Automatic modular cell culture device with perfusion-based culture solution conveying function

Abstract

The disclosed cell culture device comprises: a cell culture module comprising a cell culture unit in which an orifice plate, which forms at least one culture space, is attached to a cartridge; a culture solution supply module comprising: a pump unit for forming a supply and discharge flow of a culture solution to the cell culture unit; the culture solution storage chamber is used for accommodating at least one culture solution chamber; and a pair of culture solution tubes made of a flexible material and connecting the culture solution supply module and the cell culture module, which are spatially separated, to form a duct through which the culture solution is supplied and discharged, the cell culture module being placed in a CO2 incubator for creating a warm environment, the culture solution storage chamber creating a low-temperature environment lower than normal temperature, and the culture solution storage chamber creating a low-temperature environment lower than normal temperature. A pair of culture solution tubes exposed between the culture solution supply module and the cell culture module is in a normal-temperature environment.

Description

Technical Field

[0001] This invention relates to an automated cell culture apparatus, specifically a modular cell culture apparatus in which a cell culture module and a culture medium supply module are connected via a pipeline network, the culture medium is automatically supplied and discharged, and the apparatus can be miniaturized to the point where only the cell culture module is placed in a commercial CO2 incubator. Through perfusion-based culture medium delivery, extremely small amounts of culture medium can be precisely delivered, and the medium can be supplied without changing its texture during delivery.

[0002] This application claims priority to Korean Patent Application No. 10-2022-0042443, dated April 5, 2024, the entire contents of which are disclosed in the Korean Patent Application No. 10-2022-0042443 and are incorporated herein by reference. Background Technology

[0003] Cell culture is an indispensable technique in many fields for researching and applying cell function, properties, diseases, and treatments. However, cell culture typically requires significant time and effort, especially given the numerous risk variables such as contamination and mutations during the process, necessitating a highly precise culture environment. Furthermore, the quality and reproducibility of cell culture are highly dependent on the operator's skill and technique, thus raising concerns about ensuring the reliability and consistency of experimental results. In light of this, the demand for automated cell culture equipment has been increasing in recent years, with many companies investing considerable human and material resources in developing technologies to automate the cell culture process.

[0004] This invention addresses the aforementioned problems, and its main feature is the automation of the environment, enabling automatic replacement of the culture medium required for cell growth within the culture space. Furthermore, in addition to the aforementioned technical concepts, this invention also aims to provide additional technical elements that are not obvious to those skilled in the art. Summary of the Invention

[0005] The problem that the invention aims to solve For cell culture processes with a long time span, the cell culture module needs to be placed in a warm environment suitable for cell growth (usually around 37°C), while the culture medium supply module needs to be placed in a low-temperature environment (usually 4~8°C) to prevent the stored culture medium from deteriorating.

[0006] Thus, when considering means to maintain two spaces at different temperatures, heat transfer between the two spaces may lead to derivative problems such as energy waste and water condensation. Therefore, the main objective of this invention is to physically separate the warm space for cell culture from the low-temperature space for culture medium storage, and to achieve perfusion-based culture medium delivery using slender culture medium tubes and a gas-liquid interface delivery mechanism, thereby enabling accurate quantities of culture medium to move between the cell culture module and the culture medium supply module without waste or degradation.

[0007] Furthermore, the most critical issue in cell culture is cross-contamination caused by external contaminants. Therefore, cell culture equipment must prevent contamination at its source during use. Moreover, to improve productivity and process efficiency, all surfaces that may come into contact with the culture medium and cells should be designed to be detachable from the equipment, allowing for the use of pre-cleaned and sterilized units during culture operations. This invention addresses the aforementioned problems, and another objective is to eliminate the need for cleaning and sterilization steps during the culture process. For reference, examples of surfaces that may come into contact with the culture medium and cells include the walls of the cell culture space, the walls of the culture medium storage space, the walls of the piping for culture medium movement, and pumps and valves that generate flow.

[0008] In addition, the present invention aims to provide an automated cell culture device in which the cell culture module and the culture medium supply module are connected by a pipeline network to automatically supply and discharge culture medium, and can be miniaturized to the point that only the cell culture module is placed in a commercial CO2 incubator.

[0009] In addition, the purpose of this invention is to solve the problems of culture medium waste and deterioration by realizing a perfusion-based culture medium delivery system that utilizes a slender culture medium tube and a gas-liquid interface delivery mechanism, thereby ensuring that an accurate amount of culture medium is supplied without waste or deterioration.

[0010] Another objective of the present invention is to provide an automated cell culture apparatus that, by implementing a practical disposable pump unit, can block the risk of contamination at the source, eliminate the need for cleaning and sterilization processes, thereby improving productivity and process efficiency, and simplifying the validation process.

[0011] The technical problems of this invention are not limited to those mentioned above. Those skilled in the art can clearly understand other technical problems not mentioned through the following description.

[0012] means for solving problems In one embodiment, the cell culture apparatus provided by the present invention includes: a cell culture module comprising a cell culture section formed by mounting a well plate forming at least one culture space on a cartridge; a culture medium supply module comprising: a pump unit for supplying and draining culture medium to the cell culture section; a culture medium storage chamber for accommodating at least one culture medium chamber; and a pair of flexible culture medium tubes connecting the spatially separated culture medium supply module and the cell culture module to form a pipeline for supplying and draining the culture medium, wherein the cell culture module is placed in a CO2 incubator that creates a warm environment, the culture medium storage chamber creates a low-temperature environment below room temperature, and the pair of culture medium tubes exposed between the culture medium supply module and the cell culture module are in a room temperature environment.

[0013] The cell culture module may include: a valve module for controlling the supply and discharge of culture medium to each culture space of the well plate; and a rotating cover plate configured such that multiple culture medium channels, formed by a pair of culture medium tubes extending through the valve module, correspond to each culture space. The rotating cover plate and the culture medium channels may be positioned at the upper end of the well plate, allowing culture medium flowing in from the culture medium channels to fall from the air towards the bottom. As a specific embodiment, the multiple culture medium channels may consist of one or more supply channels and one or more discharge channels. In this case, the supply channel extends from the lower surface of the cover plate to the middle height of the culture space, and the discharge channel extends from the lower surface of the cover plate to the bottom height of the culture space.

[0014] The valve module includes multiple valve units that control the opening and closing of a pair of culture medium tubes allocated to each culture space. Each valve unit includes a movable clamp with a groove through which the culture medium tubes pass. Through linear movement of the movable clamp, the groove can squeeze the culture medium tubes to achieve closure.

[0015] An optical microscope system for monitoring cell status can be further installed at the lower end of the cell culture module. The optical microscope system may include a lens and a camera for magnifying and capturing images; the lens and camera are vertically movable for focusing. The optical microscope system may include a motorized stage for moving the lens and camera.

[0016] Additionally, a light source for an optical microscope system can be configured on the upper part of the cell culture module. The light source can be a white LED, or a red or yellow LED designed to minimize the impact on the cells.

[0017] In order to observe the entire culture space, the lens, camera and light source of the microscope system can be moved horizontally. As another embodiment, a motorized stage that can move horizontally can be set at the upper end of the microscope system, so that the lens and camera can only move vertically, and the cell culture module set at the upper end of the microscope system can move horizontally.

[0018] The pair of culture medium tubes are a supply tube and an outlet tube. After the pump unit injects a predetermined amount of culture medium into the supply tube, it pressurizes and supplies gas in sequence, thereby moving the culture medium to the culture space. The inner diameter of the supply tube should be large enough to prevent the culture medium from moving backward or forward due to gravity.

[0019] The pump unit can draw culture medium from the culture space and return it to the recovery chamber contained in the culture medium storage chamber by applying negative pressure to one end of the discharge pipe.

[0020] Furthermore, the cell culture apparatus provided by the present invention may include multiple cell culture modules connected in parallel to a culture medium supply module. By closing all valve modules respectively provided in the multiple cell culture modules and opening only the culture medium channel corresponding to the target culture space, a one-to-one correspondence between the culture medium supply module and the cell culture module as the delivery target is achieved during the generation of culture medium supply and / or recovery flow.

[0021] Furthermore, the present invention provides a culture medium supply module capable of supplying culture medium to at least one spatially separated culture space and recovering culture medium from the culture space. The culture medium supply module includes: a pump unit for forming a supply and discharge flow of culture medium for the cell culture section; and a culture medium storage chamber for accommodating a recovery chamber and at least one culture medium chamber. The culture medium storage chamber creates a low-temperature environment below room temperature. The pump unit can supply culture medium stored in the culture medium chamber to each culture space through a supply pipe and discharge it to the recovery chamber through a discharge pipe.

[0022] In one embodiment, the pump unit may include a plurality of syringe pumps, which may include a gas syringe, at least one culture medium syringe, and a recovery syringe.

[0023] The gas injector and at least one culture medium injector are connected to the middle of the tube via a T-shaped or Y-shaped branch connector. Check valves are provided upstream and downstream of the branch connector connected to the injector, thereby restricting the flow direction generated by the injector's actuation to a single direction. In the injector used for culture medium supply, the upstream end is connected to the culture medium chamber, and the downstream end is connected to the culture medium supply tube, thus allowing only unidirectional flow of culture medium from the culture medium chamber to the culture space. Components including the culture medium injector and the check valve can be connected in parallel to the supply tube.

[0024] The gas injector is connected in parallel with the culture medium injector and is positioned at the upstream end of the supply tube. The gas injector and the culture medium injector are similarly connected to the supply tube via a branch connector, and check valves are connected upstream and downstream of the branch connector, thereby restricting the flow of gas generated by the gas injector to a single direction toward the culture space.

[0025] The gas injector may have an air filter upstream of the upstream check valve.

[0026] According to the inhalation and exhalation stroke of the culture medium syringe, a predetermined amount of culture medium is introduced into the supply tube, and according to the inhalation and exhalation stroke of the gas syringe, gas is introduced into the supply tube, and the culture medium introduced into the supply tube can be transported to the culture space by the gas.

[0027] In some embodiments, the plurality of syringe pumps may include gas syringes and a plurality of culture medium syringes, with the culture medium delivery between any culture medium syringe and the gas syringe occurring at staggered times, thereby allowing the delivery of multiple culture media to the culture space in a gas-isolated state.

[0028] The recovery syringe is connected to the downstream end of the discharge tube. Check valves are provided upstream and downstream of the junction of the recovery syringe and the discharge tube, respectively, so that flow is allowed only in a single direction toward the recovery chamber.

[0029] The pump unit may have multiple pump actuators, which cause the plunger of each syringe pump to move linearly to perform suction and discharge strokes. Since the direction of flow is restricted by a check valve linked to the syringe, a volume of liquid exceeding the syringe volume can be delivered by repeatedly moving the plunger of the syringe pump.

[0030] Furthermore, the present invention provides a culture medium supply module capable of supplying culture medium to at least one spatially separated culture space. The culture medium supply module includes: a pump unit having a plurality of syringe pumps for forming a supply flow of culture medium to the cell culture section; and a culture medium storage chamber for accommodating at least one culture medium chamber. The plurality of syringe pumps include: a gas syringe connected to the upstream end of the supply tube; and at least one culture medium syringe connected in parallel downstream of the gas syringe. In the supply tube, check valves are respectively provided upstream and downstream of the confluence point of the gas syringe and each culture medium syringe, thereby allowing flow only in a single direction toward the culture space. A predetermined amount of culture medium is introduced into the supply tube through the suction and discharge strokes of the culture medium syringes, and the gas introduced into the supply tube through the suction and discharge strokes of the gas syringes causes the culture medium introduced into the supply tube to be transported to the culture space.

[0031] The plurality of syringe pumps may include gas syringes and a plurality of culture medium syringes, wherein the culture medium delivery between any culture medium syringe and the gas syringe is staggered at different times, thereby allowing the various culture media to be delivered to the culture space in a gas-isolated state.

[0032] Furthermore, the culture medium supply module may include a cooling culture medium storage chamber to prevent culture medium deterioration, and may also cool all tubing components, including the syringe and check valve, thereby preventing deterioration of the culture medium already introduced into the supply tube via the syringe. As another embodiment, the culture medium supply module can be manufactured in a small form and installed inside a commercial refrigerator or refrigeration facility, thereby maintaining the entire culture medium supply module at a low temperature.

[0033] Furthermore, since the culture medium supply module utilizes a gas-liquid interface transport mechanism to deliver the culture medium, even without directly measuring the flow rate within the tube, the presence of liquid inside the tube can be determined by using a transparent or semi-transparent tube, thus confirming whether the equipment is operating normally. As an example of equipment malfunction detection, infrared LEDs can be used to detect whether a tube is installed and whether gas or liquid is present inside it. When a command to supply or discharge culture medium is input, the state inside the supply and discharge tubes can be monitored to determine if the state changes from gas to liquid to gas, thereby confirming that the culture medium is indeed being properly delivered to the supply and discharge tubes.

[0034] Invention Effects According to the cell culture apparatus of the present invention having the above-described configuration, the cell culture module and the culture medium supply module are connected through a pipeline network to automatically supply and discharge culture medium, and the cell culture module can be miniaturized to the point that it can be placed in a commercial CO2 incubator. Therefore, the automated cell culture apparatus has excellent ease of operation.

[0035] Furthermore, this invention solves the problems of culture medium waste and deterioration by implementing a perfusion-based culture medium delivery mechanism using slender culture medium tubes and a gas-liquid interface, thereby providing an accurate quantity of culture medium without waste or deterioration.

[0036] Moreover, the present invention can provide an automated cell culture device that, by implementing a practical disposable pump unit, can block the risk of contamination at the source and eliminate the need for cleaning and sterilization processes, thereby improving productivity and process efficiency and simplifying the validation process.

[0037] The effects of the present invention are not limited to those mentioned above, and those skilled in the art can clearly understand other effects not mentioned from the following description. Attached Figure Description

[0038] Figure 1 This is a diagram showing the overall structure of a modular cell culture apparatus according to one embodiment of the present invention.

[0039] Figure 2 This is a diagram representing the cell culture module.

[0040] Figure 3 This is a diagram showing the structure of the cartridge.

[0041] Figure 4 It is a diagram showing the structure and working principle of a valve unit.

[0042] Figure 5 This is an enlarged view showing the perforated plate and cover plate installed inside the cartridge.

[0043] Figure 6 It is a cross-sectional view representing a single culture space.

[0044] Figure 7 This is a diagram representing the culture medium supply module.

[0045] Figure 8 This diagram illustrates the culture medium transport mechanism utilizing the gas-liquid interface.

[0046] Figure 9 This is a diagram used to illustrate the concept of a culture medium delivery mechanism based on multiple syringe pumps.

[0047] Figure 10This is a diagram illustrating a piping network concept based on a syringe pump that quantitatively delivers and recovers various liquids into the culture space.

[0048] Figure 11 This diagram illustrates the pre-filling process performed in a culture medium syringe.

[0049] Figure 12 This is a diagram illustrating one implementation of a pump unit.

[0050] Figure 13 This is a diagram illustrating one implementation of an automated pump unit.

[0051] Figure 14 This is a diagram showing the internal structure of the culture medium supply module.

[0052] Figure 15 This is a diagram illustrating the temperature environment of the culture medium in the modular cell culture apparatus according to the present invention.

[0053] Figure 16 This is a diagram showing the structure of connecting multiple cell culture modules to a single culture medium supply module.

[0054] Explanation of reference numerals in the attached figures 10: Cell culture apparatus; 100: Cell culture module; 110: Cell culture section; 120: Cartridge; 122: Supply pipe connection; 124: Discharge pipe connection; 130: Valve module; 132: Valve unit; 132-1: Supply valve unit; 132-2: Discharge valve unit; 134: Movable clamp; 136: Pipe groove; 137: Spring; 138: Clamp actuator; 140: Well plate; 142: Culture space; 144: Insert; 146: Membrane; 150: Cover plate; 152: Culture medium channel; 154: Supply channel; 156: Discharge channel; 160: Optical monitoring unit; 162: Optical microscope system; 200: Culture medium supply module; 210: Culture medium storage chamber; 212: Cooling unit; 220: Culture medium chamber; 222: Recovery chamber; 230: Pump unit; 232: Syringe pump; 234: Check valve; 236: Branch connector; 238: Air filter; 240: plunger; 242: pump actuator; 250: Gas injector; 252: Culture medium injector; 254: Retrieved syringe; 260: Culture medium tube; 260-M: Supervisor; 260-B: Branch Supervisor; 262: Supply pipe; 264: Discharge pipe; 300: Incubator. Detailed Implementation

[0055] The preferred embodiments of the present invention will now be described in detail with reference to the accompanying drawings. The advantages, features, and methods of achieving the present invention will become apparent from the accompanying drawings and the specific embodiments described below. However, the present invention is not limited to the embodiments disclosed below, but can be implemented in different ways. These embodiments are provided merely to complete the disclosure of the present invention and to fully inform those skilled in the art of its scope. The present invention is defined only by the scope of the claims. Throughout the specification, the same reference numerals refer to the same components.

[0056] Unless otherwise defined, all terms used in this specification (including technical and scientific terms) are to be understood in a manner commonly understood by one of ordinary skill in the art to which this invention pertains. Furthermore, terms already defined in a dictionary should not be interpreted in an idealized or excessive sense unless specifically defined therein. The terminology used in this specification is for illustrative purposes and does not limit the invention. Unless otherwise specified, the singular form in this specification includes the plural form.

[0057] The use of the terms "comprises" and / or "comprising" in this specification does not exclude the possibility that there may be one or more other components, steps, operations, and / or elements in addition to those mentioned.

[0058] It should also be noted that this invention was completed with the support of the following research projects.

[0059] Project ID: 1415187257 (20023762) Department Name: Trade, Industry and Resources Department Name of the professional organization: Korea Institute for Industrial Technology Evaluation Research Project Title: Technology Development for the Machinery and Equipment Industry - Manufacturing Basic Production Systems Research Project Title: Development of an Automated Culture System for In-situ Uniform Cell Aggregation, Proliferation, and Differentiation Based on a Material-Permeable Thin-Film Scaffold with a Three-Dimensional Concave-Concave Structure and Permeable Ultramicroperfusion Technology Contribution rate: 100 Project implementing organization: Celloid Co., Ltd. Research period: April 1, 2023 to December 31, 2026 Figure 1 This diagram illustrates the overall structure of a modular cell culture apparatus 10 (hereinafter referred to as "cell culture apparatus") according to an embodiment of the present invention. The cell culture apparatus 10 according to the present invention includes a cell culture module 100 and a culture medium supply module 200. The cell culture module 100 and the culture medium supply module 200 are connected by a pair of tubes to achieve the supply and discharge of culture medium to the cell culture module 100. The culture medium is supplied through one tube and recovered through the other tube; this network of tubes enables perfusion-based culture medium delivery.

[0060] The piping network configured to achieve perfusion-based culture medium delivery is broadly divided into supply pipes 262 that supply culture medium to the culture space 142 and discharge pipes 264 that recover culture medium from the culture space 142. That is, the culture medium pipe 260 consists of a pair of supply pipes 262 and discharge pipes 264. The piping network enables unidirectional culture medium flow as follows: culture medium stored in the culture medium chamber 220 is supplied to the culture space 142 and discharged from the culture space 142, returning to the recovery chamber 222. Based on the modules of the cell culture apparatus 10, culture medium is supplied from the culture medium supply module 200 to the cell culture module 100, and the culture medium supplied to the cell culture module 100 is recovered back to the culture medium supply module 200. The culture medium pipe 260 itself may not be composed of a single pipe; for example, it may branch midway or extend via connectors, but it is clearly divided into supply pipes 262 and discharge pipes 264 in the overall flow direction. To facilitate a clear understanding of the present invention, without needing to describe the specific components separately, the culture medium tube 260 is divided into a supply tube 262 and an outlet tube 264, and is referred to by these terms collectively.

[0061] Figure 2 This diagram illustrates a cell culture module 100. The cell culture module 100 includes a cell culture section 110 and an optical monitoring section 160 disposed at the lower part of the cell culture section 110. The cell culture section 110 includes well plates 140 made of a light-transmitting material forming a culture space 142, and the optical monitoring section 160 includes an optical microscope system 162 for acquiring images of cells cultured in the well plates 140. By utilizing the images acquired through the optical microscope system 162, remote observation of cell culture and management of the culture environment can be easily performed.

[0062] exist Figure 2In this embodiment, the well plate 140 is mounted on a cartridge 120 provided in the cell culture section 110. To facilitate operations such as replacing the well plate 140 and setting the culture medium tube 260, the cartridge 120 can be configured as a pull-out drawer-type structure. The cartridge 120 is equipped with a replaceable well plate 140 and includes a valve module 130 for controlling the inflow and outflow of culture medium, and a cover plate 150 for sealing the well plate 140.

[0063] Figure 3 This is a diagram showing the structure of the cartridge 120. (Refer to...) Figure 3 A valve module 130 is fixedly installed in the cartridge 120. The valve module 130 functions to control the flow direction of culture medium supplied to the culture spaces 142 of the well plate 140 and to control the flow direction of culture medium discharged from the culture spaces 142. When the well plate 140 has multiple culture spaces 142, the valve module 130 has a corresponding number of valve units 132. In the illustrated embodiment, the well plate 140 has six culture spaces 142, and correspondingly, the valve module 130 has seven supply valve units 132-1 and seven discharge valve units 132-2. As will be described in detail later, the cell culture apparatus 10 of the present invention employs a culture medium delivery mechanism utilizing an air-liquid interface. Thus, the seven valve units 132 consist of six valve units 132 controlling the flow direction of the culture medium and one valve unit 132 controlling the flow direction of the gas.

[0064] The front surface of the cartridge 120 has a supply tube connection 122 and a discharge tube connection 124. The supply tube connection 122 and the discharge tube connection 124 form a pipeline network connection between the cell culture module 100 and the culture medium supply module 200. A valve module 130 is disposed between the supply tube connection 122 and the discharge tube connection 124 and the well plate 140. Each culture space 142 is equipped with a valve unit 132, thereby allowing each culture space 142 to independently supply and discharge the culture medium.

[0065] Figure 4 This is a diagram illustrating the structure and working principle of valve unit 132. (For example...) Figure 4 As shown in (a), the valve unit 132 includes a movable clamp 134 capable of linearly moving vertically elastically based on a spring 137. The distal end of the movable clamp 134 has a tube groove 136 with a through-hole shape, through which the culture medium tube 260 extends to the orifice plate 140 side. (Refer to...) Figure 4In (b), under neutral conditions where no external force acts on the movable clamp 134, the movable clamp 134 is pulled upward and pressed against the culture medium tube 260 by the elastic force of the spring 137. That is, the culture medium tube 260 is blocked, thereby preventing the flow of culture medium.

[0066] Above the valve module 130, each valve unit 132 is equipped with a corresponding clamp actuator 138. In the illustrated embodiment, the valve module 130 contains fourteen (2×7) valve units 132, and correspondingly, fourteen clamp actuators 138. The clamp actuators 138 are fixedly mounted on the main body of the cell culture module 100. Each clamp actuator 138 performs linear movement in the up-down direction; when moving downwards, it compresses the spring 137 and presses the movable clamp 134. Figure 4 As shown in (b), by the downward movement of the movable clamp 134, the culture medium tube 260, which was pressed in the tube groove 136, returns to a free state. The culture medium tube 260 restores its original shape through its own elasticity, thereby forming an open state that allows the culture medium to flow.

[0067] The cartridge 120 is withdrawn from the cell culture module 100 body in a drawer-like manner. If a clamp actuator 138 is in the lowered position while the cartridge 120 is withdrawn, interference will occur when attempting to retract the cartridge 120. To prevent such malfunction of the clamp actuator 138, a recess sensor (not shown) can be provided. The clamp actuator 138 can be allowed to move downwards only when the recess sensor confirms that the cartridge 120 has been retracted, thereby preventing this problem.

[0068] Figure 5 This is an enlarged view of the perforated plate 140 and cover plate 150 mounted on the cartridge 120. In the illustrated embodiment, six independent culture spaces 142 can be arranged in a 2×3 configuration on a single perforated plate 140. This is only one example, and the number, shape, size, and configuration of the culture spaces 142 can be varied in many ways.

[0069] Furthermore, the cover plate 150 of the closed-loop structure of the sealed plate 140 has a plurality of culture medium channels 152 distributed to each culture space 142. The pair of culture medium channels 152 are divided into a supply channel 154 and a discharge channel 156. Figure 6 This is a cross-sectional view showing a single culture space 142. An insert 144 is inserted into the culture space 142, and the bottom surface of the insert 144 is formed by a membrane 146. The membrane 146 ensures the smooth flow and diffusion of substances, providing an environment in which cells can grow stably. Cells cultured on the membrane 146 cannot cross the membrane 146, but the culture medium can cross the membrane 146 to move to the drain channel 156, thus preventing cell loss at the source during culture medium replacement. Figure 6In the embodiment of culture space 142 shown, cell culture can be performed in the same way even if the cell is attached to the bottom by means of a hydrogel or the like instead of a membrane 146.

[0070] Reference Figure 6 The supply channel 154 extends downward along the inner side of the insert 144 to a middle depth, while the discharge channel 156 extends downward along the outer side of the insert 144 to a bottom depth. That is, the discharge channel 156 is longer than the supply channel 154. Because the supply channel 154 is adjacent to the inner side of the insert 144, the culture medium is supplied slowly by dropping or dripping, thus preventing splashing or excessive flow that could affect the growing cells. The discharge channel 156 extends considerably to the outside of the cell-growing insert 144 and is adjacent to the bottom of the culture space 142, thereby enabling the complete absorption and discharge of the culture medium without residue.

[0071] In addition, such as Figure 5 As shown, the culture medium tube 260 extending from the valve module 130 connects to the culture medium channel 152 provided in the cover plate 150, forming a connection structure that bends sequentially upwards, forwards, and downwards at three points. The culture medium tube 260 is a flexible tube made of polymer compounds such as polyethylene (PE), polypropylene (PP), polystyrene (PS), and silicone. Because the culture medium tube 260 is made of such a flexible material, the flow path opening and closing in the valve module 130 is performed based on the compression and reset of the tube groove 136. In the cover plate 150 of the opening and closing structure, the bending of the culture medium tube 260 at the three points causes force to act in the direction of closing the cover plate 150. Thus, the flexible culture medium tube 260 always applies force in the direction of closing the cover plate 150, thereby effectively preventing contaminants from entering the interior of the culture space 142.

[0072] Figure 7 This diagram shows the culture medium supply module 200. The culture medium supply module 200 includes a lower culture medium storage chamber 210 and a pump unit 230 disposed above the culture medium storage chamber 210. The pump unit 230 can be connected to the culture medium storage chamber 210 with an up-and-down rotating opening and closing structure. By rotating and opening the pump unit 230, it can access the culture medium storage chamber 210, thereby performing operations such as replenishing or replacing the culture medium in the culture medium chamber 220.

[0073] Pump unit 230 functions to supply culture medium stored in at least one culture medium chamber 220 contained in culture medium storage chamber 210 to cell culture module 100, and to generate flow for recovering culture medium from culture space 142 of cell culture module 100. In particular, in this invention, pump unit 230 employs a culture medium delivery mechanism utilizing an air-liquid interface. Figure 8 This diagram illustrates the culture medium transport mechanism that utilizes the gas-liquid interface.

[0074] Figure 8 (a) in the diagram represents a conventional culture medium delivery mechanism, which involves pressurizing and delivering the culture medium to the target area via a tube. Because the culture medium is delivered under pressure, the longer the tube, the greater the consumption of culture medium. Even after a single delivery, culture medium residue remains in the tube. Therefore, the risk of culture medium quality problems due to temperature variations (approximately 15-25°C based on ambient temperature) is high. To address this issue of reduced culture medium quality, it may be necessary to refrigerate the entire tube.

[0075] Figure 8 (b) indicates the concept of a culture medium delivery mechanism utilizing a gas-liquid interface employed in this invention. This mechanism involves sequentially injecting culture medium and a gas, such as air, and using the pressure of the pressurized air to deliver the culture medium to the target area. Alternatively, the sequential injection of culture medium and gas can be repeated multiple times, with the later-injected gas ensuring that the previously injected culture medium reaches the target area sequentially. Because the culture medium is delivered using gas pressure, only gas remains in the tube after each delivery, with no culture medium residue. Therefore, even at room temperature, there is virtually no concern about temperature-induced degradation of the culture medium quality.

[0076] The effective diameter of the culture medium tube 260 affects the maximum and minimum volume of culture medium that can be transported at one time. Specifically, during the transport of culture medium, if the culture medium fails to completely fill the cross-section of the culture medium tube 260, the gas (air) will not be able to continue pushing the culture medium and will instead pass directly through the culture medium tube 260. Therefore, the culture medium to be transported needs to form a liquid column within the culture medium tube 260, and the length of the liquid column needs to be at least 1.5 times the inner diameter of the culture medium tube 260. For example, in a circular pipe with an inner diameter of D, the volume of culture medium that can be transported using the method of the present invention needs to be at least [amount missing]. above.

[0077] Similarly, the maximum volume of culture medium that can be delivered by the method according to the invention also depends on the inner diameter of the culture medium tube 260. When the culture medium tube 260 is vertically arranged, in order to prevent the culture medium from flowing backward due to gravity, the surface tension between the culture medium and the culture medium tube 260 needs to be balanced with the gravity acting on the culture medium. The inner diameter of the culture medium tube 260 that satisfies this condition depends on the density of the culture medium, the surface tension, and the surface characteristics of the culture medium tube 260, i.e., the contact angle. Furthermore, in the case of a culture medium tube 260 with an excessively small inner diameter, the resistance generated on the inner wall of the culture medium tube 260 when the culture medium flows through it causes an excessive pressure drop, thereby affecting the smooth supply of the culture medium. Therefore, it is necessary to take this situation into account and limit the minimum inner diameter.

[0078] The numerical range of the inner diameter of the culture medium tube 260 will be analyzed below using several formulas.

[0079] When the culture medium tube 260 is placed vertically, in order to prevent the liquid from flowing backward under the action of gravity, the surface tension between the liquid and the culture medium tube 260 must be balanced with the gravity of the liquid. Therefore, the following inequality holds regarding the length of the liquid (liquid segment, liquid column) that can move in the culture medium tube 260.

[0080] [Formula 1]

[0081] In the above inequalities, the density (ρ) and surface tension (T) of the liquid depend on the type of liquid moving, and the contact angle (θ) depends on the interaction between the liquid and the inner wall of the culture medium tube 260. As a result, the conditions for the maximum length of the movable liquid mentioned above, along with the aforementioned conditions for the minimum flow rate of the present invention, apply together and can be reduced to the following inequalities.

[0082] [Formula 2]

[0083] At this point, the minimum length of the movable liquid is directly proportional to the inner diameter of the culture medium tube 260, and the maximum length is inversely proportional to the inner diameter. Therefore, in order to use this invention as a means of conveying different amounts of liquid, it is highly advantageous to broaden its permissible conveying range using the above conditions. Thus, in this invention, it is intended to design the inner diameter of the culture medium tube 260 to be sufficiently small. In particular, in order to make the above inequality hold and realize the technical concept proposed in this invention, it is preferable to at least make the minimum condition for the length of the movable liquid, i.e., 1.5D, smaller than the maximum condition, i.e., The value of . According to this condition, in order to realize the present invention, the pipe diameter should meet the conditions shown in [Formula 3] below.

[0084] [Formula 3]

[0085] The results of multiple investigations, including those described above, indicate that, in order to achieve long-distance transport, the culture medium tube 260 preferably has a length of 15 cm or more for smooth, lossless liquid transport, and the inner diameter of the culture medium tube 260 is preferably 10 μm or more and 5 mm or less.

[0086] In one embodiment of the present invention, Figure 7 The culture medium supply module 200 includes a pump unit 230 comprising multiple syringe pumps 232. For each syringe pump 232, there are multiple check valves 234 and multiple branch connectors 236, which are connected to a culture medium tubing 260, thereby forming a piping network. This invention is configured such that all the syringe pumps 232, check valves 234, branch connectors 236, and culture medium tubing 260 can be made from commercially available plastic off-the-shelf products, thus enabling a practical disposable pump unit 230.

[0087] In fields with extremely high requirements for hygiene and quality management, such as life sciences, pharmaceuticals, and the food and beverage industry, various pump technologies (piston pumps, diaphragm pumps, peristaltic pumps, etc.) are used to accurately deliver liquid samples or solutions. However, such reusable pumps must undergo cleaning, sterilization, and validation processes after use. This not only increases the complexity and cost of the production process but also raises the risk of cross-contamination due to incomplete cleaning.

[0088] To address these issues, single-use pump technology was developed. Single-use pumps provide the core components of the pump, including the fluid contact parts, as disposable parts. Each batch uses a brand new pump head, thus preventing contamination at the source and eliminating the need for cleaning and sterilization processes. This simplifies the production process, improves productivity and efficiency, and simplifies validation.

[0089] However, most current commercially available disposable pump solutions primarily focus on maintaining a consistent flow rate and preventing contamination. This results in insufficient performance to meet the demands for precise metering. In other words, existing disposable pump technology prioritizes flow stability and basic pumping functionality, while neglecting accurate volume measurement and metering control. This invention aims to overcome these limitations of existing disposable pumps by providing a pump unit 230 that offers high metering accuracy while preventing contamination.

[0090] Figure 9 This is a diagram used to illustrate the concept of a culture medium delivery mechanism based on multiple syringe pumps 232. Figure 9The most basic component includes a gas injector 250 and a culture medium injector 252. The arrows indicate the direction of culture medium supply; based on the flow of the culture medium, the gas injector 250 is positioned upstream, and the culture medium injector 252 is positioned downstream of it. Figure 9 In the description, if the culture medium tube 260 is divided into a main tube 260-M and a branch tube 260-B, then the main tube 260-M is connected to the supply tube connection 122 of the cell culture module 100, and the branch tube 260-B, through the branch connector 236, respectively connects the gas injector 250 and the culture medium injector 252 to the main tube 260-M. Furthermore, check valves 234 are connected upstream and downstream of the branch connector 236. Through the check valves 234, gas (air) and culture medium can only flow in the single direction indicated by the arrow, prohibiting reverse flow.

[0091] The upstream side of the gas injector 250 is open to the atmosphere, and an air filter 236 for filtering foreign matter can be provided at the inlet of the check valve 234. The upstream side of the culture medium injector 252 is connected to the culture medium chamber 220, and a vent is formed to allow atmospheric pressure to act inside the culture medium chamber 220. This vent can also be equipped with an air filter 236.

[0092] Figure 9 (a) indicates the process of drawing culture medium stored in culture medium chamber 220 into culture medium injector 252. Through the aspiration stroke of culture medium injector 252, the culture medium stored in culture medium chamber 220 passes through upstream check valve 234 and fills branch tube 260-B and culture medium injector 252. However, due to downstream check valve 234, gas in main tube 260-M is not drawn into culture medium injector 252.

[0093] Figure 9 (b) in the diagram represents the process of filling the branch tube 260-B with culture medium via the discharge stroke of the culture medium syringe 252, and then quantitatively discharging the culture medium from the culture medium syringe 252 into the main tube 260-M. This process is described in conjunction with... Figure 9 In the opposite case (a), the check valve 234 operates, and the culture medium is not discharged to the culture medium chamber 220 side. As a result, a predetermined amount of culture medium corresponding to the length of the discharge stroke of the culture medium syringe 252 is discharged to the main pipe 260-M.

[0094] Figure 9 (c) indicates the process of introducing gas into the main pipe 260-M through the intake and exhaust strokes of the gas injector 250, and using pressurized gas to transport the culture medium within the main pipe 260-M in the direction of the arrow. The process of introducing gas into the main pipe 260-M through the intake and exhaust strokes of the gas injector 250 is essentially the same as... Figure 9The processes in (a) of 9 and (b) of 9 are the same. The culture medium can be delivered to the distal end of the gas injector 250, and the volume of gas drawn in and expelled can fill the distal end of the main tube 260-M.

[0095] Figure 10 This describes a piping network concept based on a syringe pump 232 that quantitatively delivers various liquids (such as culture medium) to and recovers the culture medium within the culture space 142. The culture medium tubing 260 has a dual-channel structure consisting of a supply tubing 262 and a discharge tubing 264. One gas syringe 250 and multiple culture medium syringes 252 are connected in parallel to the supply tubing 262 (main tubing) via branch tubing 260-B, branch connector 236, and check valve 234, respectively. Figure 9 Similarly, the gas injector 250 is located at the upstream end of the supply tube 262. Various liquids introduced from each culture medium injector 252 into the supply tube 262 are delivered to the cell culture module 100 side through the suction and discharge stroke of the gas injector 250. For example, in Figure 8 As illustrated in (b), the culture medium delivery between any culture medium injector 252 and gas injector 250 can be staggered, thereby allowing multiple liquids to be sequentially delivered to the cell culture module 100 side in a gas-separated, unmixed state. Therefore, multiple liquids can be delivered through a single supply tube 262 while significantly reducing the risk of cross-contamination.

[0096] In addition, Figure 10 A recovery syringe 254 for culture medium recovery is also shown. A recovery chamber 222 is connected to the downstream end of the discharge tube 264. The recovery syringe 254 is adjacent to the recovery chamber 222 and connected in parallel with the discharge tube 264. The branch tube 260-B, branch connector 236, check valve 234, and air filter 236 have the same configuration as the culture medium syringe 252, but the difference is that the check valve 234 of the recovery syringe 254 only allows flow in a single direction from the cell culture module 100 to the recovery chamber 222. In this piping network, through the suction and discharge strokes of the recovery syringe 254, the culture medium in the culture space 142 can be collected non-backflow through the discharge tube 264 into the recovery chamber 222.

[0097] Figure 11 This diagram illustrates the pre-filling process performed in the culture medium syringe 252. Pre-filling refers to the operation of filling the delivery pipe with fluid without leaving air during fluid delivery. In this invention, to achieve accurate quantitative delivery of the culture medium, the culture medium needs to fill the section from the culture medium chamber 220 to the end of the branch pipe 260-B (before it merges into the main pipe) without gaps. This can be achieved by performing the following steps: Figure 11 The pre-charge process is shown.

[0098] The pre-filling process of the culture medium syringe 252 is completed through a total of two aspiration and expulsion (pumping) operations. Figure 11 (a) ~ Figure 11 (c) shows the extent to which the culture medium fills the culture medium chamber 220 to the middle position of the branch connector 236 during a single pumping process. In the subsequent... Figure 11 (d) to Figure 11 In the secondary pumping process shown in (e), culture medium is further drawn in to fill the end of branch line 260-B. If more culture medium is pumped than this amount, some culture medium will flow into main line 260-M, resulting in waste; if less culture medium is pumped than this amount, residual gas will cause errors in the delivery volume.

[0099] Figure 12 It indicates the actual presentation Figure 10 The diagram illustrates one embodiment of a pump unit 230 within a conceptual piping network. For ease of explanation, Figure 12 In an exemplary embodiment, a gas injector 250, two culture medium injectors 252, and a recovery injector 254 are shown. Additionally, a branch connector 236, a check valve 234, and an air filter 236 connected to the gas injector 250 are also shown. Figure 12 For ease of understanding, the culture medium chamber 220 and the recovery chamber 222 are schematically shown, and the culture medium chamber 220 and the recovery chamber 222 are housed within the culture medium storage chamber 210 (see reference). Figure 7 The arrows indicate the direction of flow of the supplied and recovered culture medium. A large amount of gas is required to deliver the culture medium to the remote culture space 142; therefore, the gas injector 250 can be larger than the culture medium injector 252. Furthermore, the recovery injector 254 uses a single injector without distinguishing between injectors that draw in liquids and those that draw in gases; therefore, it can be roughly the same size as the gas injector 250. Since the recovery chamber 222 draws in multiple culture media, a chamber with a larger capacity than the culture medium chamber 220 can be used.

[0100] Figure 13 This diagram illustrates one embodiment of the pump unit 230, corresponding to an embodiment including the pump actuator 242 of each syringe pump 232 contained in the automatically driven pump unit 230. For ease of understanding of the invention, Figure 13 The branch connector 236 and check valve 234 are omitted in the design. The structure including the branch connector 236 and check valve 234 can be referred to [reference needed]. Figure 12 .

[0101] exist Figure 13In one embodiment, the pump unit 230 includes a gas injector 250, five culture medium injectors 252, and a recovery injector 254. It has a total of seven injector pumps 232, and correspondingly, seven pump actuators 242 (see reference). Figure 7 The pump actuator 242 is a linear actuator that automatically realizes the suction and discharge strokes of the syringe pump 232 by grasping the end of the plunger 240 of the syringe pump 232 and moving it linearly.

[0102] To facilitate the disassembly of the pump actuator 242 and the plunger 240 end of the syringe pump 232, a disassembly clamp 244 can be added to the end of the plunger 240. The plunger 240 and the pump actuator 242 can be connected by passing through the disassembly clamp 244 through a shaft extending from the pump actuator 242.

[0103] Figure 14 This diagram shows the interior of the culture medium supply module 200. The internal structure is shown by removing a portion of the culture medium supply module 200, which includes a culture medium storage chamber 210 housing multiple culture medium chambers 220, and a pump unit 230 opening above the culture medium storage chamber 210 with a rotary opening and closing structure. In addition to housing the multiple culture medium chambers 220, the culture medium storage chamber 210 can also accommodate a large-capacity recovery chamber 222. At least one cooling unit 212 (e.g., a Peltier unit) is provided in the culture medium storage chamber 210 to maintain a low-temperature environment of 4–8°C. Alternatively, the low-temperature environment of the culture medium storage chamber 210 can be maintained by placing the culture medium supply module 200 in a separate cold storage compartment that creates the low-temperature environment.

[0104] The low-temperature environment of the culture medium storage chamber 210 allows the culture medium stored in the culture medium chamber 220 to maintain excellent quality over a long period. Furthermore, preferably, the culture medium tubes 260, branch connectors 236, and check valves 234 forming a piping network are also disposed within the culture medium storage chamber 210. This is because, if quantitative delivery of the culture medium is achieved via… Figure 11 During the pre-filling process, the culture medium tube 260 upstream of the culture medium syringe 252 will also be filled with culture medium. Furthermore, preferably, cold air is also supplied to a portion of the pump unit 230, which is connected to the culture medium storage chamber 210, thereby creating a suitable low-temperature environment. For this purpose, the pump unit 230, which opens with a rotary opening and closing structure, can have a heat-insulating structure that seals the culture medium storage chamber 210.

[0105] Figure 15This diagram illustrates the temperature environment of the culture medium in the cell culture apparatus 10. In the culture medium supply module 200, the temperature of the culture medium is maintained at 4-8°C due to the low-temperature environment of the culture medium storage chamber 210. The culture medium tube 260, connecting the culture medium supply module 200 to the cell culture module 100, is exposed to a room temperature environment ranging from approximately 15-25°C. The cell culture module 100, typically placed in a CO2 incubator, maintains a temperature range of approximately 36-37°C. Therefore, during the supply of culture medium, a natural temperature regulation of the culture medium is achieved, progressing from low temperature to room temperature to a rise in culture temperature. This prevents the supply of excessively cold culture medium to the culture space 142, thereby preventing cells from experiencing thermal shock that could lead to functional decline or damage. Conversely, if the culture medium is stored at room temperature for too long, various proteins and other substances within the culture medium may decompose, making it difficult to create the environment required for cell culture. However, in this invention, such a problem of rising culture medium temperature can be solved by adjusting the delivery rate of the pressurized culture medium.

[0106] Figure 16 This diagram conceptually illustrates the configuration of multiple cell culture modules 100 connected to a single culture medium supply module 200. Supply pipes 262 and discharge pipes 264 extending from the culture medium supply module 200 are connected in parallel to the multiple cell culture modules 100. The process of supplying culture medium from a single cell culture module 200 to a target cell culture module 100 is controlled by a valve module 130 provided in each cell culture module 100.

[0107] That is, the valve modules 130 of all cell culture modules 100 except the cell culture module 100 which is the target of the culture medium are cut off from the flow of the culture medium. Thus, although it is a one-to-many pipeline network connecting one culture medium supply module 200 and multiple cell culture modules 100, in the actual delivery of the culture medium, a one-to-one correspondence is achieved through the coordinated opening and closing of multiple valve modules 130.

[0108] During the operation of the cell culture apparatus 10, the absolute time for delivering the culture medium is relatively short. Therefore, by configuring a one-to-many piping network that connects multiple cell culture modules 100 to a culture medium supply module 200, the culture medium supply module 200 can be used more effectively.

[0109] On the other hand, the culture medium supply module 200 uses a gas-liquid interface transport mechanism to deliver the culture medium. Therefore, even without directly measuring the flow rate inside the tube, the presence of liquid inside the tube can be determined by using a transparent or semi-transparent tube, thus indicating whether the culture medium supply module 200 is operating normally. As an embodiment for detecting abnormalities in the culture medium supply module 200, an infrared LED or other monitoring device can be used to determine whether the tube is properly installed and whether gas or liquid is present inside the tube. When a command to supply or discharge culture medium is input, the state inside the supply and discharge tubes can be monitored to determine whether the state changes from gas to liquid to gas, thereby confirming whether the culture medium is indeed being properly delivered to the culture medium supply and discharge tubes. Although not shown separately, the infrared LED or other monitoring device can be configured to directly contact the tube (by mounting the infrared LED or other monitoring device on a clamp that can hold the tube and irradiating or photographing it at close range), or it can be configured to irradiate light or photograph the tube at a distance separated from it.

[0110] The embodiments of the present invention have been described above with reference to the accompanying drawings. However, those skilled in the art will understand that the present invention can be implemented in other specific forms without changing its technical concept or essential features. Therefore, it should be understood that the embodiments described above are illustrative in all respects and not restrictive.

Claims

1. A cell culture apparatus, wherein, The cell culture apparatus includes: A cell culture module, comprising a cell culture section formed by mounting a well plate that creates at least one culture space onto a cartridge; A culture medium supply module includes: a pump unit for supplying and discharging culture medium to the cell culture section; and a culture medium storage chamber for accommodating at least one culture medium chamber; and A pair of flexible culture medium tubes connect the spatially separated culture medium supply module and cell culture module, forming a conduit for the supply and discharge of the culture medium. The cell culture module is placed in a CO2 incubator that creates a warm environment, the culture medium storage chamber creates a low temperature environment below room temperature, and a pair of culture medium tubes exposed between the culture medium supply module and the cell culture module are in a room temperature environment.

2. The cell culture apparatus according to claim 1, wherein, The pair of culture medium tubes serve as a supply tube and an outlet tube. The pump unit sequentially pressurizes and supplies a predetermined amount of culture medium and gas to the supply pipe, thereby supplying the culture medium to the culture space. The inner diameter of the supply tube is such that it can prevent the culture medium from flowing back due to gravity.

3. The cell culture apparatus according to claim 2, wherein, The pump unit draws culture medium from the culture space by applying negative pressure to the discharge pipe and returns it to the recovery chamber contained in the culture medium storage chamber.

4. The cell culture apparatus according to claim 1, wherein, The cell culture apparatus includes multiple cell culture modules connected in parallel to a culture medium supply module. By controlling the opening and closing of valve modules in each of the multiple cell culture modules, a one-to-one correspondence is achieved between the culture medium supply module and the cell culture module that is the delivery target during the generation of culture medium supply and / or recovery flow.

5. A culture medium supply module capable of supplying culture medium to at least one spatially separated culture space and recovering culture medium from said culture space, wherein, The culture medium supply module includes: a pump unit for supplying and draining culture medium to the cell culture section; and a culture medium storage chamber accommodating a recovery chamber and at least one culture medium chamber. The culture medium storage chamber is designed to maintain a low-temperature environment below room temperature. The pump unit supplies the culture medium stored in the culture medium chamber to each of the culture spaces through the supply pipe, and discharges it to the recovery chamber through the discharge pipe.

6. The culture medium supply module according to claim 5, wherein, The pump unit includes multiple syringe pumps. The plurality of syringe pumps include a gas syringe, at least one culture medium syringe, and a recovery syringe.

7. The culture medium supply module according to claim 6, wherein, The gas injector and at least one culture medium injector are connected in parallel with the supply tube. Check valves are installed upstream and downstream of the junction with the supply pipe.

8. The culture medium supply module according to claim 7, wherein, The gas injector is positioned at the upstream end of the supply tube. The check valves installed in the gas injector and the culture medium injector allow flow in only one direction toward the culture space.

9. The culture medium supply module according to claim 8, wherein, The gas injector has an air filter upstream of the upstream check valve.

10. The culture medium supply module according to claim 8, wherein, A predetermined amount of culture medium is introduced into the supply tube through the suction and discharge stroke of the culture medium syringe. The gas introduced into the supply tube through the inhalation and exhalation stroke of the gas injector delivers the culture medium introduced into the supply tube to the culture space.

11. The culture medium supply module according to claim 10, wherein, The plurality of syringe pumps includes gas syringes and a plurality of culture medium syringes. The delivery of culture medium between any culture medium syringe and the gas syringe is staggered, thereby allowing the various culture media to be delivered to the culture space in a gas-isolated state.

12. The culture medium supply module according to claim 6, wherein, The recovery syringe is connected to the downstream end of the discharge tube. Check valves are provided upstream and downstream of the junction of the recovery syringe and the discharge tube, respectively, thereby allowing flow only in a single direction toward the recovery chamber.

13. The culture medium supply module according to claim 6, wherein, The pump unit has multiple pump actuators to cause the plunger of each syringe pump to move linearly to perform the suction and discharge strokes.

14. A culture medium supply module capable of supplying culture medium to at least one spatially separated culture spaces, wherein, The culture medium supply module includes: The pump unit includes multiple syringe pumps for forming a supply flow of culture medium to the cell culture section; and a culture medium storage chamber for accommodating at least one culture medium chamber. The plurality of syringe pumps includes a gas syringe connected to the upstream end of the supply tube, and at least one culture medium syringe connected in parallel to the downstream side of the gas syringe. Check valves are installed upstream and downstream of the junction of the gas injector and each culture medium injector with the supply tube, respectively, to allow flow only in a single direction toward the culture space. A predetermined amount of culture medium is introduced into the supply tube through the suction and discharge stroke of the culture medium syringe, and the gas introduced into the supply tube through the suction and discharge stroke of the gas syringe delivers the culture medium introduced into the supply tube to the culture space.

15. The culture medium supply module according to claim 14, wherein, The plurality of syringe pumps includes gas syringes and a plurality of culture medium syringes. The delivery of culture medium between any culture medium syringe and the gas syringe is staggered, thereby allowing the various culture media to be delivered to the culture space in a gas-isolated state.

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