Automated cell culture device and method

Abstract

The present invention belongs to the field of cell culture technology, and particularly relates to an automated cell culture device and method. In the device, a peristaltic pump and a plurality of electromagnetic tube clamps are installed on the front side of a partition A, and connectors are installed on the left and right ends of the peristaltic pump, respectively. The connector is used to connect to the pipeline of a liquid storage bag, and the electromagnetic tube clamp is used to clamp the pipeline of the liquid storage bag. The front side of the partition B is rotatably connected with a swing frame, and the front and rear sides of the swing frame are respectively rotatably connected to the inner side wall of the box body and the partition B. The inside of the swing frame is rotatably connected with a fixed seat, and the left and right sides of the fixed seat are respectively rotatably connected to the inner side wall of the swing frame. Two receiving grooves are arranged on the top of the fixed seat, and an adapter plate is clamped in the receiving groove. A fixed groove is arranged on the top of the adapter plate, and a heating plate is installed on the top of the open cover, the front side of the box body, the left and right sides of the box body, and the partition B. The device can realize automated cell culture, reduce the requirements for environmental cleanliness, avoid cell contamination, and improve cell inoculation uniformity.

Description

Technical Field

[0001] The present invention belongs to the technical field of cell culture, and in particular relates to an automated cell culture device and method. Background Art

[0002] Cell culture refers to a method of simulating the in vivo environment (sterility, suitable temperature, pH and certain nutritional conditions, etc.) in vitro to enable cells to survive, grow, reproduce and maintain their main structures and functions. Cell culture technology is an important and commonly used technology in cell biology research methods. Through cell culture, we can not only obtain a large number of cells, but also study cell signal transduction, cell anabolism, cell growth and proliferation, etc.

[0003] At present, the existing cell culture equipment has a low degree of automation, and operators are required to manually add a variety of cell culture reagents into the cell culture bottle during the culture process to operate the cells. The manual operation is cumbersome, and this culture method needs to be carried out in a clean environment, otherwise cell contamination is likely to occur, which increases the production cost of the factory. In addition, the current cell culture process is prone to uneven shaking of the cell culture bottle, resulting in the problem of uneven distribution of the cultured cells. Summary of the invention

[0004] The present invention provides an automated cell culture device and method to solve the above problems.

[0005] The technical solution of the present invention is achieved in this way:

[0006] An automated cell culture device, comprising a box, the top of which is hinged with an opening cover, a partition A and a partition B are respectively installed inside the box from top to bottom, a peristaltic pump and a plurality of electromagnetic tube clamps are installed on the front side of the partition A, left and right ends of the peristaltic pump are respectively installed with connectors, the connectors are used to connect with the pipeline of a liquid storage bag, the electromagnetic tube clamps are used to clamp the pipeline of the liquid storage bag, the peristaltic pump realizes liquid inlet and liquid discharge by forward and reverse rotation, and the electromagnetic tube clamps are used to control the on and off of the flow of liquid in the pipeline;

[0007] The front side of the partition B is rotatably connected to a swing frame, and the front and rear sides of the swing frame are respectively rotatably connected to the inner side wall of the box body and the partition B, and the interior of the swing frame is rotatably connected to a fixed seat, and the left and right sides of the fixed seat are respectively rotatably connected to the inner side wall of the swing frame, and the top of the fixed seat is provided with two receiving grooves, and an adapter plate is clamped in the receiving groove, and the top of the adapter plate is provided with a fixing groove, and the receiving groove and the fixing groove are used to install and fix two different types of cell culture bottles;

[0008] The top of the open cover, the front side of the box body, the left and right sides of the box body, and the partition B are respectively installed with heating plates.

[0009] Through the above technical scheme, the cooperation between the peristaltic pump and the electromagnetic tube clamp in the present invention can realize automatic liquid delivery to the cell culture bottle and recovery of waste liquid, avoiding cell contamination caused by multiple manual operations, and at the same time can reduce the requirements of the entire culture device on environmental cleanliness during use, thereby reducing production costs.

[0010] The swing frame and the fixing seat of the present invention are designed to rotate the cell culture bottle in both the X-axis and Y-axis directions, thereby improving the uniformity of shaking the cells in the cell culture bottle. The matching of the receiving slot, the adapter plate and the fixing slot can be compatible with two types of cell culture bottles at the same time, thereby improving the versatility of the entire culture device. The entire incubator is heated on six sides to improve the uniformity of heating during the cell culture process.

[0011] Optionally, a rotating shaft A is respectively installed on the front and rear sides of the swing frame. The rotating shaft A on the front side of the swing frame is rotatably connected to the side wall of the box body through a bearing, and the rotating shaft A on the rear side of the swing frame passes through the partition B and is transmission-connected to a motor A, and the motor A is installed on the rear side of the partition B.

[0012] Through the above technical solution, the motor A can drive the swing frame to rotate in the left and right directions, which is conducive to fully mixing the liquid in the cell culture bottle.

[0013] Optionally, a rotating shaft B is respectively installed on the left and right ends of the fixed seat, and a plurality of bearing seats are respectively installed on the left and right ends of the bottom of the fixed seat. The rotating shaft B passes through the center of the bearing seat. A motor B is installed at the bottom of the fixed seat, and the output shaft of the motor B is transmission-connected to one of the ends of the rotating shaft B.

[0014] Through the above technical solution, the motor B can drive the fixing seat to rotate in the front-rear direction, which is beneficial to fully mix the liquid in the cell culture bottle in different directions.

[0015] Optionally, a plurality of elastic buckles are installed at intervals on the inner side wall of the accommodating groove and the inner side wall of the fixing groove.

[0016] Through the above technical solution, the elastic buckle can improve the stability and firmness of the fixing and installation of the cell culture bottle by the accommodating groove and the fixing groove.

[0017] Optionally, a plurality of rotating fixing devices are installed at circumferential intervals on the top of the fixing seat and the top of the adapter plate, and the rotating fixing devices include a support rod and a rotating block, the top of the support rod is rotatably connected to the rotating block, one end of the top of the rotating block is provided with a slope, and the bottom of the support rod is fixedly connected to the top of the fixing seat and the top of the adapter plate.

[0018] Through the above technical solution, the rotating fixing device can further fix the cell culture bottle to prevent the cell culture bottle from being separated during the shaking process.

[0019] Optionally, a support frame is installed at the bottom of the fixed seat, and two parallel and spaced guide rails are installed inside the support frame. Sprocket A and motor C are respectively installed at both ends of the bottom of the fixed seat, and sprocket B is installed at the output end of motor C. An annular chain is sleeved between the sprocket A and the sprocket B, and a clamping block is fixedly connected to the annular chain, a U-shaped rod is fixedly connected to one side of the clamping block, an LED lamp is installed on the U-shaped rod, and the LED lamp is located above the accommodating groove, and a mounting plate is fixedly connected to the upper end of the clamping block, and a microscope is installed on the top of the mounting plate, the LED lamp and the camera of the microscope are arranged correspondingly above and below, the bottom of the mounting plate is slidably connected to the two guide rails, and observation holes are respectively opened at the front and rear ends of the bottom of the accommodating groove, and the microscope is located below the observation hole.

[0020] Through the above technical solution, the motor C can drive the U-shaped rod to move through the ring chain and the clamp block, and then drive the LED light and the microscope to move synchronously, so as to take pictures of the cells in the cell culture bottle, facilitate the calculation of the cell growth confluence, and judge the cell growth status.

[0021] Optionally, hooks are installed on the left and right sides of the box respectively, and the hooks include a fixed plate, and both ends of the fixed plate are rotatably connected to a movable plate, and a plurality of L-shaped openings are spaced apart at the lower end of the movable plate, and a semicircular groove is formed on the inner side wall of the L-shaped opening.

[0022] Through the above technical solution, the L-shaped opening and the semicircular groove are used to hang the liquid storage bag.

[0023] Optionally, a handle A is installed on the front side of the opening cover, and an observation window is provided on the side of the opening cover.

[0024] Through the above technical solution, the handle A is convenient for opening the cover, and the observation window is helpful for the staff to observe the liquid delivery of the cell culture bottle inside the box and the color change of the cell culture medium.

[0025] An automated cell culture method, the specific operation steps are as follows:

[0026] S1. Start: Start the entire culture device, set the temperature inside the box to 37°C, and the CO2 concentration to 5%;

[0027] S2. Installation of cell culture flask and liquid storage bag: Install the cell culture flask in the receiving tank or the fixing tank, and connect the pipelines of liquid storage bag E and liquid storage bag F to the corresponding electromagnetic tube clamps respectively;

[0028] S3, second liquid storage bag installation: after the cells in liquid storage bag A are re-thawed, the pipelines of liquid storage bag A and liquid storage bag B are connected to the connector of the peristaltic pump and the corresponding electromagnetic tube clamp respectively;

[0029] S4, cell inoculation: the peristaltic pump transfers the cell suspension in the liquid storage bag A and the cell culture medium in the liquid storage bag B to two cell culture bottles in equal parts, the swing frame is tilted, and after each layer of liquid in the cell culture bottle is evenly divided, the swing frame is reset to maintain a horizontal state;

[0030] S5. Cell observation: During the cell culture process, the camera of the microscope takes a picture of the side of the cell culture bottle every 24 hours to calculate the cell growth confluence and determine the cell harvesting time;

[0031] S6, third liquid storage bag installation: connect the pipeline of liquid storage bag C and the pipeline of liquid storage bag D to the connector of the peristaltic pump and the corresponding electromagnetic pipe clamp respectively;

[0032] S7, cell harvesting: after the peristaltic pump transfers the liquid in the cell culture bottle into the liquid storage bag E, the peristaltic pump equally distributes the cleaning solution in the liquid storage bag D into two cell culture bottles, the swing frame is tilted to clean the cells in the cell culture bottle, and after the cleaning is completed, the peristaltic pump transfers the liquid in the cell culture bottle into the liquid storage bag E again;

[0033] The peristaltic pump divides the digestion solution in the liquid storage bag C into two cell culture bottles. The swing frame is tilted and kept horizontal, and the digestion solution is evenly distributed on the bottom of the culture bottle. After the horizontal state is kept still, the peristaltic pump divides the cleaning solution in the liquid storage bag D into two cell culture bottles. The swing frame is tilted to mix the cells in the cell culture bottles, and the cell suspension in the cell culture bottles is transferred into the liquid storage bag F by the peristaltic pump.

[0034] After the second cleaning as above, the peristaltic pump transfers the cell suspension in the cell culture bottle into the liquid storage bag F.

[0035] Through the above technical scheme and the above culture method, the degree of automation is high, the cell inoculation is evenly distributed, the volume of each layer of culture medium in the cell culture bottle is consistent, the cell yield is high, and the consistency between batches is high; the closed pipeline operation eliminates cell contamination.

[0036] After adopting the above technical solution, the beneficial effects of the present invention are:

[0037] The cell culture device of the present invention can realize automated cell culture. During the cell culture process, the electromagnetic tube clamp cooperates with the peristaltic pump to realize automatic liquid addition and waste liquid recovery in the cell culture bottle, avoiding the tedious manual operation and cell contamination problems, and can also reduce the cleanliness requirements of the environment during the use of the culture device. At the same time, the swing frame and the fixed seat can be rotated and tilted in both directions along the X-axis and the Y-axis, so that the cell culture bottle can be rotated and shaken in all directions, improving the uniformity of the cells in the cell culture bottle, and improving the quality and efficiency of cell culture. BRIEF DESCRIPTION OF THE DRAWINGS

[0038] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative labor.

[0039] Figure 1 The structure of the culture device in the embodiment is shown in FIG. Figure Ⅰ ;

[0040] Figure 2 The structure of the culture device in the embodiment is shown in FIG. Figure II ;

[0041] Figure 3 The structure of the culture device in the embodiment is shown in FIG. Figure III ;

[0042] Figure 4 The structure of the culture device in the embodiment is shown in FIG. Figure IV ;

[0043] Figure 5 The structure of the swing frame and the fixed seat in the embodiment is shown in FIG. Figure Ⅰ ;

[0044] Figure 6 The structure of the swing frame and the fixed seat in the embodiment is shown in FIG. Figure II ;

[0045] Figure 7 The structure of the swing frame and the fixed seat in the embodiment is shown in FIG. Figure III ;

[0046] Figure 8 is a schematic structural diagram of a fixing seat in an embodiment;

[0047] Fig. 9 yes Figure 1 Enlarged view of point A in the middle.

[0048] Description of the accompanying drawings: 1. Box body; 2. Open cover; 3. Partition A; 4. Partition B; 5. Electromagnetic tube clamp; 6. Pipeline plate; 7. Peristaltic pump; 8. Motor A; 9. Motor B; 10. Motor C; 11. Swing frame; 12. Fixed seat; 13. Rotating axis A; 14. Rotating axis B; 15. Accommodating groove; 16. Adapter plate; 17. Fixed groove; 18. Heating plate; 19. Heating wire; 20. Bearing seat; 21. Elastic buckle; 22. Support rod; 23. Rotating block; 24. Support frame; 2 5. Protective cover; 26. Guide rail; 27. Sprocket A; 28. Sprocket B; 29. ​​Ring chain; 30. Clamp; 31. Mounting plate; 32. U-shaped rod; 33. LED light; 34. Microscope; 35. Observation hole; 36. Fixed plate; 37. Moving plate; 38. L-shaped opening; 39. Semicircular groove; 40. Observation window; 41. Handle A; 42. Handle B; 43. Adjustable bracket; 44. U-shaped notch; 45. Support column; 46. Support seat; 47. Spacer; 49. Strip hole. DETAILED DESCRIPTION

[0049] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0050] The embodiments of the present application disclose an automated cell culture device and method.

[0051] Example

[0052] according to Figures 1 to 9 As shown, an automated cell culture device includes a box body 1, and hooks are installed on the left and right sides of the box body 1, respectively. The hooks include a fixed plate 36, and both ends of the fixed plate 36 are rotatably connected to a movable plate 37, and a plurality of L-shaped openings 38 are spaced apart at the lower end of the movable plate 37. The inner side wall of the L-shaped opening 38 is provided with a semicircular groove 39, and the L-shaped opening 38 and the semicircular groove 39 are used to hang a liquid storage bag.

[0053] There are six liquid storage bags in total, namely liquid storage bag A, liquid storage bag B, liquid storage bag C, liquid storage bag D, liquid storage bag E and liquid storage bag F. Liquid storage bag A, liquid storage bag B, liquid storage bag C and liquid storage bag D are used to hold cell fluid, culture medium, digestion fluid-pancreatic enzyme, and cleaning fluid-physiological saline respectively. Liquid storage bag E and liquid storage bag F are empty bags, which are used to collect waste fluid and cell fluid respectively. Digestion fluid is pancreatic enzyme, which changes cells from an adherent state to a suspended state. The cleaning fluid has two functions. One is to dilute the digestion fluid to weaken the effect of the digestion fluid on cells; the other is to clean the cells remaining in the culture bottle.

[0054] An opening cover 2 is hinged on the top of the box body 1 , a handle A41 is installed on the front side of the opening cover 2 , and an observation window 40 is arranged on the side of the opening cover 2 .

[0055] A controller is installed at the lower front end of the box 1. The controller is provided with a display screen. The controller includes a barcode scanning system and a control system. The barcode scanning system is used to scan the barcodes of items used in the cell culture process and transmit the barcode information to the control system. The controller, the barcode scanning system and the control system are all prior arts and are not within the scope of protection of the present invention, and will not be described in detail here.

[0056] The four corners of the bottom of the box body 1 are threadedly connected with adjustable brackets 43, and the adjustable brackets 43 can adjust the levelness of the entire box body 1. A plurality of U-shaped notches 44 are provided on the side of the bottom of the box body 1.

[0057] The interior of the box 1 is respectively installed with partitions A3 and B4 from top to bottom, a peristaltic pump 7 is installed on the front side of the partition A3, and connectors are installed on the left and right ends of the peristaltic pump 7, respectively, and the connectors are used to connect with the pipelines of different liquid storage bags, and two rows of electromagnetic tube clamps 5 are installed on the front side of the partition A3, and multiple electromagnetic tube clamps 5 in each row are arranged at intervals, and the electromagnetic tube clamps 5 are used to fix and clamp the pipelines of multiple liquid storage bags. Two pipeline plates 6 are installed on the front side of the partition A3, which are arranged in parallel and at intervals, and each row of electromagnetic tube clamps 5 corresponds to the pipeline plate 6 one by one. The peristaltic pump 7 is used to transport the liquid in the liquid storage bag connected thereto, and the peristaltic pump 7 realizes the liquid inlet and outlet by forward and reverse rotation, and the electromagnetic tube clamp 5 is used to control the on and off of the flow of liquid in the pipeline.

[0058] The peristaltic pump 7 and the electromagnetic tube clamp 5 are both prior art and will not be described in detail here.

[0059] A cell culture station is installed inside the box 1, and the cell culture station is located on the front side of the partition B4. The cell culture station includes a swing frame 11. Handles B42 are provided at the left and right ends of the top of the swing frame 11. Rotating shafts A13 are respectively installed on the front and rear sides of the swing frame 11. The ends of the two rotating shafts A13 are rotatably connected to the partition B4 and the side wall of the box 1 through bearings. One end of the rotating shaft A13 passes through the partition B4 and is transmission-connected to a motor A8. The motor A8 is located on the rear side of the partition B4. The motor A8 drives the rotation of the rotating shaft A13 to drive the entire swing frame 11 to rotate in the left and right directions.

[0060] Four vertically distributed support columns 45 are installed on the rear side of the partition B4. The four support columns 45 are distributed in a cube. Two of the support columns 45 are fixedly connected to the partition B4, and the bottoms of the remaining two support columns 45 are fixedly connected to the bottom inner side of the box body 1. The top of the four support columns 45 is detachably connected to a support seat 46 by bolts, and the side of the support seat 46 is fixedly connected to a motor A8. The output shaft of the motor A8 passes through the support seat 46, the partition B4 and the rotating shaft A13 for transmission connection.

[0061] A fixed seat 12 is installed inside the swing frame 11, and a rotating shaft B14 is installed at the left and right ends of the fixed seat 12 respectively. A number of bearing seats 20 are installed at the left and right ends of the bottom of the fixed seat 12 respectively. One end of the rotating shaft B14 is rotatably connected to the side wall of the swing frame 11 through a bearing, and the other end of the rotating shaft B14 passes through the center of the bearing seat 20. A motor B9 is installed at the bottom of the fixed seat 12, and the output shaft of the motor B9 is transmission-connected to the end of one of the rotating shafts B14. The motor B9 drives the rotation of the rotating shaft B14 to drive the entire fixed seat 12 to rotate in the front and rear directions.

[0062] The design of motor A8 and motor B9 enables the fixing seat 12 to rotate both left and right as well as front and back, that is, the cell culture bottle can rotate both left and right as well as front and back, thereby ensuring that the cell suspension in the cell culture bottle can be fully mixed during the cell culture process, thereby improving the efficiency and uniformity of cell culture.

[0063] A partition bar 47 is provided in the middle of the interior of the fixing seat 12, and the partition bar 47 divides the interior of the fixing seat 12 into two receiving grooves 15. The receiving grooves 15 are used to install and fix the cell culture bottles. A plurality of elastic buckles 21 are installed at intervals on the inner side walls of the receiving grooves 15. The elastic buckles 21 deform and squeeze the side walls of the cell culture bottles located in the receiving grooves 15, thereby fixing the cell culture bottles in the receiving grooves 15. A plurality of rotating fixing devices are installed at intervals along the circumferential direction on the top of the fixing seat 12, and three rotating fixing devices are distributed around each receiving groove 15.

[0064] An adapter plate 16 is installed inside the receiving groove 15, and the side wall of the adapter plate 16 is in contact with the elastic buckle 21, and a fixing groove 17 is provided on the top of the adapter plate 16. A plurality of elastic buckles 21 are arranged at intervals along the circumferential direction on the inner side wall of the fixing groove 17, and a plurality of rotating fixing devices are installed on the top of the adapter plate 16, and the rotating fixing devices are distributed along the periphery of the fixing groove 17. The receiving groove 15 and the fixing groove 17 are used to fix and install two types of cell culture flasks, namely, T175 cell culture flasks and five-layer cell factories, respectively.

[0065] The rotating fixing device includes a support rod 22 and a rotating block 23. The top of the support rod 22 is rotatably connected to the rotating block 23. An inclined surface is provided at one end of the top of the rotating block 23. The bottom of the support rod 22 is fixedly connected to the top of the fixing seat 12 and the top of the adapter plate 16. The distance between the rotating block 23 and the top of the fixing seat 12, and the distance between the rotating block 23 and the adapter plate 16 match the thickness of the cell culture bottle to be fixed.

[0066] A support frame 24 is installed at the bottom of the fixed seat 12, and two parallel guide rails 26 are installed inside the support frame 24. A protective cover 25 is installed at the bottom of the fixed seat 12. The support frame 24 is located inside the protective cover 25. A plurality of waist-shaped holes are opened at the bottom of the protective cover 25. The waist-shaped holes are located above the water tray for heat diffusion. After the water in the water tray evaporates, it enters through the waist-shaped holes for heat dissipation. A strip hole 49 is opened on the rear side of the protective cover 25. A connecting rod is fixedly connected to one end of the bottom of the fixed seat 12, and a sprocket A27 is rotatably connected to the lower end of the connecting rod. A motor C10 is fixedly connected to the other end of the bottom of the fixed seat 12, and a sprocket B28 is installed at the output end of the motor C10. An annular chain 29 is sleeved between the sprocket A27 and the sprocket B28. The annular chain 29 is fixedly connected with a clamping block 30. A U-shaped rod 32 is fixedly connected to one side of the clamping block 30. The U-shaped rod 32 is vertically arranged. The upper end of the U-shaped rod 32 is located above the fixing seat 12. An LED lamp 33 is installed on the upper end of the U-shaped rod 32. The lower end of the U-shaped rod 32 passes through the bar hole 49 and is fixedly connected to the clamping block 30. The upper end of the clamping block 30 is fixedly connected with a mounting plate 31. A microscope 34 is installed on the top of the mounting plate 31. The LED lamp 33 and the camera of the microscope 34 are arranged correspondingly up and down. Two sliders are fixedly connected to the bottom of the mounting plate 31. The two sliders are respectively slidably connected to the two guide rails 26. Observation holes 35 are respectively opened at the front and rear ends of the bottom of the accommodating groove 15, and the microscope 34 is located below the observation hole 35.

[0067] A water tray is installed on the inner side wall of the bottom of the box body 1. The water tray is used to hold sterile water. The water tray is located below the cell culture station.

[0068] The box body 1 is provided with a heating plate 18 with heating on six sides. The top of the open cover 2, the lower end of the front side of the box body 1, the lower ends of the left and right sides of the box body 1, and the rear side of the partition B4 are respectively installed with heating plates 18. The heating plate 18 is provided with a heating wire 19, and the heating wire 19 is distributed in a continuous U-shaped bend. The heating wire 19 indirectly heats the cell culture bottle by heating the air inside the box body 1. The heating wire 19 controls the start and stop of heating through the control system. The circuit of the heating wire 19 adopts a dual-circuit heating method, including a main silicone insulated heating wire and a spare silicone insulated heating wire. Both heating wires 19 are controlled by the control system to form a dual-circuit heating method. When the main silicone insulated heating wire fails, the control system can start the spare silicone insulated heating wire for heating, so that the heating plate 18 has two heating modules, which are spare for each other.

[0069] An automated cell culture method comprises the following steps:

[0070] S1. Start up: Start the whole culture device, add 1L of sterile water to the water tray, modify the parameters of the culture device: set the temperature to 37°C, and the CO2 concentration to 5%; heat the heating wire 19 to keep the temperature of the whole culture at 37°C. Adding sterile water to the water tray can keep the humidity inside the culture device ≥90%RH.

[0071] Scanning barcodes: After the consumables (cell culture bottles and liquid storage bags) are unpacked, the barcode scanning system scans the barcodes on the consumables, and the barcode information is entered into the control system;

[0072] S2. Installation of consumables: The cell culture flask is installed and fixed using the receiving groove 15 or the fixing groove 17 according to the model, the elastic buckle 21 clamps the side wall of the cell culture flask, the rotating block 23 presses the top of the cell culture flask by rotation, the pipeline plate 6 and the electromagnetic tube clamp 5 are installed, and the pipelines of the liquid storage bag E and the pipelines of the liquid storage bag F are respectively installed using a sterile pipe connection machine, and the pipelines of the liquid storage bag E and the liquid storage bag F are respectively installed in the corresponding electromagnetic tube clamps 5, and the liquid storage bag E and the liquid storage bag F are hung on the hook of the equipment.

[0073] S3, second liquid storage bag installation: After the cells in liquid storage bag A are thawed, use a sterile pipe connection machine to connect and install the pipelines of liquid storage bag A and liquid storage bag B, and install the pipelines of liquid storage bag A and liquid storage bag B with the connector of the peristaltic pump and the corresponding electromagnetic pipe clamp 5 respectively;

[0074] S4, cell inoculation: click on the cell inoculation program of the control system, input the volume of the cell suspension and the volume of the cell culture medium, the electromagnetic tube clamp 5 holds the liquid storage bag A together with the cell culture bottle, and the cell suspension in the liquid storage bag A is equally divided into two cell culture bottles through the peristaltic pump 7; the electromagnetic tube clamp 5 switches to hold the liquid storage bag B together with the cell culture bottle, and the cell culture medium in the liquid storage bag B is equally divided into two cell culture bottles through the peristaltic pump 7; the motor A drives the swing frame 11 to rotate and shake, and the motor B drives the fixed seat 12 to rotate and shake, and the rotation angles of the swing frame 11 and the fixed seat 12 are both between -30° and 30°. After the cell culture bottle shakes evenly, the motor A drives the swing frame 11 to rotate 90°. After each layer of liquid in the cell culture bottle is evenly divided, the motor A drives the swing frame 11 to reset and keep it horizontal;

[0075] S5. Cell observation: During the cell culture process, the camera of the microscope 34 takes pictures every 24 hours, and the control system calculates the cell growth confluence. The operator determines the cell harvesting time according to the confluence;

[0076] S6, third liquid storage bag installation: Use a sterile pipe connection machine to connect and install the pipelines of liquid storage bag C and liquid storage bag D respectively, and install the pipelines of liquid storage bag C and liquid storage bag D with the connector of the peristaltic pump and the corresponding electromagnetic pipe clamp 5 respectively;

[0077] S7. Cell harvesting: Click the cell harvesting program of the control system, input the volume of liquid and cleaning solution, and transfer the liquid in the cell culture bottle into the liquid storage bag E through the peristaltic pump 7; the electromagnetic tube clamp 5 switches to keep the liquid storage bag D together with the cell culture bottle, and divides the cleaning solution in the liquid storage bag D into two cell culture bottles through the peristaltic pump 7, the motor A drives the swing frame 11 to rotate and shake, and the motor B drives the fixed seat 12 to rotate and shake. The rotation angles of the swing frame 11 and the fixed seat 12 are both between -30° and 30°, and the cells in the cell culture bottle are washed. After the washing is completed, the cell culture bottle shakes evenly, and the motor A drives the swing frame 11 to rotate 90°, and transfers the liquid in the cell culture bottle into the liquid storage bag E through the peristaltic pump 7; the electromagnetic tube clamp 5 switches to keep the liquid storage bag C together with the cell culture bottle, and divides the digestion solution in the liquid storage bag C into two cell culture bottles through the peristaltic pump 7, and the motor A Drive the swing frame 11 to rotate and shake, and the motor B drives the fixed seat 12 to rotate and shake. The rotation angles of the swing frame 11 and the fixed seat 12 are both between -30° and 30°. Keep it horizontal, and the digestive solution is evenly distributed on the bottom of the culture bottle. Let it stand horizontally for 3-5 minutes. Switch the electromagnetic tube clamp 5 to keep the liquid storage bag D together with the cell culture bottle, and divide the cleaning solution in the liquid storage bag D into two cell culture bottles by the peristaltic pump 7. Drive the swing frame 11 to rotate and shake, and the motor B drives the fixed seat 12 to rotate and shake. The rotation angles of the swing frame 11 and the fixed seat 12 are both between -30° and 30°. Mix the cells in the cell culture bottle. Drive the swing frame 11 to rotate 90° by the motor A, and transfer the cell suspension in the cell culture bottle into the liquid storage bag F by the peristaltic pump 7. Perform a second cleaning as above, transfer the cell suspension in the cell culture bottle into the liquid storage bag F by the peristaltic pump 7, and the cell harvesting is completed.

[0078] Liquid storage bag A, liquid storage bag B, liquid storage bag C, liquid storage bag D, liquid storage bag E and liquid storage bag F are connected to the cell culture bottle and the connector of the peristaltic pump through sterile connectors and pipelines according to the sequence requirements in the culture process, so as to transport the liquid.

[0079] In the description of the present invention, it should be understood that the terms "longitudinal", "lateral", "upper", "lower", "front", "back", "left", "right", "vertical", "horizontal", "top", "bottom", "inside", "outside" and the like indicate orientations or positional relationships based on the orientations or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the present invention and simplifying the description, rather than indicating or implying that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore cannot be understood as a limitation on the present invention. In the description of the present invention, unless otherwise specified and limited, it should be noted that the terms "installed", "connected" and "connected" should be understood in a broad sense, for example, it can be a mechanical connection or an electrical connection, or it can be the internal communication of two elements, it can be a direct connection, or it can be an indirect connection through an intermediate medium. For ordinary technicians in this field, the specific meanings of the above terms can be understood according to specific circumstances.

[0080] The above are only preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. An automated cell culture device, characterized in that: It comprises a box body, the top of which is hinged with an opening cover, the interior of which is respectively installed with a partition A and a partition B from top to bottom, a peristaltic pump and a plurality of electromagnetic tube clamps are installed on the front side of the partition A, and connectors are respectively installed on the left and right ends of the peristaltic pump, the connectors are used to connect with the pipeline of the liquid storage bag, the electromagnetic tube clamps are used to clamp the pipeline of the liquid storage bag, the peristaltic pump realizes liquid inlet and liquid discharge by forward and reverse rotation, and the electromagnetic tube clamps are used to control the on and off of the flow of liquid in the pipeline; The front side of the partition B is rotatably connected to a swing frame, and the front and rear sides of the swing frame are respectively rotatably connected to the inner side wall of the box body and the partition B, and the interior of the swing frame is rotatably connected to a fixed seat, and the left and right sides of the fixed seat are respectively rotatably connected to the inner side wall of the swing frame, and the top of the fixed seat is provided with two receiving grooves, and an adapter plate is clamped in the receiving groove, and the top of the adapter plate is provided with a fixing groove, and the receiving groove and the fixing groove are used to install and fix two different types of cell culture bottles; The top of the open cover, the front side of the box, the left and right sides of the box, and the partition B are respectively installed with heating plates; The front and rear sides of the swing frame are respectively provided with a rotating shaft A, the rotating shaft A on the front side of the swing frame is rotatably connected to the side wall of the box body through a bearing, and the rotating shaft A on the rear side of the swing frame passes through the partition B and is transmission-connected to a motor A, and the motor A is installed on the rear side of the partition B; The left and right ends of the fixed seat are respectively installed with a rotating shaft B, and the left and right ends of the bottom of the fixed seat are respectively installed with a plurality of bearing seats, the rotating shaft B passes through the center of the bearing seat, and the bottom of the fixed seat is installed with a motor B, and the output shaft of the motor B is drivingly connected to the end of one of the rotating shafts B; The inner side wall of the receiving groove and the inner side wall of the fixing groove are both installed with a plurality of elastic buckles at intervals; The top of the fixing seat and the top of the adapter plate are both installed with several rotating fixing devices at intervals along the circumferential direction, and the rotating fixing devices include a support rod and a rotating block. The top of the support rod is rotatably connected to the rotating block, and one end of the top of the rotating block is provided with an inclined surface. The bottom of the support rod is fixedly connected to the top of the fixing seat and the top of the adapter plate.

2. An automated cell culture device according to claim 1, characterized in that: A support frame is installed at the bottom of the fixed seat, and two parallel and spaced guide rails are installed inside the support frame. Sprocket A and motor C are respectively installed at two ends of the bottom of the fixed seat, and sprocket B is installed at the output end of the motor C. An annular chain is sleeved between the sprocket A and the sprocket B, and a clamping block is fixedly connected to the annular chain. A U-shaped rod is fixedly connected to one side of the clamping block, and an LED lamp is installed on the U-shaped rod. The LED lamp is located above the accommodating groove, and a mounting plate is fixedly connected to the upper end of the clamping block. A microscope is installed on the top of the mounting plate. The LED lamp and the camera of the microscope are arranged correspondingly up and down, and the bottom of the mounting plate is slidably connected to the two guide rails. Observation holes are respectively opened at the front and rear ends of the bottom of the accommodating groove, and the microscope is located below the observation holes.

3. An automated cell culture device according to claim 2, characterized in that: Hooks are installed on the left and right sides of the box body respectively, and the hooks include a fixed plate, and the two ends of the fixed plate are rotatably connected to a movable plate respectively, and a plurality of L-shaped openings are spaced apart at the lower end of the movable plate, and a semicircular groove is opened on the inner side wall of the L-shaped opening.

4. The automated cell culture device according to claim 1, characterized in that: A handle A is installed on the front side of the opening cover, and an observation window is arranged on the side of the opening cover.

5. An automated cell culture method, characterized in that: The cell culture is carried out using the culture device of claim 3, and the specific operation steps are as follows: S1. Start: Start the entire culture device, set the temperature inside the box to 37°C, and the CO2 concentration to 5%; S2. Installation of cell culture flask and liquid storage bag: Install the cell culture flask in the receiving tank or the fixing tank, and connect the pipelines of liquid storage bag E and liquid storage bag F to the corresponding electromagnetic tube clamps respectively; S3, second liquid storage bag installation: after the cells in liquid storage bag A are thawed, the pipelines of liquid storage bag A and liquid storage bag B are connected to the connector of the peristaltic pump and the corresponding electromagnetic tube clamp respectively; S4. Cell inoculation: The peristaltic pump transfers the cell suspension in the liquid storage bag A and the cell culture medium in the liquid storage bag B to two cell culture bottles in equal parts. The rotation angles of the swing frame and the fixed seat are between -30° and 30°. After the cell culture bottles are shaken evenly, the swing frame rotates 90°. After each layer of liquid in the cell culture bottles is evenly divided, the swing frame is reset to maintain a horizontal state. S5. Cell observation: During the cell culture process, the camera of the microscope takes a picture of the side of the cell culture bottle every 24 hours to calculate the cell growth confluence and determine the cell harvesting time; S6, third liquid storage bag installation: connect the pipeline of liquid storage bag C and the pipeline of liquid storage bag D to the connector of the peristaltic pump and the corresponding electromagnetic tube clamp respectively; S7, cell harvesting: after the peristaltic pump transfers the liquid in the cell culture bottle into the liquid storage bag E, the peristaltic pump divides the cleaning solution in the liquid storage bag D into two cell culture bottles, and the rotation angles of the swing frame and the fixed seat are both between -30° and 30° to clean the cells in the cell culture bottle. After the cleaning is completed, the swing frame rotates 90°, and the peristaltic pump transfers the liquid in the cell culture bottle into the liquid storage bag E again; The peristaltic pump divides the digestion solution in the liquid storage bag C into two cell culture bottles, and the rotation angles of the swing frame and the fixed seat are between -30° and 30°. After keeping horizontal, the digestion solution is evenly distributed on the bottom of the culture bottle. After the horizontal state is calm, the peristaltic pump divides the cleaning solution in the liquid storage bag D into two cell culture bottles, and the rotation angles of the swing frame and the fixed seat are between -30° and 30°. The cells in the cell culture bottles are mixed, and the swing frame rotates 90°, and the cell suspension in the cell culture bottle is transferred into the liquid storage bag F by the peristaltic pump; After the second cleaning as above, the peristaltic pump transfers the cell suspension in the cell culture bottle into the liquid storage bag F.

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