Flow path system for cell culture device and cell culture device

Abstract

The present application discloses a flow path system for a cell culture device and a cell culture device, the flow path system for a cell culture device comprises: a cell culture chip, the cell culture chip having a culture chamber; and a liquid supply mechanism, the liquid supply mechanism being used to supply culture medium stock solution to the culture chamber, the liquid supply mechanism indirectly supplies culture medium stock solution to the culture chamber and the culture medium stock solution in the culture chamber is circulatory. The flow path system for a cell culture device proposed in the present application can provide cells or tissues with fluid shear forces close to physiological conditions more realistically; ensure that nutrients are uniformly and continuously delivered to cells, avoid local nutrient depletion or accumulation of metabolites, thereby facilitating long-term stable growth and maintenance of activity of cells, and facilitating normal functional expression of cells; it can also provide a more stable and lasting environment, provide more time point data for long-term experiments, and at the same time, effectively avoid contamination of cells or tissues.

Description

Technical Field

[0001] The present application relates to the field of bioengineering, and in particular to a flow path system for a cell culture device and a cell culture device. Background Art

[0002] As an in vitro research method, cell culture technology provides a unique platform for biomedical research. Researchers can control and study life processes in an in vitro environment. Cells cultured in vitro can be used in disease model research, drug screening and other research. In the past, cell culture was mostly carried out on a two-dimensional plane, which could not completely replicate the three-dimensional structure and complex environment of cells in the body, and it was difficult to reproduce the physiological functions of complete organs.

[0003] In recent years, with the rise of microfluidic organ chip technology, researchers have made major breakthroughs in the field of cell culture. Microfluidic organ chips simulate the complex three-dimensional structure and physiological functions of the microenvironment in the body, providing cells with a growth environment that is closer to the natural state. By simulating blood flow through a microfluidic system, the delivery of nutrients and the excretion of metabolites, as well as signal exchange between cells, can be achieved, thereby more realistically reflecting the physiological and pathological states of organs or tissues. In the body, the flow of blood between organs or tissues is a closed system, providing fluid shear force to tissues or organs. Although microfluidic organ chips can currently provide fluid shear force to tissues, they have not yet achieved a closed circulation flow state of blood similar to that in the body, in order to more realistically reflect the growth state of tissues or organs in the body.

[0004] Therefore, it is necessary to provide a fluid system that can realize closed circulation flow of culture medium inside a cell culture device. Summary of the invention

[0005] According to the first aspect of the present application, a flow path system for a cell culture device is proposed, which includes: a cell culture chip, which has a culture chamber; and a liquid supply mechanism, which is used to supply culture medium stock solution to the culture chamber, and the liquid supply mechanism indirectly supplies culture medium stock solution to the culture chamber and the culture medium stock solution in the culture chamber is circulatory.

[0006] Specifically, the liquid supply mechanism includes: a circulation container for circulatedly supplying culture medium stock solution to the culture chamber; and a stock solution container for providing culture medium stock solution to the circulation container.

[0007] Specifically, the flow path system further includes: a waste liquid container for collecting waste liquid in the flow path system; and a cleaning container for storing liquid for cleaning the flow path system.

[0008] Specifically, the number of the circulation containers is at least one.

[0009] Specifically, the flow path system further includes: a pump, which is used to provide power for the flow of liquid in the flow path system, and the number of the pump is at least one.

[0010] Specifically, the number of the cell culture chip is 1, and the circulation container is connected to the inlet and outlet of the cell culture chip through pipelines respectively.

[0011] Specifically, there are multiple cell culture chips, and the multiple cell culture chips are connected in series through pipelines, and the pipelines of the circulation container are respectively connected to the inlet of the first cell culture chip and the outlet of the last cell culture chip; or multiple cell culture chips are independently arranged, and there are multiple circulation containers, and the circulation containers are respectively connected to the inlet and outlet of the corresponding cell culture chips.

[0012] According to a second aspect of the present application, a cell culture device is proposed, which includes: a flow path system for the cell culture device, an imaging system and an analysis and detection system, and the flow path system is the above-mentioned flow path system for the cell culture device.

[0013] Specifically, the imaging system includes: a microscope lens, a camera, a common light source and a fluorescence excitation light source; the microscope lens, the camera and the fluorescence excitation light source are arranged below the cell culture chip, and the common light source is arranged above the cell culture chip, and the cell culture chip is the above-mentioned cell culture chip.

[0014] Specifically, the analysis and detection system includes a pipetting device for sucking liquid in the circulation container for detection; a detection system for detecting the liquid, and the circulation container is the above-mentioned circulation container.

[0015] The flow system for a cell culture device proposed in the present application can more realistically simulate the in vivo environment by setting up a flow system on a cell culture chip that allows the circulation of culture medium stock solution, thereby providing cells or tissues with shear forces close to those under physiological conditions; the circulating supply of culture medium stock solution can ensure that nutrients are evenly and continuously delivered to cells, avoiding local nutrient depletion or accumulation of metabolites, thereby facilitating long-term stable growth and maintenance of cell activity, and facilitating the normal functional expression of cells; the circulation of culture medium stock solution to the cell culture chip reduces the waste of culture medium, provides a more stable and lasting environment, provides more time point data for long-term experiments, and at the same time, can effectively avoid contamination of cells or tissues.

[0016] Other features and advantages of the present application will be described in detail in the subsequent detailed description. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] The accompanying drawings constituting a part of the present application are used to provide a further understanding of the present application, and the schematic implementation modes and descriptions of the present application are used to explain the present application. In the accompanying drawings:

[0018] Figure 1 Schematic diagram of a flow system for a cell culture device according to the present application.

[0019] Figure 2 Schematic diagram of multiple cell culture chips working independently in this application.

[0020] Figure 3 This is a schematic diagram of the connection between multiple cell culture chips and the circulation container when working independently in this application.

[0021] Figure 4 This is a schematic diagram of multiple cell culture chips working in series in this application.

[0022] Figure 5 This is a schematic diagram of the connection between multiple cell culture chips and a circulation container when working in series according to the present application.

[0023] Figure 6 Schematic diagram of the cell culture device according to the present application.

[0024] Explanation of the numbers in the accompanying drawings: 1-cell culture chip; 2-circulation container; 3-stock liquid container; 4-waste liquid container; 5-microscope lens; 6-camera; 7-ordinary light source; 8-fluorescence excitation light source; 9-pipette device; 10-disinfectant container; 11-clean water container; 12-multi-channel switching valve; 13-circulation pump; 14-waste liquid pump; 15-stock liquid pump; 16-guide rail; 17-enzyme reader; 18-enzyme plate; 19-automatic plate ejector; 20-robotic arm; 21-disinfection part; 22-control board; 23-host; 24-display screen; 25-inlet; 26-telescopic tube; 27-circulation container cover; 28-chip culture platform; 29-cell culture chip station. DETAILED DESCRIPTION

[0025] The technical solution of the present application will be described in detail below with reference to the accompanying drawings and in combination with the implementation modes.

[0026] The establishment of organoids provides a good model for in vitro research on diseases and interactions between multiple organs. However, organs or tissues are surrounded by blood vessels in the body, and blood circulation provides fluid shear force for cells, provides nutrients for tissues, and removes metabolic waste. In current research, microfluidic chips are used to provide fluid shear force for organoids, but a closed blood circulation similar to that in the body has not yet been achieved to more realistically reflect the true state of tissues or organs in the body.

[0027] Based on this, according to the first aspect of the present application, a flow path system for a cell culture device is proposed, which may include: a cell culture chip 1, which has a culture chamber; and a liquid supply mechanism, which is used to supply culture medium stock solution to the culture chamber, and the liquid supply mechanism indirectly supplies culture medium stock solution to the culture chamber and the culture medium stock solution in the culture chamber is circulatory.

[0028] The flow system for the cell culture device in the present application provides a dynamic culture environment for cells, which is helpful for the maintenance and research of cell functions. The dynamic circulation flow provides cells with sufficient nutrients and oxygen, removes metabolic waste, and maintains a stable pH value in the cell culture environment. At the same time, the circulation flow of the culture medium also reduces the consumption and waste of the culture medium.

[0029] In order to provide culture medium to the cell culture chip 1, specifically, the liquid supply mechanism may include: a circulation container 2 for circulating and supplying culture medium stock solution to the culture chamber; a stock solution container 3: for providing culture medium stock solution to the circulation container 2. The circulation container 2 has at least four openings, which are used to communicate with the culture chamber of the cell culture chip 1 to form a closed-loop cavity, discharge the waste liquid in the circulation container 2, and communicate with the stock solution container 3. The circulation container 2 is not directly connected to the outside world; the stock solution container 3 can be in a suitable form that can provide culture medium stock solution to the circulation container 2, for example, it can be a centrifuge tube, and the required volume of culture medium is provided according to actual needs. When working, it is only necessary to place the stock solution container 3 on the injection port 25, and the injection port 25 can be provided with a suitable pipeline to transfer the culture medium stock solution to the circulation container 2, for example, it can be a common hose and a hard pipe, preferably a telescopic tube 26. The telescopic tube 26 can be used to transfer the culture medium stock solution when needed, and the degree of change can be changed according to the height of the stock solution container 3 to transfer the culture medium stock solution in the stock solution container 3 to avoid insufficient culture medium stock solution in the circulation container 2. After the raw liquid container 3 provides the culture medium raw liquid to the circulation container 2, the circulation container 2 and the culture chamber form a closed loop. The closed circulation flow of the circulation container 2 provides culture medium for the cell culture chip 1, which can avoid the communication between the raw liquid container 3 and the internal environment of the culture chamber, reducing the possibility of cell contamination. At the same time, the circulating flow of the culture medium between the circulation container 2 and the culture chamber simulates the blood flow state in the body, and more realistically reflects the living environment of cells or tissues in the body.

[0030] In order to ensure the sterile environment of the flow system for the next organ chip culture, specifically, the flow system can also include: a waste liquid container 4 for collecting waste liquid in the flow system; a cleaning container for storing liquid for cleaning the flow system. The above-mentioned waste liquid container 4 is used to empty all liquids in the circulation container 2, pipelines, and culture chambers after the organ chip culture is completed to avoid affecting the next organ chip culture; in order to avoid microbial contamination caused by liquid residue in the flow system, the flow system needs to be cleaned after the flow system empties the culture medium. In order to ensure the cleaning effect, the above-mentioned cleaning container includes a disinfectant container 10 and a clean water container 11; the disinfectant container 10 can prevent the flow system from being contaminated, and the clean water container 11 is used to provide clean water (for example: deionized water) to clean the disinfectant of the flow system to avoid residues causing adverse effects on the next organ chip culture.

[0031] In order to form a closed flow path with the above-mentioned culture chamber, specifically, the number of the circulation container 2 can be at least 1, and the outlet and inlet of the circulation container 2 correspond to the inlet and outlet of the cell culture chip 1 respectively. The number of the above-mentioned circulation containers 2 can be set according to the number of outlets and inlets on the cell culture chip 1. For example, if the cell culture chip 1 has 2 inlets and 2 outlets, 2 circulation containers 2 need to be set to correspond thereto, so as to provide different liquids for the cell culture chip 1 to meet the experimental and cell growth requirements. In this application, a cell culture chip 1 with 2 inlets and 2 outlets is taken as an example.

[0032] The pipeline connecting the circulation container 2 with the raw liquid container 3, the disinfectant container 10 and the clean water container 11 is switched to be connected with the raw liquid container 3 or the disinfectant container 10 and the clean water container 11 through a multi-channel switching valve 12. The number of the multi-channel switching valves 12 is determined according to the number of raw liquid containers 3 and the cell culture chip 1.

[0033] In order to promote the flow of liquid in the flow path system, specifically, the flow path system may further include: a pump, which is used to provide power for the flow of liquid in the flow path system, and the number of the pump is at least one. The number of the pumps is set according to the positions between the stock liquid container 3, the circulation container 2, and the cleaning container; the number of the pumps can be 1 and / or 2, and the pump includes a circulation pump 13. The circulation pump 13 needs to be set between the circulation container 2 and the cell culture chip 1 to provide power for the flow of liquid in the circulation container 2. In addition, the rate at which the circulation pump 13 pumps out the liquid can also be adjusted according to experimental needs. The pumping rate of the circulation pump 13 is 2.5μL / min-10μL / min; the circulation container 2 in the flow path system discharges the waste liquid to the waste liquid container 4 by gravity, and the stock liquid container 3 and the cleaning container use gravity to provide the culture medium stock liquid and the cleaning liquid to the circulation container 2; the number of the pumps can be 3, 4, 5 or 6, and the pump includes a circulation pump 13, a waste liquid pump 14 and a stock liquid pump 15. The waste liquid pump 14 is used to pump the waste liquid in the flow path system out to the waste liquid container 4, and the stock liquid pump 15 pumps the culture medium stock liquid or the cleaning solution into the circulation container 2. The number of the pumps is determined according to the number and positional relationship of the cell culture chips 1 and the positional relationship among the original liquid container 3 , the waste liquid container 4 , the cleaning container and the circulation container 2 .

[0034] The above is combined with this application Figure 1 A detailed description of the fluidic system used in the cell culture apparatus is shown.

[0035] As in this application Figure 1-Figure 5 As shown, in order to carry out organ chip culture, specifically, the number of the cell culture chip 1 can be one, and the circulation container 2 is connected to the inlet and outlet of the cell culture chip 1 through pipelines. The number of the cell culture chips 1 can be multiple, and multiple cell culture chips 1 are connected in series through pipelines, and the pipelines of the circulation container 2 are respectively connected to the inlet of the first cell culture chip 1 and the outlet of the last cell culture chip 1; or multiple cell culture chips 1 are independently arranged, and the number of the circulation containers 2 is multiple, and the circulation containers 2 are respectively connected to the inlet and outlet of the corresponding cell culture chips 1. In this application, four cell culture chips 1 are taken as an example.

[0036] As in this application Figure 6As shown, according to the second aspect of the present application, a cell culture device is proposed, and the cell culture device may include: a flow path system for the cell culture device, an imaging system and an analysis and detection system, and the flow path system is the above-mentioned flow path system for the cell culture device. Organ chip culture usually takes a long time, such as 7 days or 14 days. The specific culture time is determined according to the cell type and growth state. During the organ chip culture, it is necessary to observe the growth state of the cells in the cell culture chip 1. An imaging system is set inside the cell culture device, and the cells can be observed in the original position of the organ chip without suspending the closed circulation flow of the culture medium stock solution; in addition. The setting of the detection system can timely evaluate the parameters of the culture medium stock solution, such as evaluating cell metabolism, cytotoxicity, detecting specific biomarkers, changes in nutrients, etc. Combining the flow path system with the imaging system and the analysis and detection system can realize the highly integrated culture of the cell culture chip 1, reduce human errors, and improve experimental efficiency.

[0037] In order to better record the state of the organ chip, specifically, the imaging system may include: a microscope lens 5, a camera 6, an ordinary light source 7 and a fluorescent excitation light source 8; the microscope lens 5, the camera 6 and the fluorescent excitation light source 8 may be arranged below the cell culture chip 1, and the ordinary light source 7 may be arranged above the cell culture chip 1, and the cell culture chip 1 is the above-mentioned cell culture chip 1. When there are multiple organ chips, before the experiment starts, the position zero point of the imaging system is calibrated, and the imaging system can be accurately moved along the guide rail 16 to the position of the corresponding cell culture chip 1 to accurately record the growth state of each organ chip; when only the tissue or cell morphology needs to be observed, the ordinary light source 7 can be used, and when it is necessary to observe fluorescently labeled molecules, tissues or cell structures, the fluorescent excitation light source 8 is used, and the fluorescent excitation light source 8 can excite wavelengths of different lengths as needed.

[0038] In order to detect the culture medium at regular intervals, specifically, the analysis and detection system may include a pipetting device 9 for aspirating the liquid in the circulation container 2 for detection; a detection system for detecting the liquid, and the circulation container 2 is the above-mentioned circulation container 2. The above-mentioned pipetting device 9 can be an appropriate device to achieve the purpose of sampling in the circulation container 2, for example, it can be a pipette gun; in order to ensure that the culture medium in the circulation container 2 is not evaporated and to ensure sterility, a circulation container cover 27 is also provided on the top of the circulation container 2. When sampling is required, the circulation container cover 27 is opened; the circulation container cover 27 can be assisted by a mechanical arm or in the form of an automatic switch; the above-mentioned pipette gun can be moved by means of a guide rail or a mechanical wall to complete sampling. In this application, the guide rail 16 is taken as an example; the above-mentioned detection system can be an ELISA instrument 17. The ELISA instrument 17 does not require other operations. The liquid is sucked onto the ELISA plate 18, and then the ELISA plate 18 is placed in the ELISA instrument 17 to complete the operation. The ELISA plate 18 is placed in the automatic plate ejector 19. When detection is required, it is automatically ejected. After the sample is added, it is transmitted to the ELISA instrument 17 through the mechanical wall 20 for detection. The ELISA instrument 17 can simultaneously detect and record the culture medium of different cell culture chips together, saving detection time and ensuring the consistency of detection standards.

[0039] In order to ensure that the cell culture device operates better, the cell culture device also includes an environmental disinfection component 21, a control system, a heating device and a gas device. The disinfection component 21 can be an appropriate component that can meet the internal disinfection of the cell culture device, such as an ozone generator and / or an ultraviolet lamp; the control system is used to control the operation of the internal equipment of the cell culture device, such as controlling the operation and disinfection of the flow system, controlling the imaging system to reach the position of the cell culture chip 1 to be observed along the guide rail 16 and complete the photo taking work, and controlling the sampling, placement of samples to be tested, and detection of the analysis and detection system; the heating device and the gas device can also set parameters on the control system according to experimental needs; the control system includes a control board 22, a host 23 and a display screen 24, and the control board 22, the host 23 and the display screen 24 are connected to other devices through wires. When in use, it only needs to be set on the display screen 24 to complete the above-mentioned workflow inside the cell culture device.

[0040] When organ chip culture is required, cells are inoculated into the cell culture chip 1, and then the cell culture chip 1 is placed on the cell culture chip station 29, and then the pipeline of the circulation container 2 is connected to the outlet and inlet of the cell culture chip 1 respectively. The number of the above-mentioned cell culture chip stations 29 is set according to actual needs. In this application, 4 stations are taken as an example. The cell culture chip stations 29 are set on the chip culture platform 28, and then the control system is used to mobilize the corresponding system work to complete the organ chip culture process.

[0041] The flow path system for a cell culture device proposed in the present application can more realistically simulate the in vivo environment by setting a flow path system on the cell culture chip 1 that allows the culture medium stock solution to circulate, thereby providing cells or tissues with shear forces close to those under physiological conditions; the circulating supply of culture medium stock solution can ensure that nutrients are evenly and continuously delivered to cells, avoiding local nutrient depletion or accumulation of metabolites, thereby facilitating long-term stable growth and maintenance of cell activity, and facilitating the normal functional expression of cells; the culture medium stock solution is circulated and supplied to the cell culture chip 1, reducing the waste of culture medium, providing a more stable and lasting environment, providing more time point data for long-term experiments, and at the same time, effectively avoiding contamination of cells or tissues.

[0042] The preferred embodiments of the present application are described in detail above; however, the present application is not limited to the specific details in the above embodiments. Within the technical concept of the present application, a variety of simple modifications can be made to the technical solution of the present application, and these simple modifications all fall within the protection scope of the present application.

[0043] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, this application will not further describe various possible combinations.

[0044] In addition, the various implementation modes of the present application may be arbitrarily combined, and as long as they do not violate the concept of the present application, they should also be regarded as the contents disclosed by the present invention.

Claims

1. A flow path system for a cell culture device, the flow path system for a cell culture device comprising: A cell culture chip (1), wherein the cell culture chip (1) has a culture chamber; and a liquid supply mechanism, the liquid supply mechanism being used to supply the culture medium stock solution to the culture chamber, The waste liquid container (4) is used to collect waste liquid in the flow path system; the cleaning container is used to store liquid for cleaning the flow path system; the characteristics are: The liquid supply mechanism indirectly supplies the culture medium stock solution to the culture chamber, and the culture medium stock solution in the culture chamber is circulatable; Wherein, the liquid supply mechanism comprises: A circulation container (2) is used for circulating the culture medium stock solution to the culture chamber and circulating with the culture medium in the culture chamber; A stock liquid container (3) is used to provide the circulation container (2) with a culture medium stock liquid; after the stock liquid container (3) provides the culture medium stock liquid to the circulation container (2), the circulation container (2) and the culture chamber form a closed-loop cavity.

2. The flow path system for a cell culture device according to claim 1, characterized in that: The number of the circulation container (2) is at least one.

3. The flow path system for a cell culture device according to claim 1, characterized in that: The flow path system further includes: a pump, which is used to provide power for the flow of liquid in the flow path system, and the number of the pump is at least one.

4. The flow path system for a cell culture device according to claim 1, characterized in that: The number of the cell culture chip (1) is one, and the circulation container (2) is connected to the inlet and outlet of the cell culture chip (1) respectively through pipelines.

5. The flow path system for a cell culture device according to claim 1, characterized in that: There are multiple cell culture chips (1), the multiple cell culture chips (1) are connected in series via pipelines, and the pipeline of the circulation container (2) is respectively connected to the inlet of the first cell culture chip (1) and the outlet of the last cell culture chip (1); or A plurality of cell culture chips (1) are independently arranged, the number of the circulation containers (2) is plural, and the circulation containers (2) are respectively connected to the inlet and outlet of the corresponding cell culture chip (1).

6. A cell culture device, characterized in that The cell culture device comprises: a flow path system for a cell culture device, an imaging system and an analysis and detection system. The flow path system is the flow path system for a cell culture device according to any one of claims 1 to 5.

7. The cell culture device according to claim 6, characterized in that: The imaging system comprises: a microscope lens (5), a camera (6), a common light source (7) and a fluorescence excitation light source (8); the microscope lens, the camera and the fluorescence excitation light source are arranged below the cell culture chip (1), the common light source (7) is arranged above the cell culture chip (1), and the cell culture chip (1) is the cell culture chip (1) according to claim 1.

8. The cell culture device according to claim 7, characterized in that: The analysis and detection system comprises a liquid transfer device (9) for aspirating liquid in the circulation container (2) for detection; and a detection system for detecting the liquid, wherein the circulation container (2) is the circulation container (2) according to claim 2.

Images