Device for culturing NK (Natural Killer) cells in vitro

Abstract

The utility model discloses an in-vitro NK cell culture device, and belongs to the technical field of cell culture, and the in-vitro NK cell culture device is characterized by comprising a culture box with a controller, a cell culture system arranged in the culture box, used for culturing NK cells, quantitatively adding a complete culture medium into the culture box and maintaining the proliferation requirement of the NK cells, and electrically connected with the controller; the cell monitoring system is arranged on the incubator, communicates with the cell culture system, is used for sampling regularly and detecting the growth state, concentration and activity of the NK cells, and is electrically connected with the controller, and the system has the effects of improving the culture efficiency, stabilizing the culture environment, achieving intelligent management and being free of pollution.

Description

Technical Field

[0001] This application relates to the field of cell culture technology, and more specifically to an in vitro NK cell culture device. Background Technology

[0002] Natural killer cells (NK cells) are important immune cells in the body, possessing a wide range of immune functions, including anti-tumor and antiviral activity. In recent years, with the rise of cell immunotherapy, the application prospects of NK cells have become increasingly broad, especially in the fields of cancer immunotherapy and antiviral therapy, where NK cells have gradually attracted attention as an immunotherapeutic tool.

[0003] Currently, large-scale in vitro culture and expansion of NK cells mainly relies on manual replacement of culture flasks / bags and the artificial addition of cytokines and culture media. During cell culture, cells are cultured in small-volume flasks, then transferred to large-volume flasks, and finally to large cell culture bags. This requires multiple replacements of cell culture containers of different capacities to accommodate the cell culture volume, which easily leads to cell contamination.

[0004] In view of this, the inventors have designed an in vitro NK cell culture device to solve the above problems. Utility Model Content

[0005] To address the shortcomings of existing technologies, the purpose of this invention is to provide an in vitro NK cell culture device. Its advantages are that it effectively solves the problems of easy contamination and cumbersome operation in artificial cell culture by culturing NK cells in a semi-automatic manner, thereby improving culture efficiency, providing a stable culture environment, and enabling intelligent management.

[0006] To achieve the above objectives, this utility model provides the following technical solution: an in vitro NK cell culture device, comprising an incubator with a controller.

[0007] The cell culture system, set inside an incubator, is used to culture NK cells and quantitatively add complete culture medium to the incubator. It has a constant temperature setting function for NK cell culture and growth factors to maintain the proliferation requirements of NK cells and is electrically connected to the controller.

[0008] The cell monitoring system, installed on the incubator and connected to the cell culture system, is used to periodically sample and detect the growth status, concentration, and activity of NK cells, and is electrically connected to the controller.

[0009] By adopting the above technical solution, when NK cell culture is required, the incubator is first placed in a sterile environment or sterile room. Then, the cell culture system and cell monitoring system are set through the control box to maintain a relatively constant temperature for the cultured NK cells and the complete culture medium. Next, the NK cells and complete culture medium are placed into the cell culture system for culture. After the cell monitoring system detects the growth status, concentration, and activity of the NK cells in the cell culture system, the detected values ​​are fed back to the controller. The controller then controls the cell culture system to add complete culture medium to the NK cells, thereby maintaining the NK cells in the cell culture system in an optimal state.

[0010] In summary, the intelligent control culture achieved by using the controller, cell culture system, and cell monitoring system on the incubator can effectively avoid contamination caused by manual addition of complete culture medium, and achieve the goals of greatly improving culture efficiency, providing a stable culture environment, and intelligent management.

[0011] The present invention is further configured as follows: the cell culture system includes a material chamber disposed on an incubator and separated from the outside world, a first temperature sensor disposed in the material chamber, a first heater disposed in the material chamber, a first cooler disposed in the material chamber, a culture medium bag disposed in the material chamber and containing complete culture medium, a mononuclear cell bag disposed in the material chamber and containing mononuclear cells, a culture chamber disposed on an incubator and separated from the outside world, a second temperature sensor disposed in the culture chamber, a second heater disposed in the culture chamber, a second cooler disposed in the culture chamber, a cell culture bag disposed in the culture chamber for culturing NK cells, and a conveying system disposed on an incubator for conveying materials in the material chamber to the culture chamber. The cell culture bag in the culture chamber is connected to a cell monitoring system. The first temperature sensor, the first heater, the first cooler, the second temperature sensor, the second heater, the second cooler, and the conveying system are all electrically connected to a controller.

[0012] By adopting the above technical solution, when NK cell culture needs to be carried out in a cell culture bag, the controller controls the first temperature sensor, the first heater, and the first cooler to maintain a constant temperature in the material chamber within a set range of 2~8°C. The controller also controls the first temperature sensor, the first heater, and the first cooler to maintain a constant temperature in the culture chamber within a set range of 37°C±1°C. The controller can control the conveying system to transport mononuclear cells from the mononuclear cell bag to the cell culture bag. Then, according to the set value, the conveying system transports the complete culture medium from the cell culture bag to the cell culture bag and stops working after reaching the transport value. When the cell monitoring system detects that the cell density in the cell culture bag is too high, the cell monitoring system transmits a signal to the controller, which then controls the conveying system to transport the complete culture medium from the culture medium bag to the cell culture bag and stops working after reaching the transport value, thereby effectively keeping the NK cells in the optimal state.

[0013] The present invention is further configured such that: the delivery system includes a single nucleated cell conduit disposed in the incubator and used to connect the single nucleated cell bag and the cell culture bag, a culture medium conduit disposed in the incubator and used to connect the culture medium bag and the cell culture bag, and two delivery pumps and two sealing solenoid valves respectively disposed on the single nucleated cell conduit and the culture medium conduit, wherein the two delivery pumps and the sealing solenoid valves are electrically connected to the controller.

[0014] By adopting the above technical solution, the controller controls the delivery pump and sealing solenoid valve on the mononuclear cell conduit. The delivery pump delivers mononuclear cells from the mononuclear cell bag to the cell culture bag through the mononuclear cell conduit. The delivery pump and sealing solenoid valve on the culture medium conduit deliver the complete culture medium from the culture medium bag to the cell culture bag through the culture medium conduit, and stop working after the delivery value is reached. When the cell monitoring system detects that the cell density in the cell culture bag is too high, the cell monitoring system transmits a signal to the controller, and the controller controls the delivery pump and sealing solenoid valve on the culture medium conduit to deliver the complete culture medium from the culture medium bag to the cell culture bag through the culture medium conduit, and stop working after the delivery value is reached.

[0015] The present invention is further configured such that: the opening of the mononuclear cell bag is connected to the mononuclear cell conduit via a threaded interface; the opening of the culture medium bag is connected to the culture medium conduit via a threaded interface; and each opening of the cell culture bag is connected to the corresponding mononuclear cell conduit and culture medium conduit via a threaded interface.

[0016] By adopting the above technical solution, the threaded interface facilitates the installation and sealing of the openings of the mononuclear cell bag and the culture medium bag, and the corresponding mononuclear cell conduit and culture medium conduit.

[0017] The present invention is further configured such that: the mononuclear cells in the mononuclear cell bag are separated by centrifugation using a mononuclear cell separation device, the mononuclear cell separation device comprising, from top to bottom, a PBS bag, a blood bag for holding blood, a separation bag for holding separation fluid, and a connecting tube, the PBS bag and the blood bag being connected by the connecting tube, and the blood bag and the separation bag being connected by the connecting tube.

[0018] By adopting the above technical solution, when it is necessary to separate mononuclear cells in blood, the connecting tube between the PBS bag and the blood bag is first tightly sealed with clamps to separate the PBS bag from the blood bag. Then, the mononuclear cell separation device is placed on a centrifuge. The blood in the blood bag is separated from top to bottom into a plasma layer, a white membrane layer, a separating fluid layer, and a red blood cell and granulocyte layer by centrifugation. The white membrane layer and plasma layer remain in the blood bag, while the separating fluid layer and the red blood cell and granulocyte layer remain in the separation bag. The connecting tube between the blood bag and the separation bag is separated by heat sealing. The clamps are then opened, and the PBS bag and the blood bag are centrifuged again to separate the mononuclear cells into the PBS bag. The connecting tube between the PBS bag and the blood bag is then heat sealed, and the PBS bag is separated from the blood bag, thus completing the separation of mononuclear cells. Finally, the PBS bag is assembled with the cell culture bag for use, i.e., the PBS bag is a mononuclear cell bag.

[0019] The present invention is further configured such that the cell culture bag, the mononuclear cell bag, and the culture medium bag are all made of EVA material.

[0020] The present invention is further configured such that: the cell monitoring system includes a cell viability analyzer for detecting NK cells, a detection tube disposed on an incubator for delivering NK cells and culture medium to the detection port of the cell viability analyzer, a three-way solenoid valve disposed between the detection tube and the cell culture system, and a liquid pump disposed on the side of the detection tube away from the three-way solenoid valve. One end of the detection tube is connected to the cell culture system through the three-way solenoid valve, and the other end is located at the detection port of the cell viability analyzer. The controller is electrically connected to the cell viability analyzer and the three-way solenoid valve.

[0021] By adopting the above technical solution, when it is necessary to detect NK cells in the cell culture system, the controller controls the three-way solenoid valve to connect the detection tube to the cell culture system. Then, the liquid pump extracts NK cells and culture medium from the cell culture system. After a certain amount of NK cells and culture medium are extracted, the controller controls the three-way solenoid valve to connect the detection tube to the outside. Under the action of the liquid pump, the NK cells and culture medium enter the detection port of the cell viability analyzer along the detection tube. The cell viability analyzer then detects the NK cells and culture medium and uploads the detection data to the controller.

[0022] In summary, this utility model has the following advantages:

[0023] 1. Intelligent control of culture is achieved through the controller, cell culture system, and cell monitoring system on the incubator. This effectively avoids contamination caused by the manual addition of cytokines, culture media, etc., and achieves the goals of greatly improving culture efficiency, providing a stable culture environment, and intelligent management.

[0024] 2. When mononuclear cells need to be placed in a cell culture bag for culture, the controller controls the first temperature sensor, the first heater, and the first cooler to maintain a constant temperature in the material chamber within a set range of 2~8°C. The controller also controls the first temperature sensor, the first heater, and the first cooler to maintain a constant temperature in the culture chamber within a set range of 37°C±1°C. The controller can control the delivery system to transport the mononuclear cells from the mononuclear cell bag to the cell culture bag. Then, the delivery system transports the complete culture medium from the cell culture bag to the cell culture bag and stops working after reaching the delivery value. When the cell monitoring system detects that the cell density in the cell culture bag is too high, the cell monitoring system transmits a signal to the controller, which then controls the delivery system to transport the complete culture medium from the culture medium bag to the cell culture bag and stops working after reaching the delivery value, thereby effectively keeping the NK cells in optimal condition.

[0025] 3. When it is necessary to detect NK cells in the cell culture system, the controller controls the three-way solenoid valve to connect the detection tube to the cell culture system. Then, the liquid pump extracts NK cells and culture medium from the cell culture system. After a certain amount of NK cells and culture medium are extracted, the controller controls the three-way solenoid valve to connect the detection tube to the outside. Under the action of the liquid pump, the NK cells and culture medium enter the detection port of the cell viability analyzer along the detection tube. The cell viability analyzer detects the NK cells and culture medium and uploads the detection data to the controller. Attached Figure Description

[0026] Figure 1 This is a schematic diagram of the overall structure of this embodiment;

[0027] Figure 2 This is a schematic diagram highlighting the structure of the incubator in this embodiment;

[0028] Figure 3 This is a schematic diagram of the cell culture system in this embodiment;

[0029] Figure 4 This is a schematic diagram of the single nucleus cell separation device in this embodiment.

[0030] Explanation of reference numerals in the attached diagram: 1. Incubator; 2. Controller; 3. Touchscreen display; 4. Material bin; 5. First temperature sensor; 6. First heater; 7. First cooler; 8. Mononuclear cell bag; 9. Culture medium bag; 10. Culture chamber; 11. Second temperature sensor; 12. Second heater; 13. Second cooler; 14. Cell culture bag; 15. Mononuclear cell conduit; 16. Culture medium conduit; 17. Transfer pump; 18. Cell viability analyzer; 19. Detection tube; 20. Three-way solenoid valve; 21. Liquid pump; 22. Gas sensor; 23. Carbon dioxide tube; 24. Carbon dioxide tank; 25. Exhaust pipe; 26. Gas tube solenoid valve; 27. Sealing solenoid valve; 28. PBS bag; 29. ​​Blood bag; 30. Separation bag; 31. Connecting tube. Detailed Implementation

[0031] The present invention will be further described in detail below with reference to the accompanying drawings.

[0032] An in vitro NK cell culture device, such as Figure 1 , 2 As shown, it includes an incubator 1 with a controller 2, and a touch screen 3 is installed on the controller 2.

[0033] The cell culture system is installed in the incubator 1 and is used to culture NK cells and quantitatively add complete culture medium to the incubator 1. It has a constant temperature setting function for NK cell culture and complete culture medium to maintain the proliferation requirements of NK cells and is electrically connected to the controller 2.

[0034] A cell monitoring system, installed on incubator 1 and connected to the cell culture system, is used to periodically sample and detect the growth status, concentration, and activity of NK cells, and is electrically connected to controller 2.

[0035] like Figure 1 , 2As shown, the cell culture system includes a material chamber 4 disposed on an incubator 1 and isolated from the outside; a first temperature sensor 5 disposed within the material chamber 4; a first heater 6 disposed within the material chamber 4; a first cooler 7 disposed within the material chamber 4; a culture medium bag 9 disposed within the material chamber 4 and containing complete culture medium; a mononuclear cell bag 8 disposed within the material chamber 4 and containing mononuclear cells; a culture chamber 10 disposed on an incubator 1 and isolated from the outside; a second temperature sensor 11 disposed within the culture chamber 10; a second heater 12 disposed within the culture chamber 10; a second cooler 13 disposed within the culture chamber 10; and a culture medium bag 9 disposed within the material chamber 4 and containing mononuclear cells; a culture chamber 10 disposed on an incubator 1 and isolated from the outside; a second temperature sensor 11 disposed within the culture chamber 10; a second heater 12 disposed within the culture chamber 10; and a second cooler 13 disposed within the culture chamber 10. The culture chamber 10 contains cell culture bags 14 for culturing NK cells, and a conveying system mounted on the incubator 1 for conveying materials from the material bin 4 to the culture chamber 10. The cell culture bags 14 in the culture chamber 10 are connected to a cell monitoring system. The first temperature sensor 5, the first heater 6, the first cooler 7, the second temperature sensor 11, the second heater 12, the second cooler 13, and the conveying system are all electrically connected to the controller 2. The first heater 6 and the second heater 12 are A39TH compressed air constant temperature heaters, and the first cooler 7 and the second cooler 13 are Minichiller 300 OLE. The cell culture bags 14, the mononuclear cell bags 8, and the culture medium bags 9 are all made of EVA material.

[0036] When NK cell culture needs to be performed in cell culture bag 14, the controller 2 controls the first temperature sensor 5, the first heater 6, and the first cooler 7 to maintain a constant temperature in the material chamber 4 within the set range of 2~8°C. The controller 2 also controls the first temperature sensor 5, the first heater 6, and the first cooler 7 to maintain a constant temperature in the culture chamber 10 within the set range of 37°C±1°C. The controller 2 can control the conveying system to transport mononuclear cells from the mononuclear cell bag 8 to the cell culture bag 14. Then, according to the set value, the conveying system transports complete culture medium from the culture medium bag 9 to the cell culture bag 14 and stops working after reaching the transport value. When the cell monitoring system detects that the cell density in the cell culture bag 14 is too high, the cell monitoring system transmits a signal to the controller 2, and the controller 2 controls the conveying system to transport complete culture medium from the culture medium bag to the cell culture bag 14 and stops working after reaching the transport value, thereby effectively keeping the NK cells in the optimal state.

[0037] Furthermore, such as Figure 2 , 3As shown, the delivery system includes a mononuclear cell conduit 15 disposed in the incubator 1 and used to connect the mononuclear cell bag 8 and the cell culture bag 14, a culture medium conduit 16 disposed in the incubator 1 and used to connect the culture medium bag 9 and the cell culture bag 14, and two delivery pumps 17 and two sealing solenoid valves 27 respectively disposed on the mononuclear cell conduit 15 and the culture medium conduit 16. The two delivery pumps 17 and the sealing solenoid valves 27 are electrically connected to the controller 2.

[0038] The controller 2 controls the delivery pump 17 and sealing solenoid valve 27 on the mononuclear cell conduit 15. The delivery pump 17 delivers mononuclear cells from the mononuclear cell bag 8 to the cell culture bag 14 through the mononuclear cell conduit 15. The delivery pump 17 and sealing solenoid valve 27 on the culture medium conduit 16 deliver the complete culture medium from the culture medium bag to the cell culture bag 14 through the culture medium conduit 16 and stop working after the delivery value is reached. When the cell monitoring system detects that the cell density in the cell culture bag 14 is too high, the cell monitoring system transmits a signal to the controller 2. The controller 2 controls the delivery pump 17 and sealing solenoid valve 27 on the culture medium conduit 16 to deliver the complete culture medium from the culture medium bag 9 to the cell culture bag 14 through the culture medium conduit 16 and stop working after the delivery value is reached.

[0039] like Figure 2 , 3 As shown, the opening of the mononuclear cell bag 8 is connected to the mononuclear cell conduit 15 via a threaded interface, the opening of the culture medium bag 9 is connected to the culture medium conduit 16 via a threaded interface, and each opening of the cell culture bag 14 is connected to the corresponding mononuclear cell conduit 15 and culture medium conduit 16 via a threaded interface. The threaded interface facilitates the installation and sealing of the openings of the mononuclear cell bag 8, the culture medium bag 9, and each opening of the cell culture bag 14 with the corresponding mononuclear cell conduit 15 and culture medium conduit 16.

[0040] It is worth mentioning that, such as Figure 4As shown, the mononuclear cells in the mononuclear cell bag 8 are separated by centrifugation using a mononuclear cell separation device. The mononuclear cell separation device, from top to bottom, includes a PBS bag 28, a blood bag 29 for holding blood, a separation bag 30 for holding separation fluid, and a connecting tube 31. The PBS bag 28 and blood bag 29 are connected by the connecting tube 31, and the blood bag 29 and separation bag 30 are also connected by the connecting tube 31. When it is necessary to separate the mononuclear cells in the blood, the connecting tube 31 between the PBS bag 28 and blood bag 29 is first tightly sealed with clamps, separating the PBS bag 28 and blood bag 29. Then, the mononuclear cell separation device is placed on a centrifuge. The blood in the blood bag 29, through the separation fluid and centrifugation, separates from top to bottom into a plasma layer, a white membrane layer, a separation fluid layer, and a red blood cell and granulocyte layer. The white membrane layer and plasma layer remain in the blood bag 29, while the separation fluid layer and the red blood cell and granulocyte layer remain in the separation bag 30. 0. Separate the blood bag 29 and the separation bag 30 by heat sealing the connecting tube 31 between them; open the clamp, then centrifuge the PBS bag 28 and the blood bag 29 to separate the mononuclear cells into the PBS bag 28. Then heat seal the connecting tube 31 between the PBS bag 28 and the blood bag 29 and separate the PBS bag 28 and the blood bag 29 to complete the separation of the mononuclear cells. Finally, assemble the PBS bag 28 with the cell culture bag 14 for use. That is, the PBS bag 28 is the mononuclear cell bag 8.

[0041] like Figure 1 , 2 As shown, the cell monitoring system includes a cell viability analyzer 18 for detecting NK cells, a detection tube 19 mounted on an incubator 1 for delivering NK cells and culture medium to the detection port of the cell viability analyzer 18, a three-way solenoid valve 20 positioned between the detection tube 19 and the cell culture system, and a liquid pump 21 positioned on the side of the detection tube 19 away from the three-way solenoid valve 20. The cell viability analyzer 18 is a Vi-CELL XR Beckman Coulter viability analyzer. One end of the detection tube 19 is connected to the cell culture system via the three-way solenoid valve 20, and the other end is located at the detection port of the cell viability analyzer 18. The controller 2 is electrically connected to both the cell viability analyzer 18 and the three-way solenoid valve 20. The cell viability analyzer 18 is a Vi-CELL XR Beckman Coulter viability analyzer.

[0042] When it is necessary to detect NK cells in the cell culture system, the controller 2 controls the three-way solenoid valve 20 to connect the detection tube 19 to the cell culture system. Then, the liquid pump 21 extracts NK cells and culture medium from the cell culture system. After a certain amount of NK cells and culture medium are extracted, the controller 2 controls the three-way solenoid valve 20 to connect the detection tube 19 to the outside. Under the action of the liquid pump 21, the NK cells and culture medium enter the detection port of the cell viability analyzer 18 along the detection tube 19. The cell viability analyzer 18 detects the NK cells and culture medium and uploads the detection data to the controller 2.

[0043] like Figure 1 , 2 As shown, the incubator 1 is equipped with a gas regulation system connected to the culture chamber 10. The gas regulation system includes a gas sensor 22 installed inside the culture chamber 10, a carbon dioxide pipe 23 installed on the incubator 1 for supplying carbon dioxide to the inside of the culture chamber 10, a carbon dioxide tank 24 installed outside the incubator 1 and connected to the outer end of the carbon dioxide pipe 23, an exhaust pipe 25 installed on the incubator 1 for discharging the gas inside the culture chamber 10, and gas pipe solenoid valves 26 installed on the carbon dioxide pipe 23 and the exhaust pipe 25 respectively. The gas sensor 22 and the gas pipe solenoid valves 26 are electrically connected to the controller 2.

[0044] When the gas sensor 22 detects that the carbon dioxide concentration in the culture chamber 10 is lower than the set value range, the controller 2 can open the carbon dioxide gas pipe solenoid valve 26, and the carbon dioxide in the carbon dioxide tank 24 enters the culture chamber 10 through the carbon dioxide pipe 23. After the carbon dioxide concentration in the culture chamber 10 is within the set value range, the controller 2 can close the gas pipe solenoid valve 26 of the carbon dioxide pipe 23.

[0045] The working process and beneficial effects of this utility model are as follows: When NK cell culture is required, the incubator 1 is first placed in a sterile environment or sterile room. The cell culture system and cell monitoring system can be set through the control box to keep the cultured NK cells and the complete culture medium at a relatively constant temperature. Then, mononuclear cells and complete culture medium are placed into the cell culture system for culture. After the cell monitoring system detects the growth status, concentration and activity of NK cells in the cell culture system, the detected values ​​are fed back to the controller 2. The controller 2 controls the cell culture system to add complete culture medium to the NK cells, thereby keeping the NK cells in the cell culture system in the optimal state.

[0046] In summary, the intelligent control culture formed by the controller 2, cell culture system, and cell monitoring system on the incubator 1 can effectively avoid contamination caused by manual addition of complete culture medium, and achieve the goals of greatly improving culture efficiency, providing a stable culture environment, and intelligent management.

[0047] The above description is only a preferred embodiment of the present utility model and is not intended to limit the present utility model. Any modifications, equivalent substitutions, improvements, etc., made within the design concept of the present utility model should be included within the protection scope of the present utility model.

Claims

1. An in vitro NK cell culture device, characterized in that: Including an incubator (1) with a controller (2), The cell culture system is set in the incubator (1) for culturing NK cells and quantitatively adding complete culture medium to the incubator (1). It has a constant temperature setting function for culturing NK cells and complete culture medium to maintain the proliferation requirements of NK cells and is electrically connected to the controller (2). The cell monitoring system is set on the incubator (1) and connected to the cell culture system. It is used to periodically sample and detect the growth status, concentration and activity of NK cells. It is electrically connected to the controller (2).

2. The in vitro NK cell culture device according to claim 1, characterized in that: The cell culture system includes a material chamber (4) disposed on an incubator (1) and isolated from the outside world, a first temperature sensor (5) disposed in the material chamber (4), a first heater (6) disposed in the material chamber (4), a first cooler (7) disposed in the material chamber (4), a culture medium bag (9) disposed in the material chamber (4) and containing complete culture medium, a mononuclear cell bag (8) disposed in the material chamber (4) and containing mononuclear cells, a culture chamber (10) disposed on an incubator (1) and isolated from the outside world, a second temperature sensor (11) disposed in the culture chamber (10), and a first temperature sensor (5) disposed in the culture chamber (10). The system includes a second heater (12), a second cooler (13) located in the culture chamber (10), a cell culture bag (14) located in the culture chamber (10) for culturing NK cells, and a conveying system located on the incubator (1) for conveying materials from the material chamber (4) to the culture chamber (10). The cell culture bag (14) in the culture chamber (10) is connected to the cell monitoring system. The first temperature sensor (5), the first heater (6), the first cooler (7), the second temperature sensor (11), the second heater (12), the second cooler (13), and the conveying system are all electrically connected to the controller (2).

3. The in vitro NK cell culture device according to claim 2, characterized in that: The delivery system includes a mononuclear cell conduit (15) disposed in the incubator (1) and used to connect the mononuclear cell bag (8) and the cell culture bag (14), a culture medium conduit (16) disposed in the incubator (1) and used to connect the culture medium bag (9) and the cell culture bag (14), and two delivery pumps (17) and two sealing solenoid valves (27) respectively disposed on the mononuclear cell conduit (15) and the culture medium conduit (16). The two delivery pumps (17) and the sealing solenoid valves (27) are electrically connected to the controller (2).

4. The in vitro NK cell culture device according to claim 3, characterized in that: The opening of the mononuclear cell bag (8) is connected to the mononuclear cell conduit (15) via a threaded interface. The opening of the culture medium bag (9) is connected to the culture medium conduit (16) via a threaded interface. Each opening of the cell culture bag (14) is connected to the corresponding mononuclear cell conduit (15) and culture medium conduit (16) via a threaded interface. The mononuclear cells in the mononuclear cell bag (8) are separated by centrifugation using a mononuclear cell separation device. The mononuclear cell separation device includes, from top to bottom, a PBS bag (28), a blood bag (29) for holding blood, a separation bag (30) for holding separation fluid, and a connecting tube (31). The PBS bag (28) and the blood bag (29) are connected by the connecting tube (31). The blood bag (29) and the separation bag (30) are connected by the connecting tube (31).

5. The in vitro NK cell culture device according to claim 2, characterized in that: The cell culture bag (14), the mononuclear cell bag (8), and the culture medium bag (9) are all made of EVA material.

6. The in vitro NK cell culture device according to claim 1, characterized in that: The cell monitoring system includes a cell viability analyzer (18) for detecting NK cells, a detection tube (19) set on an incubator (1) for delivering NK cells and culture medium to the detection port of the cell viability analyzer (18), a three-way solenoid valve (20) set between the detection tube (19) and the cell culture system, and a liquid pump (21) set on the side of the detection tube (19) away from the three-way solenoid valve (20). One end of the detection tube (19) is connected to the cell culture system through the three-way solenoid valve (20), and the other end is located at the detection port of the cell viability analyzer (18). The controller (2) is electrically connected to the cell viability analyzer (18) and the three-way solenoid valve (20).

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