential for large-scale cell preparation.

Description

Technical Field

[0001] This invention relates to the field of biological cell culture technology, specifically a three-dimensional cell culture device with automatic medium exchange function. Background Technology

[0002] Cell culture often requires a stable temperature and humidity environment. Existing incubators, microgravity simulation devices, and medium-change devices are not integrated. During medium changes, the culture dishes must first be removed from the microgravity simulation device, and sometimes even completely removed from the incubator. The medium change is performed outside the incubator before returning the dishes to the incubator. This process can lead to the escape of insulating gases from the incubator, causing relatively drastic changes in internal temperature and humidity, and even incubator contamination. More importantly, during medium changes, the culture dishes are detached from the biaxial motion system for an extended period, forcibly interrupting the simulated microgravity mechanical effects on the cells, resulting in discontinuous mechanical stimulation. This uncontrollable process of medium changes severely impacts the stability of the culture environment.

[0003] With the development of cell therapy, tissue engineering and large-scale cell preparation, automated cell culture reactors are gradually becoming a trend, which emphasize continuous nutrient supply, online medium exchange and stable culture conditions in closed or semi-closed systems.

[0004] However, existing microgravity simulation culture devices generally lack deep integration of automated medium-change systems, making it difficult to simultaneously meet the requirements of continuous microgravity mechanical effects and automated online medium-change. Furthermore, automated culture reactors with medium-change capabilities lack the means to adapt to and scale up cell culture and expansion processes under simulated microgravity mechanical environments.

[0005] For example, in Chinese patent document CN104031835A, the cell replacement medium shell rotates synchronously with the cell culture medium shell. While this ensures that the microgravity mechanical effects are not interrupted during the medium replacement process, the cell replacement medium shell also rotates synchronously during non-medium replacement times, reducing the expansion space and weight of the cell culture medium shell, and also consuming more energy per unit cell culture process. Therefore, the volume and weight of the cell culture medium shell using this approach are severely limited, making it unsuitable for large-scale cell culture and medium replacement operations. It is only suitable for small-scale cell culture scenarios in the laboratory and not for large-scale cell preparation scenarios.

[0006] Similar structures are also found in Chinese patent documents with publication numbers CN117089456A, CN220132231U, and CN222476617U. If the replacement fluid device rotates synchronously with the culture device throughout the entire culture cycle, it will continuously occupy culture space and increase energy consumption, which is not conducive to large-scale cell preparation.

[0007] Therefore, it is crucial to provide a cell culture device that integrates three-dimensional cell culture and automatic medium exchange functions in a closed space, and has the potential for large-scale cell preparation. Summary of the Invention

[0008] The purpose of this invention is to provide a three-dimensional cell culture device with automatic medium exchange function to solve the problems mentioned in the background art.

[0009] To achieve the above objectives, the present invention provides the following technical solution:

[0010] A three-dimensional cell culture device with automatic medium change function includes: a box; the box is equipped with a rotating component, a medium change component and a control and display component;

[0011] The internal space of the housing is divided into a rotating cavity, a lifting cavity, and a control cavity; the rotating cavity and the lifting cavity are adjacent to each other vertically.

[0012] The rotating component is installed in the rotating cavity; the rotating component is a dual-shaft structure, on which multiple culture bottles can be installed simultaneously; by controlling the two sets of mutually perpendicular motors to work together, the rotation angle and speed of multiple culture bottles in three-dimensional space can be controlled simultaneously to achieve batch three-dimensional culture;

[0013] The liquid changing assembly is installed in the lifting chamber; the culture bottle has a cap assembly that matches the liquid changing interface of the liquid changing assembly; when the rotating assembly stops at a predetermined position, the liquid changing interface of the liquid changing assembly rises from the lifting chamber into the mouth of the culture bottle in the rotating chamber through a progressive lifting structure to perform batch liquid changing operations on multiple culture bottles.

[0014] The control and display component is installed in the control cavity and is used to control the rotation motor of the rotation component, the lifting motor and pump motor of the fluid exchange component, and the coordinated operation of other related electronic components.

[0015] Preferably, the rotating assembly includes a rotating frame and a fixed box. The rotating frame is rotatably mounted in the rotating cavity by a first synchronous motor, and the fixed box is rotatably mounted on the rotating frame by a second synchronous motor. The fixed box and the rotating frame are coplanar and perpendicular to each other, forming a double rotating axis structure.

[0016] Preferably, the fixing box is used to mount multiple culture bottles via a snap-fit ​​structure, and the central axis of the culture bottles is perpendicular to the cross-section of the fixing box.

[0017] Preferably, the culture flask has a liquid exchange chamber protruding into the flask opening at the center of the cap, with the open end of the liquid exchange chamber facing the cap and its cross-section smaller than that of the internal cavity. A perforated permeable membrane seat is fixedly disposed at the bottom of the liquid exchange chamber, and a cylindrical piston seat is fixedly disposed at the center of the bottom of the permeable membrane seat. A piston is slidably disposed inside the piston seat, and a spring is disposed between the bottom end of the piston and the inner wall of the bottom end of the piston seat. The permeable membrane is sleeved on the outer wall of the piston seat and fixedly attached to the surface of the permeable membrane seat near the inside of the flask. A through piston vent is disposed at the center of the piston.

[0018] When the spring pushes the piston outward, the head of the piston can completely block the open end of the liquid exchange chamber, sealing the internal space of the bottle; when the piston is pushed into the internal space of the liquid exchange chamber, the liquid exchange chamber is connected to the outside.

[0019] Preferably, the fluid replacement assembly includes a lift, a circulating fluid pump, connecting hoses, and a lifting platform;

[0020] The elevator and the circulating fluid pump are fixedly installed at the bottom of the lifting chamber; the lifting platform is installed at the top of the elevator; when the elevator rises, the lifting platform can rise from the lifting chamber into the rotating chamber; the connecting hose connects the circulating fluid pump to the fluid exchange head installed inside the lifting platform.

[0021] Preferably, the top of the lifting platform is detachably equipped with an abutment plate, which abuts against the cap of the culture bottle when the lifting platform rises to the target height.

[0022] Preferably, the fluid exchange head is detachably mounted on the lifting plate; a lead screw motor is provided at the center of the bottom of the lifting platform; the lifting plate is threaded onto the lead screw of the lead screw motor, and the lifting plate can be controlled to slide up and down inside the lifting platform by controlling the rotation of the lead screw motor; an extension hole is provided on the abutment plate; an abutment block is also provided at the center of the lower surface of the abutment plate, so that a gap is left between the upper surface of the lifting plate and the lower surface of the abutment plate;

[0023] When the rotating assembly rotates to the predetermined position, the lifting platform and the lifting plate rise progressively, allowing the fluid exchange head to pass through the extension hole and push the piston into the internal space of the fluid exchange chamber, thereby achieving fluid exchange docking.

[0024] Preferably, the liquid changing head is cylindrical in shape, and the outer diameter of the upper cylindrical part of the liquid changing head is consistent with the inner diameter of the open end of the liquid changing chamber on the bottle cap; an injection port and a return port are respectively opened on both sides of the central axis of the liquid changing head, and the horizontal opening of the injection port is higher than the horizontal opening of the return port; the upper cylindrical part of the liquid changing head is also provided with a liquid changing head vent corresponding to the piston vent; during the liquid changing process, the liquid changing head vent is always connected to the piston vent and the outside.

[0025] Preferably, the circulating liquid pump is internally equipped with a fixed circulating pipe and multiple dispensing pumps;

[0026] The fixed circulation pipe is divided into parallel inlet and outlet pipes, with its main interface passing through the box and connected to the interface of the nutrient solution machine; its branch interfaces are connected to the dispensing pump; the interface of the dispensing pump is detachably connected to the connecting hose; a hose through groove is provided at the bottom of the lifting platform, so that the connecting hose can pass through the bottom surface of the lifting platform and extend into the interior of the lifting platform to connect with the liquid exchange head.

[0027] During liquid replacement, the fixed circulation tube, the dispensing pump, the connecting hose, the liquid replacement head, and the culture flask form a liquid flow channel inside the box.

[0028] Preferably, the cap of the culture flask is threaded to the body of the flask.

[0029] Compared with the prior art, the beneficial effects of the present invention are:

[0030] 1. The three-dimensional cell culture device with automatic liquid changing function provided by the present invention adopts a modular partition design, in which the rotating component and the liquid changing component are respectively set in the rotating cavity and the lifting cavity of the closed box, so that the rotating function and the liquid changing function are both unified and relatively independent in space;

[0031] During non-medium change time, the descent of the medium change assembly completely releases the rotation space, allowing more culture flasks to be installed on the rotating assembly, providing the potential for large-scale cell preparation; during medium change time, without opening the chamber door, the medium change assembly progressively rises into the rotating chamber and docks with the culture flasks, realizing batch automatic medium change function inside the closed chamber.

[0032] 2. The three-dimensional cell culture device with automatic liquid changing function provided by the present invention improves the existing bottle cap structure of culture flasks and provides a corresponding liquid changing interface structure, realizing the standardization and large-scale docking process between culture flasks and liquid changing components. By controlling the motor to work together, the rotation function and liquid changing function can be seamlessly connected in time, which greatly reduces the time waste and risks caused by manual operation.

[0033] 3. The three-dimensional cell culture device with automatic medium replacement function provided by this invention does not require the device to rotate while replacing the culture medium, effectively freeing up culture space and reducing the energy consumption required for the rotation process. Currently, the design of one device's fixed box can install 16 culture flasks on one side, meaning that a single box can simultaneously complete the rotation and medium replacement of up to 32 culture flasks in a single culture cycle, greatly improving the space and energy utilization rate of the cell culture process. In the future, the number of culture flasks per box can be increased according to production needs, and it has extremely high expectations for large-scale cell preparation. Attached Figure Description

[0034] Figure 1 This is a rotating view of a three-dimensional cell culture device with automatic medium changing function.

[0035] Figure 2 A cross-sectional view of the center of a rotating chamber of a three-dimensional cell culture device with automatic fluid exchange function;

[0036] Figure 3 This is a diagram of the box's exterior.

[0037] Figure 4 This is a sectional view of the internal structure of the enclosure;

[0038] Figure 5 This is an external view of the rotating component;

[0039] Figure 6 Exploded view of the rotating component;

[0040] Figure 7 Exploded view of the fixture assembly;

[0041] Figure 8 Explosion diagram of a culture flask;

[0042] Figure 9 A cross-sectional view of the center of the culture bottle in its installed state;

[0043] Figure 10 A detailed cross-sectional view of section A of the cap assembly of the culture flask;

[0044] Figure 11 This is an external view of the fluid exchange assembly;

[0045] Figure 12 This is a schematic diagram of the fluid exchange assembly connection;

[0046] Figure 13 This is an external view of the fluid changer head;

[0047] Figure 14 Diagram showing the installation and connection of the fluid changer;

[0048] Figure 15 Cross-sectional view of the tank center in the raised state of the fluid exchange assembly;

[0049] Figure 16 This is a detailed cross-sectional view of section B of the lifting platform;

[0050] Figure 17 A cross-sectional view of the center of the front row of culture flasks during medium exchange.

[0051] Figure 18 This is a detailed cross-sectional view of point C during the fluid exchange process.

[0052] In the diagram: 1. Housing; 1a. Rotating cavity; 1b. Lifting cavity; 1c. Control cavity; 11. Front wall; 11a. Touch screen slot; 12. Side wall; 13. Rear wall; 13a. Pipe hole; 14. Horizontal partition; 14a. Lifting port; 15. Vertical partition; 16. Door; 17. First motor base; 17a. First motor slot; 18. Lifting slot; 2. Rotating assembly; 21. Rotating frame; 21a. Second shaft hole; 22. Fixing box 22a. Culture flask bottom groove; 22b. Insertion hole; 23. First synchronous motor; 24. Second synchronous motor; 25. First synchronous shaft; 26. Second synchronous shaft; 27. Second motor base; 28. Fixing plate; 28a. Culture flask fixing hole; 28b. Insertion post; 28c. Disassembly block; 29. ​​Rotary tank; 3. Liquid changing assembly; 31. Lifting mechanism; 31a. Telescopic rod; 31b. Lifting rod; 32. Circulating liquid pump; 32a 32b. Fixed circulation tube; 33a. Separating pump; 34. Connecting hose; 35a. Connecting cap; 36b. Connector; 37. Lifting platform; 38a. Hoses through groove; 39b. Connecting seat; 30c. Abutment plate; 31d. Expansion hole; 32f. Slide groove; 32g. Fluid changing head; 33a. Fluid changing head thread; 33b. Fixing ring; 34c. Injection port; 35d. Return port; 35e. Fluid changing head vent; 32g. Lifting plate; 33g. a. Lead screw threaded hole; 36b. Fluid changing head threaded hole; 36c. Slider; 37. Lead screw motor; 37a. Lead screw thread; 4. Culture flask; 41. Flask body; 41a. Protruding ring; 41b. Flask mouth thread; 42. Flask cap; 42a. Fluid changing chamber; 42b. Flask cap thread; 42c. Permeable membrane seat; 42d. Piston seat; 43. Permeable membrane; 44. Piston; 44a. Piston vent; 45. Spring; 5. Control and display assembly. Detailed Implementation

[0053] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings in the embodiments of the present invention.

[0054] Please see Figure 1 and Figure 2 The present invention provides a three-dimensional cell culture device with automatic liquid exchange function, including a box 1; a rotating component 2, a liquid exchange component 3 and a control and display component 5 are installed inside the box 1.

[0055] Specifically, such as Figure 3and Figure 4 As shown, the three side walls of the housing 1 are the front wall 11, the side wall 12, and the rear wall 13. A horizontal partition 14 is detachably installed between the front wall 11 and the rear wall 13, and a lifting port 14a is opened in the center of the horizontal partition 14. A longitudinal partition 15 is fixedly installed inside the housing 1 near the side wall 12. The horizontal partition 14 and the longitudinal partition 15 divide the internal space of the housing 1 into a rotating cavity 1a, a lifting cavity 1b, and a control cavity 1c. The rotating cavity 1a and the lifting cavity 1b are adjacent vertically, and the control cavity 1c is adjacent to the rotating cavity 1a and the lifting cavity 1b horizontally.

[0056] The rotating assembly is installed in the rotating chamber 1a. The rotating assembly 2 is a dual-shaft structure, on which multiple culture flasks 4 can be installed simultaneously. By controlling the two sets of motors with perpendicular rotating shafts to work together, the rotation angle and speed of multiple culture flasks 4 in three-dimensional space can be controlled simultaneously, realizing batch three-dimensional culture and improving cell culture efficiency.

[0057] The medium exchange assembly 3 is installed in the lifting chamber 1b; the culture flask 4 has a cap assembly that matches the medium exchange interface of the medium exchange assembly 3; when the rotating assembly 2 stops at a predetermined position, the medium exchange interface of the medium exchange assembly 3 rises from the lifting chamber 1b into the mouth of the culture flask 4 in the rotating chamber 1a through a progressive lifting structure, performing batch medium exchange operations on multiple culture flasks 4. Through the coordinated work of the medium exchange assembly 3 and the rotating assembly 2, the sudden change in the culture environment caused by opening the container for medium exchange is avoided, while improving the medium exchange efficiency and shortening the time required for medium exchange during cell culture.

[0058] The control and display component 5 is installed in the control cavity 1c and is used to control the rotation motor of the rotation component 2, the lifting motor and pump motor of the fluid exchange component 3, and the coordinated operation of other related electronic components.

[0059] Furthermore, a door 16 is provided on the front wall 11 in the area directly opposite the rotating cavity 1a. The door 16 can be opened or closed to connect or isolate the rotating cavity 1a from the external space of the chamber 1, facilitating the installation and disassembly of the culture flask 4, or the cleaning and maintenance of other facilities inside the chamber 1. The center of the door 16 is a glass window, allowing for easy observation of the interior of the chamber 1. In the embodiment illustrated in the present invention, the door 16 is axially connected to the chamber 1 to achieve rotational opening and closing; in other embodiments of the present invention, the door 16 and the chamber 1 have other connection methods to achieve more opening and closing methods, such as push-pull opening and closing or flip-opening and closing.

[0060] A touch screen groove 11a is provided on the front wall 11, directly opposite the control cavity 1c, so that the control touch screen of the control display component 5 can protrude outside the housing 1 for convenient control operation. A removable maintenance plate is installed on the side wall 12 on the other side of the control cavity 1c. When necessary, the maintenance plate can be removed to perform hardware maintenance on the control display component 5.

[0061] The rotating cavity 1a is symmetrically fixed with first motor bases 17 on both sides, and first motor slots 17a are correspondingly provided on the first motor bases 17 on both sides; one of the rotating shafts in the dual-axis rotating structure of the rotating assembly 2 is installed in the rotating cavity 1a through the first motor bases 17 and the first motor slots 17a.

[0062] Lifting grooves 18 are correspondingly provided on the front wall 11 and rear wall 13 of the lifting chamber 1b. The lifting component of the liquid changing assembly 3 is slidably disposed in the lifting groove 18. In the initial state, the liquid changing assembly 3 is completely retracted in the lifting chamber 1b, opposite to the rotation center of the rotating assembly 2. When it is necessary to change the nutrient solution in the culture bottle 4, the lifting component of the liquid changing assembly 3 is operated to rise in the lifting groove 18, so that the lifting port 14a passing through the center of the transverse partition 14 abuts against the bottle cap of the culture bottle 4 on the rotating assembly 2, thereby realizing the automatic liquid changing function.

[0063] A pipe hole 13a is also provided on the rear wall 13 corresponding to the lifting chamber 1b. The pumping component of the liquid changing component 3 is connected to the external nutrient solution machine through the pipe hole 13a to discharge the old nutrient solution and pump in the new nutrient solution.

[0064] It should be noted that necessary electrical pathways are provided inside the walls of enclosure 1, and the surface is treated with waterproof insulation to prevent leakage. The rear wall 13 corresponding to control cavity 1c is also equipped with a power interface, a data transmission and maintenance interface, and a power switch.

[0065] like Figure 5 and Figure 6 As shown, the rotating assembly 2 includes a rotating frame 21 and a fixed box 22. The rotating frame 21 is rotatably mounted in the rotating cavity 1a by a first synchronous motor 23, and the fixed box 22 is rotatably mounted on the rotating frame 21 by a second synchronous motor 24. The rotating axes of the fixed box 22 and the rotating frame 21 are coplanar and perpendicular to each other, forming a double rotating axis structure.

[0066] The fixing box 22 is used to mount multiple culture bottles 4 via a snap-fit ​​structure, with the central axis of the culture bottles 4 perpendicular to the cross-section of the fixing box 22. When the rotating frame 21 and the fixing box 22 rotate simultaneously around their respective rotation axes, they can drive the culture bottles 4 to rotate in three dimensions, simulating a microgravity environment.

[0067] Specifically, the first synchronous motor 23 is installed in the first motor slot 17a, and its output end is fixedly connected to the first synchronous shaft 25. The first synchronous shaft 25 is fixedly connected to the frame of the rotating frame 21. When the first synchronous motor 23 rotates synchronously, it drives the rotating frame 21 to rotate around its rotation axis through the first synchronous shaft 25.

[0068] A second motor mount 27 is mounted on the frame of the rotating frame 21 parallel to its rotation axis, and a second shaft hole 21a is opened perpendicular to its own rotation axis. A second synchronous motor 24 is mounted inside the second motor mount 27, with its output end extending into the second shaft hole 21a. A second synchronous shaft 26 passes through the second shaft hole 21a and is fixedly connected to the output end of the second synchronous motor 24. The fixed box 22 has a generally square structure, and the second synchronous shaft 26 is cross-shaped, passing through the fixed box 22 and being fixedly connected to its body. When the second synchronous motor 24 rotates synchronously, it drives the fixed box 22 to rotate around its rotation axis via the second synchronous shaft 26.

[0069] Because the central axis of the culture flask 4 is perpendicular to the cross-section of the fixed box 22, and the rotation axis of the fixed box 22 is perpendicular to the rotation axis of the rotating frame 21, when both the rotating frame 21 and the fixed box 22 are rotating, the culture flask 4 rotates in three dimensions in the spherical space formed by the two perpendicular rotation surfaces. By controlling the rotational speed and direction of the first synchronous motor 23 and the second synchronous motor 24, different vector components can be formed in the three-dimensional spherical space, simulating a microgravity environment for the culture flask 4 mounted on the fixed box 22.

[0070] In the illustrations of this invention, the rotating frame 21 is a rectangular frame structure and the fixed box 22 is a square structure, as an example. In other embodiments of this invention, the rotating frame 21 and the fixed box 22 can be other symmetrical structures, such as rhomboid, circular, or elliptical structures, as long as the rotation axes of the rotating frame 21 and the fixed box 22 are coplanar and perpendicular to each other. The first synchronous motor 23 is mounted on the rotation axis of the rotating frame 21, and the second synchronous motor 24 is mounted on the rotation axis of the fixed box 22. That is, the rotating frame 21 and the fixed box 22 can form two concentric and perpendicular rotational surfaces, forming a three-dimensional spherical rotational space with controllable angular velocity.

[0071] A rotating slot 29 is provided on the rotating frame 21, corresponding to the center of the first motor slot 17a. The rotating slot 29 covers the body of the first synchronous motor 23 that is exposed in the rotating cavity 1a. Necessary electrical paths are opened inside the rotating frame 21, the second motor base 27, and the rotating slot 29, and the outside is insulated.

[0072] like Figures 7-9 As shown, the fixing box 22 has several culture bottle bottom grooves 22a symmetrically opened on both sides of the box body. The culture bottle bottom grooves 22a are used to place the bottom body of the culture bottle 4. The inner diameter of the culture bottle bottom groove 22a matches the outer diameter of the bottom of the culture bottle 4, and a rubber layer is provided on the inner side to increase the friction with the outer wall of the culture bottle 4.

[0073] The fixing plate 28 matches the surface shape of the fixing box 22, and a culture flask fixing hole 28a corresponding to the center of the culture flask bottom groove 22a is formed through it. The inner diameter of the culture flask fixing hole 28a is smaller than the inner diameter of the culture flask bottom groove 22a. The fixing plate 28 is detachably installed on the surface of the fixing box 22.

[0074] Specifically, the four corners and center of the mounting box 22 are provided with insertion holes 22b, and the mounting plate 28 is provided with insertion posts 28b corresponding to the centers of the insertion holes 22b, with the insertion posts 28b and insertion holes 22b being overload-matched. The outer surface of the insertion posts 28b and the inner surface of the insertion holes 22b can be made of rough rubber material to increase the friction of the contact surfaces. When the insertion posts 28b are inserted into the insertion holes 22b, the rubber surface of the insertion posts 28b fills the inside of the insertion holes 22b, so that the four corners and center of the mounting plate 28 can be firmly installed on the surface of the mounting box 22. The four sides of the mounting plate 28 are provided with disassembly blocks 28c that protrude from the edge lines of the mounting box 22, making it easy to remove the mounting plate 28 from the mounting box 22. The disassembly blocks 28c do not affect the rotational movement of the mounting box 22 and the rotating frame 21.

[0075] The culture flask 4 includes a body 41 and a cap 42. A protruding ring 41a is provided on the bottom outer wall of the body 41, the height and outer diameter of which match the bottom groove 22a of the culture flask. The outer diameter of the other parts of the body 41 is consistent with the inner diameter of the culture flask fixing hole 28a. The protruding ring 41a is engaged between the bottom groove 22a and the fixing hole 28a, meaning the culture flask 4 is stably engaged between the fixing plate 28 and the fixing box 22.

[0076] In this invention, the fixing box 22 serves to fix the culture flasks 4 in batches, so that multiple culture flasks 4 are fixed on the same reference plane at the same time, and rotate or stop synchronously to achieve the purpose of batch three-dimensional cell culture, increase the number of cells that can be cultured in a single three-dimensional culture process, and make it possible for cell companies to supply cultured cells in batches.

[0077] In other embodiments of the present invention, the fixing plate 28 and the fixing box 22 can also be detachably connected in other ways to accommodate different rotation speeds. When higher rotation speeds are required, a more robust threaded connection can be selected between the fixing plate 28 and the fixing box 22.

[0078] like Figure 10 As shown, the cap 42 of the culture flask 4 is threadedly connected to the body 41. Specifically, the outer side of the mouth of the body 41 is provided with a mouth thread 41b, and the inner side of the cap 42 is provided with a cap thread 42b that matches the mouth thread 41b. The cap 42 is threadedly connected to the body 41 through the mouth thread 41b and the cap thread 42b.

[0079] A liquid exchange chamber 42a protruding into the bottle opening is provided at the center of the bottle cap 42. The liquid exchange chamber 42a is an open end in the direction of the bottle cap 42, and its open end cross-section is smaller than the internal cavity cross-section. The illustration of this invention uses a conical liquid exchange chamber 42a as an example.

[0080] A perforated permeable membrane seat 42c is fixedly installed at the bottom of the fluid exchange chamber 42a. A cylindrical piston seat 42d is fixedly installed at the center of the bottom of the permeable membrane seat 42c. A piston 44 is slidably installed inside the piston seat 42d. A spring 45 is installed between the bottom end of the piston 44 and the inner wall of the bottom end of the piston seat 42d. The longitudinal section of the piston 44 is an irregular T-shape, with the vertical section of the T-shape as the tail end and the horizontal section of the T-shape as the head end. The tail end of the piston 44 is engaged inside the piston seat 42d, and piston movement occurs inside the piston seat 42d. The head end of the piston 44 matches the open end section of the fluid exchange chamber 42a. When the spring 45 inside the piston seat 42d pushes the piston 44 towards the bottle cap 42, the head end of the piston 44 completely seals the open end of the fluid exchange chamber 42a, sealing the internal space of the bottle body 41. When piston 44 is pushed into the internal space of liquid exchange chamber 42a, the tail end of piston 44 compresses spring 45, and liquid exchange chamber 42a is connected to the outside, that is, the internal space of bottle body 41 is connected to the outside.

[0081] The piston 44 has a through piston vent 44a at its center, which allows the internal air pressure of the piston seat 42d to communicate with the external air pressure, making it easier for the piston 44 to be pushed in and ejected under normal air pressure conditions.

[0082] The permeable membrane 43 is fitted onto the outer wall of the piston seat 42d and fixedly attached to the surface of the permeable membrane seat 42c near the body 41. When the culture flask 4 is inverted, the medium exchange chamber 42a is an inverted conical cavity that is larger at the top and smaller at the bottom, facilitating the accumulation of liquid from the body 41 of the culture flask 4 towards the bottom of the cap 42. At this time, the cells are retained inside the body 41 by the permeable membrane 43, and the nutrient solution flows into the medium exchange chamber 42a through the permeable membrane 43 and the perforated permeable membrane seat 42c, avoiding cell loss during medium exchange.

[0083] like Figures 11-18As shown, the fluid exchange assembly 3 includes a lift 31, a circulating fluid pump 32, a connecting hose 33, and a lifting platform 34. The lift 31 and the circulating fluid pump 32 are fixedly installed at the bottom of the lifting chamber 1b. The lifting platform 34 is installed at the top of the lift 31, and its side wall cross-section is consistent with the lifting port 14a at the center of the transverse partition 14. When the center of the plane of the lifting port 14a is opposite to the intersection center of the rotation axis of the fixed box 22 and the rotating frame 21, the lifting platform 34 can rise from the lifting chamber 1b into the rotating chamber 1a when the lift 31 rises, and is opposite to the center of the fixed box 22. The connecting hose 33 connects the circulating fluid pump 32 to the fluid exchange head 35 installed inside the lifting platform 34. When the fixed box 22 is stopped at an angle with the box surface facing downwards, the lifting platform 34 rises into the rotating chamber 1a and can abut against the cap 42 of the culture bottle 4 located on the lower side. After the fluid exchange head 35 rises, it can extend into the cap 42 to complete the fluid exchange operation.

[0084] The elevator 31 has two sets of intersecting telescopic rods 31a at both ends. The top ends of the two sets of telescopic rods 31a are rotatably fitted with two lifting rods 31b. The two ends of the two lifting rods 31b can slide up and down in the lifting groove 18 inside the lifting cavity 1b.

[0085] The lifting rod 31b passes through and is fixedly installed in the connecting seat 34b at the bottom of the lifting platform 34. The connecting seat 34b is provided with a baffle along the central axis of the lifting rod 31b. The rotating end of the telescopic rod 31a, which is sleeved on the outer periphery of the lifting rod 31b, is restricted between the baffles of the connecting seat 34b, so that the telescopic rod 31a can only make circular telescopic movements in the vertical plane, and avoids the rotating head of the telescopic rod 31a from deviating from the central axis of the lifting rod 31b.

[0086] When the lifting platform 31 is operated by the control display component 5, the two sets of telescopic rods 31a extend and retract by the same length at the same time, so that the two lifting rods 31b slide the same distance in the lifting groove 18, keeping the lifting platform 34 horizontally raised and lowered as a whole.

[0087] A detachable abutment plate 34c is installed on the top of the lifting platform 34. When the lifting platform 34 rises to the target height, the abutment plate 34c abuts against the cap 42 of the culture bottle 4. The function of the abutment plate 34c is to ensure that the caps 42 of the culture bottle 4 are all in the same horizontal position in the direction of liquid exchange.

[0088] The fluid change head 35 is detachably mounted on the lifting plate 36; the lifting plate 36 is slidably disposed inside the lifting platform 34, and is consistent with the internal cross-section of the lifting platform 34, with a screw hole 36a opened in the center.

[0089] A lead screw motor 37 is installed at the bottom center of the lifting platform 34, and a lead screw thread 37a is provided on the outer wall of the upper lead screw of the lead screw motor 37. The lifting plate 36 is threaded onto the lead screw of the lead screw motor 37 through the lead screw thread hole 36a and engages with the lead screw thread 37a. By controlling the rotation of the lead screw motor 37, the lifting plate 36 can be controlled to slide up and down inside the lifting platform 34.

[0090] When the lead screw motor 37 rotates, the lifting plate 36 moves up and down relative to the lead screw motor 37 under the meshing action of the lead screw hole 36a and the lead screw thread 37a, so that the fluid changing head 35 installed on the lifting plate 36 moves up and down synchronously relative to the lifting platform 34.

[0091] Multiple extension holes 34d are provided on the abutment plate 34c, corresponding to the center of the culture flask bottom groove 22a of the fixing box 22. When the lifting plate 36 rises to the target height, the liquid changing head 35 can extend through the extension holes 34d and out of the abutment plate 34c. By controlling the display component 5 to operate the lead screw motor 37 to rotate, the lifting distance of the lifting plate 36 relative to the lifting platform 34 can be finely adjusted, thereby finely adjusting the height of the liquid changing head 35 extending out of the abutment plate 34c.

[0092] When the rotating assembly 2 rotates to the predetermined position, the lifting platform 34 and the lifting plate 36 rise progressively, allowing the liquid exchange head 35 to pass through the extension hole 34d and push the piston 44 into the internal space of the liquid exchange chamber 42a, thereby achieving liquid exchange docking.

[0093] The lifting plate 36 is equipped with sliders 36c on all four sides, and the inner wall of the lifting platform 34 is provided with corresponding grooves 34f. The sliders 36c slide in the grooves 34f, making the vertical displacement of the lifting plate 36 inside the lifting platform 34 more stable. A stop block is also provided at the center of the lower surface of the abutment plate 34c, so that the upper surface of the lifting plate 36 and the lower surface of the abutment plate 34c cannot be completely fitted together, and there is always a gap in the middle.

[0094] In embodiments of the present invention, the detachable design of the contact plate 34c makes it a replaceable accessory. By setting contact plates 34c of different heights, the height of the cap 42 in contact with the culture flask 4 can be adjusted to meet the contact and liquid replacement needs of culture flasks 4 of different heights. Furthermore, part of the side wall of the lifting platform 34 can be hollowed out or detachable, facilitating the disassembly, cleaning, or replacement of the lifting plate 36, liquid replacement head 35, and connecting hose 33 inside the lifting platform 34.

[0095] The fluid changing head 35 is cylindrical in shape. A fluid changing head thread 35a is provided on the outer wall of the lower end of the column, and a retaining ring 35b is provided between the fluid changing head thread 35a and the upper end of the column. The outer diameter of the upper end of the fluid changing head 35 is the same as the inner diameter of the open end of the fluid changing chamber 42a on the bottle cap 42, meaning the upper end of the fluid changing head 35 can penetrate the open end of the fluid changing chamber 42a and extend into its interior. A fluid changing head vent 35e, corresponding to the piston vent 44a, is formed on the upper end of the fluid changing head 35. The vertical section of the vent 35e coincides with the central axis of the piston vent 44a, extending from the center of the fluid changing head 35 to a position near the retaining ring 35b, where it becomes a horizontal section. The horizontal section of the vent 35e extends horizontally through the bottom of the upper end of the fluid changing head 35 and is always located in the gap between the upper surface of the lifting plate 36 and the lower surface of the abutment plate 34c. When the fluid exchange head 35 abuts against and pushes the piston 44 upward, the fluid exchange head vent 35e and the piston vent 44a are connected, and the fluid exchange head vent 35e and the piston vent 44a are connected to the outside through the gap between the upper surface of the lifting plate 36 and the lower surface of the abutment plate 34c, so as to ensure that the air pressure inside the piston seat 42d is consistent with the air pressure outside.

[0096] The fluid exchange head 35 has an injection port 35c and a return port 35d on both sides of its central axis. The injection port 35c and the return port 35d have horizontal openings at the upper end of the column of the fluid exchange head 35, with the horizontal opening of the injection port 35c being higher than the horizontal opening of the return port 35d.

[0097] Multiple liquid changer screw holes 36b are provided on the lifting plate 36, corresponding to the center of the culture flask bottom groove 22a of the fixing box 22. The liquid changer screw holes 36b are threadedly matched with the liquid changer thread 35a, so that the liquid changer 35 can be threadedly installed on the lifting plate 36. The length of the liquid changer thread 35a is greater than the thickness of the lifting plate 36. When the liquid changer 35 is installed on the lifting plate 36, its bottom liquid changer thread 35a is exposed outside the bottom of the lifting plate 36 and is threadedly connected to the connecting hose 33.

[0098] Connecting heads 33b are fixedly installed at both ends of the connecting hose 33. Both the hose 33 and the connecting heads 33b have two cavities inside that communicate with the injection port 35c and the return port 35d of the fluid exchange head 35. A connecting cap 33a is slidably and rotatably installed on the outer wall of the connecting head 33b. The inner wall of the connecting cap 33a is provided with a thread that matches the thread 35a of the fluid exchange head, so that the connecting head 33b can be threadedly connected to the fluid exchange head 35.

[0099] The circulating fluid pump 32 is internally equipped with a fixed circulating pipe 32a and multiple dispensing pumps 32b. The fixed circulating pipe 32a is divided into parallel inlet and outlet pipes, with its main interface passing through the pipe hole 13a on the rear wall 13 and connecting to the nutrient solution machine outside the housing 1. Its branch interfaces are connected to the dispensing pumps 32b. During fluid replacement, under the action of the pumps of the external nutrient solution machine, the new nutrient solution flows into the dispensing pumps 32b through the inlet pipe of the main interface of the fixed circulating pipe 32a. The dispensing pumps 32b pump the new nutrient solution into the target culture bottle 4, while simultaneously pumping the old nutrient solution flowing out of the target culture bottle 4 into the outlet pipe of the fixed circulating pipe 32a and collecting it back into the collection device.

[0100] The interface of the separatory pump 32b is configured with the same size and specifications to detachably connect to the two tubing cavities of the connector 33b, and is also threaded to the connector cap 33a with the same thread specifications. That is, the interface of the separatory pump 32b is detachably connected to the connecting hose 33.

[0101] The bottom of the lifting platform 34 is provided with a hose through groove 34a, so that the connecting hose 33 can pass through the bottom surface of the lifting platform 34 and extend into the interior of the lifting platform 34 to connect with the fluid changing head 35.

[0102] When culture flask 4 is inverted, the medium exchange chamber 42a is wider at the top and narrower at the bottom, trapping cells in the upper flask body 41 through membrane 43. When the medium exchange head 35 abuts against the piston seat 42d, causing its tail end to completely retract into the piston seat 42d, the horizontal openings of the inlet 35c and return 35d extend completely into the medium exchange chamber 42a, and the open end of the medium exchange chamber 42a is blocked by the column of the medium exchange head 35. As new nutrient solution is continuously injected into the flask body 41 from the inlet 35c, the old nutrient solution flows out from the return 35d.

[0103] In this invention, the insertion hole 22b and the insertion post 28b ensure that the center of the fixing box 22 coincides with the center of the fixing plate 28, thereby ensuring that the center of the culture bottle bottom groove 22a and the center of the culture bottle fixing hole 28a coincide. The lifting port 14a provided on the transverse partition 14 ensures that the rising center of the lifting platform 34 coincides with the center of the fixing box 22, and the side wall of the lifting platform 34 ensures that the center of the abutment plate 34c and the lifting plate 36 coincide, thereby ensuring that the extension hole 34d and the liquid changing head screw hole 36b coincide with the central axis of the culture bottle bottom groove 22a.

[0104] The number of connecting hoses 33 and medium-changing heads 35 is the same as the number of culture flasks 4 that require medium replacement. Connecting hoses 33 and medium-changing heads 35 that do not require medium replacement can be removed from the lifting plate 36, and the corresponding interfaces on the dispensing pump 32b are sealed with a cap. The cap has a similar structure to the connector 33b, and the internal tubing can be solid. During medium replacement, the fixed circulation tube 32a, dispensing pump 32b, connecting hoses 33, medium-changing heads 35, and culture flasks 4 form a liquid flow channel inside the housing 1 for the entry of new nutrient solution and the return of old nutrient solution.

[0105] When using this invention, the rotation parameters and time for microgravity simulation are set on the touch screen, and the microgravity simulation program is started, which will enable the rotating component 2 to rotate automatically and in coordination.

[0106] When a medium change is needed, the automatic medium change program is activated, pausing the microgravity simulation program. The stopping angles of the first synchronous motor 23 and the second synchronous motor 24 are adjusted synchronously, ensuring that the culture flask 4 requiring medium change faces the transverse partition 14. According to the specifications of the culture flask 4, the lifting platform 34 is raised through the lifting port 14a until the contact plate 34c abuts against the cap 42. Then, the lead screw motor 37 rotates, raising the lifting plate 36. The medium change head 35 pushes the piston 44 at the center of the cap 42 until the tail end of the piston 44 is completely retracted into the piston seat 42d. At this point, both the injection port 35c and the return port 35d at the upper end of the medium change head 35 extend into the medium change chamber 42a. The corresponding dispensing pump 32b then starts working, pumping out the old nutrient solution from the culture flask 4 and pumping in the new nutrient solution.

[0107] After the liquid replacement is completed, the dispensing pump 32b stops working. The lead screw motor 37 rotates in the reverse direction, causing the lifting plate 36 to descend, and the liquid replacement head 35 to retract from the bottle cap 42. The piston 44, pushed by the spring 45, re-closes the bottle cap 42. After the liquid replacement head 35 has completely retracted into the lifting platform 34, the lifting mechanism 31 is controlled to retract the entire lifting platform 34 back into the lifting chamber 1b. Then the microgravity simulation program is resumed, causing the rotating frame 21 and the fixed box 22 to rotate again.

[0108] Cell culture cycles typically last for more than ten days, during which the nutrient solution needs to be changed every few days. Each medium change usually takes only a few minutes, and the brief pause in rotation during the medium change process generally does not affect the overall microgravity culture effect. This invention provides a three-dimensional cell culture device with automatic medium changing function. It integrates microgravity simulation and medium changing functions within a relatively enclosed culture chamber, ensuring that the medium changing process does not excessively affect the airflow, temperature, and humidity environment inside the culture chamber, thus maintaining a highly stable culture environment during cell culture. Furthermore, the medium replacement device does not need to rotate; instead, it adopts a downward-divided modular design and provides standardized batch medium changing interfaces and culture flasks, effectively freeing up culture space while reducing the energy consumption required for the rotation process, demonstrating high potential for large-scale cell preparation.

Claims

1. A three-dimensional cell culture device with automatic medium change function, characterized in that, include: Box (1); The box (1) is equipped with a rotating assembly (2), a liquid changing assembly (3) and a control and display assembly (5). The internal space of the housing (1) is divided into a rotating cavity (1a), a lifting cavity (1b) and a control cavity (1c); the rotating cavity (1a) and the lifting cavity (1b) are adjacent vertically. The rotating component (2) is installed in the rotating cavity (1a); the rotating component (2) is a dual-shaft structure, on which multiple culture bottles (4) can be installed simultaneously; by controlling the two sets of motors with mutually perpendicular rotating shafts to work together, the rotation angle and speed of multiple culture bottles (4) in three-dimensional space can be controlled simultaneously to achieve batch three-dimensional culture; The liquid exchange assembly (3) is installed in the lifting chamber (1b); the culture bottle (4) has a cap assembly that matches the liquid exchange interface of the liquid exchange assembly (3); when the rotating assembly (2) is stopped at a predetermined position, the liquid exchange interface of the liquid exchange assembly (3) is raised from the lifting chamber (1b) into the mouth of the culture bottle (4) in the rotating chamber (1a) through a progressive lifting structure to perform batch liquid exchange operations on multiple culture bottles (4); The control and display component (5) is installed in the control cavity (1c) and is used to control the rotation motor of the rotation component (2), the lifting motor and pump motor of the liquid exchange component (3), and the coordinated operation of other related electronic components.

2. The three-dimensional cell culture device with automatic medium change function according to claim 1, characterized in that, The rotating assembly (2) includes a rotating frame (21) and a fixed box (22). The rotating frame (21) is rotatably mounted in the rotating cavity (1a) by a first synchronous motor (23). The fixed box (22) is rotatably mounted on the rotating frame (21) by a second synchronous motor (24). The rotating axes of the fixed box (22) and the rotating frame (21) are coplanar and perpendicular to each other, forming a double rotating axis structure.

3. A three-dimensional cell culture device with automatic medium change function according to claim 2, characterized in that, The fixing box (22) is used to install multiple culture bottles (4) through a snap-fit ​​structure, and the central axis of the culture bottle (4) is perpendicular to the cross-section of the fixing box (22).

4. A three-dimensional cell culture device with automatic medium change function according to claim 3, characterized in that, The culture flask (4) has a liquid exchange chamber (42a) protruding into the center of the cap (42). The liquid exchange chamber (42a) is open in the direction of the cap (42), and its open end cross-section is smaller than the internal cavity cross-section. A perforated permeable membrane seat (42c) is fixedly provided at the bottom of the liquid exchange chamber (42a). A cylindrical piston seat (42d) is fixedly provided at the center of the bottom of the permeable membrane seat (42c). A piston (44) is slidably provided inside the piston seat (42d). A spring (45) is provided between the bottom end of the piston (44) and the inner wall of the bottom end of the piston seat (42d). The permeable membrane (43) is sleeved on the outer wall of the piston seat (42d) and fixedly attached to the surface of the permeable membrane seat (42c) near the inside of the bottle body (41). A through piston vent (44a) is provided at the center of the piston (44). When the spring (45) pushes the piston (44) outward, the head end of the piston (44) can completely block the open end of the liquid exchange chamber (42a) and seal the internal space of the bottle body (41); when the piston (44) is pushed into the internal space of the liquid exchange chamber (42a), the liquid exchange chamber (42a) is connected to the outside.

5. A three-dimensional cell culture device with automatic medium change function according to claim 4, characterized in that, The fluid exchange assembly (3) includes a lift (31), a circulating fluid pump (32), a connecting hose (33), and a lifting platform (34). The elevator (31) and the circulating fluid pump (32) are fixedly installed at the bottom of the lifting chamber (1b); the lifting platform (34) is installed at the top of the elevator (31); when the elevator (31) rises, the lifting platform (34) can rise from the lifting chamber (1b) into the rotating chamber (1a); the connecting hose (33) connects the circulating fluid pump (32) to the fluid exchange head (35) installed inside the lifting platform (34).

6. A three-dimensional cell culture device with automatic medium change function according to claim 5, characterized in that, The top of the lifting platform (34) is detachably equipped with an abutment plate (34c). When the lifting platform (34) rises to the target height, the abutment plate (34c) abuts against the cap (42) of the culture bottle (4).

7. A three-dimensional cell culture device with automatic medium change function according to claim 6, characterized in that, The fluid exchange head (35) is detachably mounted on the lifting plate (36); a screw motor (37) is provided at the center of the bottom of the lifting platform (34); the lifting plate (36) is threaded onto the screw of the screw motor (37), and the lifting plate (36) can be controlled to slide up and down inside the lifting platform (34) by controlling the rotation of the screw motor (37); an extension hole (34d) is provided on the abutting plate (34c); an abutting block is also provided at the center of the lower surface of the abutting plate (34c), so that there is a gap between the upper surface of the lifting plate (36) and the lower surface of the abutting plate (34c); When the rotating assembly (2) rotates to a predetermined position, the lifting platform (34) and the lifting plate (36) rise progressively, so that the liquid exchange head (35) can pass through the extension hole (34d) and push the piston (44) into the internal space of the liquid exchange chamber (42a) to achieve liquid exchange docking.

8. A three-dimensional cell culture device with automatic medium change function according to claim 7, characterized in that, The liquid changing head (35) is cylindrical in shape. The outer diameter of the upper end of the liquid changing head (35) is consistent with the inner diameter of the open end of the liquid changing chamber (42a) on the bottle cap (42). The liquid changing head (35) has an injection port (35c) and a return port (35d) on both sides of the central axis. The horizontal opening of the injection port (35c) is higher than the horizontal opening of the return port (35d). The upper end of the liquid changing head (35) is also provided with a liquid changing head vent (35e) corresponding to the piston vent (44a). During the liquid changing process, the liquid changing head vent (35e) is always connected to the piston vent (44a) and the outside.

9. A three-dimensional cell culture device with automatic medium change function according to claim 8, characterized in that, The circulating liquid pump (32) is internally equipped with a fixed circulating pipe (32a) and multiple dispensing pumps (32b); The fixed circulation pipe (32a) is divided into parallel inlet pipe and outlet pipe, and its main interface passes through the box (1) and is connected to the interface of the nutrient solution machine; its branch interface is connected to the dispensing pump (32b); the interface of the dispensing pump (32b) is detachably connected to the connecting hose (33); the bottom of the lifting platform (34) is provided with a hose through groove (34a) so that the connecting hose (33) can pass through the bottom surface of the lifting platform (34) and extend into the interior of the lifting platform (34) to connect with the liquid exchange head (35). During liquid replacement, the fixed circulation tube (32a), the dispensing pump (32b), the connecting hose (33), the liquid replacement head (35), and the culture flask (4) form a liquid flow channel inside the box (1).

10. A three-dimensional cell culture device with automatic medium change function according to claim 1, characterized in that, The cap (42) of the culture flask (4) is threadedly connected to the body (41).

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