Cell culture system based on multi-sensor data fusion and flow-controllable microfluidic chip

Abstract

The present invention belongs to the field of cell culture technology and relates to a cell culture system with multi-sensor data fusion and flow-controllable microfluidic chip. The system comprises: a microfluidic chip, a liquid exchange device, a gas injection device, a multi-sensor data fusion subsystem, and a constant temperature subsystem. The microfluidic chip is provided with a culture fluid injection port and a cell culture chamber. The cell culture fluid flow channel from the culture fluid injection port to the cell culture chamber is in the shape of a multi-branched fluid channel. The liquid exchange device pipeline is connected to the culture fluid injection port, the gas injection device pipeline is connected to the gas injection port, and the multi-sensor data fusion subsystem and the constant temperature subsystem are electrically connected to a circuit board respectively. The present invention realizes monitoring and control of the cell culture environment, can monitor the cell status of multiple different cell culture chambers in real time, and promptly adjust the culture parameters according to model feedback to ensure the consistency of the cell populations in the multiple cell culture chambers, thereby improving the repeatability and reliability of the experimental results.

Description

Technical Field

[0001] The invention belongs to the technical field of cell culture, and relates to a cell culture system of multi-sensor data fusion and flow-controllable microfluidic chip. Background Art

[0002] Three-dimensional cell culture is a cell culture technology that improves the understanding and study of cell behavior by simulating the in vivo environment. This technology can maintain the inherent characteristics of cells, allowing for more accurate observation of cell behavior and exploration of its mechanisms. Three-dimensional cell culture is achieved by constructing three-dimensional scaffolds or using materials such as hydrogels to provide cells with a three-dimensional growth space that is closer to the in vivo environment, helping cells maintain their physiological functions and morphology. However, the culture process is relatively complex and the cost is relatively high. At the same time, although three-dimensional culture improves the cell growth environment, it is still difficult to accurately simulate the complex physiological microenvironment in the body, and the culture process is difficult to monitor in real time and precisely control.

[0003] In recent years, the emergence of microfluidic chip technology has brought new solutions to three-dimensional cell culture. This technology utilizes microfabrication techniques to construct tiny channels and chambers on a chip, enabling precise control of the cell microenvironment and enabling cell culture, manipulation, and analysis. Its advantages are significant, including the ability to mimic the physiological microenvironment in the body, providing cells with growth conditions that more closely resemble real-life conditions; enabling real-time monitoring and precise control of the cell culture process; and significantly reducing reagent and cell usage, lowering experimental costs.

[0004] However, existing cell culture systems based on microfluidic chips still have some shortcomings. First, in terms of multi-channel control, it is impossible to accurately control the flow rate of culture fluid input to multiple culture chambers at the same time, which increases the error between experimental results. Second, in terms of cell parameter monitoring, it is difficult to simultaneously monitor multiple cell parameters, such as cell metabolic activity, gene expression, protein secretion, etc., which limits the comprehensive understanding of the physiological state of cells. In addition, due to subtle differences between cell communities cultured in microfluidic chips, even under the same culture conditions, the observed results will still be different, which affects the repeatability and reliability of the experimental results to a certain extent.

[0005] Therefore, a device or method is needed to solve the above technical problems by monitoring and controlling the cell culture environment in real time and ensuring the consistency of the experimental environment of multiple cell culture chambers. Summary of the Invention

[0006] The technical solution adopted by the present invention to solve the technical problem is: a cell culture system with multi-sensor data fusion and flow-controllable microfluidic chip, including:

[0007] A microfluidic chip for monitoring and / or controlling cell culture parameters within a cell culture chamber disposed within the microfluidic chip; the cell culture parameters include: chamber flow rate, carbon dioxide concentration, oxygen concentration, temperature, metabolic activity, gene expression, and protein secretion; the chamber flow rate, carbon dioxide concentration, oxygen concentration, and temperature are monitored and controlled in real time, and metabolic activity, gene expression, and protein secretion are monitored in real time;

[0008] a liquid exchange device, used to inject cleaning liquid into the microfluidic chip for cleaning during the cleaning phase and subsequently aspirate the cleaning liquid, and also used to inject cell suspension and culture liquid into the microfluidic chip during the culture phase;

[0009] A gas injection device is used to quickly dry the microfluidic chip during the cleaning phase and to deliver culture gas to the cell culture chamber for cell culture during the culture phase; the culture gas includes oxygen and carbon dioxide;

[0010] The multi-sensor data fusion subsystem is used to monitor and analyze cell parameters in multiple cell culture chambers, thereby comprehensively and accurately understanding the physiological status of cells. It is also used to predict the culture results of different cell populations and adjust the culture conditions of each cell culture chamber in advance to reduce the result deviation caused by differences in cell populations, ensure the consistency of cell culture, and thus improve the repeatability and reliability of experimental results.

[0011] Constant temperature subsystem, used to control the temperature of the cell culture chamber to ensure the stability of the cell culture environment;

[0012] The microfluidic chip is provided with a culture fluid injection port and a cell culture chamber, and the cell culture fluid flow path from the culture fluid injection port to the cell culture chamber is in the form of multiple bifurcated fluid channels; the cell culture chamber is provided with a circuit board, a fluorescence sensor, a temperature sensor, a carbon dioxide sensor, an oxygen sensor, a serpentine resistance wire, and a gas injection port; the fluorescence sensor, the temperature sensor, the carbon dioxide sensor, the oxygen sensor, and the serpentine resistance wire are electrically connected to the circuit board respectively;

[0013] The cell culture system is equipped with multiple microfluidic chips;

[0014] The pipeline of the liquid exchange device is connected to the culture fluid injection port, the pipeline of the gas injection device is connected to the gas injection port, and the multi-sensor data fusion subsystem and the constant temperature subsystem are electrically connected to the circuit board respectively.

[0015] Preferably, the cell culture fluid flow channel from the culture fluid injection port to the cell culture chamber includes: the culture fluid injection port is respectively connected to the inlets of the two front-section flow channels, the outlets of the two front-section flow channels are respectively connected to the inlet of the differential balance flow channel, the outlet of the differential balance flow channel is connected to the inlet of the flow limiting flow channel, and the outlets of the flow limiting flow channel are respectively connected to the cell culture chamber and the flow reservoir.

[0016] More preferably, the microfluidic chip is provided with N culture fluid injection ports, 1≤N≤100, and N is an integer; the N culture fluid injection ports correspond to 2N front-end flow channels, 2N differential balance flow channels, 2N flow-limiting flow channels, 2N cell culture chambers, and 2N flow reservoirs, respectively.

[0017] More preferably, the connection point where the culture fluid injection port connects to the two front flow channels is a "Y"-shaped bifurcation; the "Y"-shaped bifurcation has better flow balance and distribution, so that the flow in the front flow channel can be distributed more evenly;

[0018] The connection points where the flow-through and flow-limiting channels connect the cell culture chamber and the flow reservoir are respectively formed into a "Y"-shaped bifurcation; the "Y"-shaped bifurcation has better flow distribution, so that the flow between the cell culture chamber and the flow reservoir can be further distributed and adjusted.

[0019] More preferably, the two front flow channels and the two differential balancing flow channels are arranged side by side along the direction of fluid flow, and a differential balancing diaphragm is arranged between the two differential balancing flow channels. The differential balancing diaphragm is made of elastic nanomaterials. The differential balancing diaphragm divides a fluid pipeline into two parallel differential balancing flow channels along the direction of fluid flow; the differential balancing diaphragm is used to adjust the flow rate between adjacent differential balancing flow channels.

[0020] More preferably, the connection mode of the flow-limiting channel connecting the cell culture chamber and the flow storage tank includes: the flow-limiting channel is provided with: a front-section flow-limiting channel, a cantilever beam, and a diversion pile in sequence along the fluid flow direction; the cantilever beam is made of elastic deformation material, and the cantilever beam is arranged on the left and right sides of the inner wall of the front-section flow-limiting channel; fluid passage openings are reserved on the upper and lower sides of the cantilever beam; the cantilever beam is used to reduce the liquid flow rate by elastic deformation when the liquid flow rate is too fast; therefore, the cantilever beam can be connected to the inner wall of the front-section flow-limiting channel in a variety of ways, such as a bracket or rib connection; fluid passage openings can be preset on the upper, lower, left and right sides of the cantilever beam, but at least fluid passage openings must be reserved on the upper and lower sides; the diversion pile is arranged at the "Y"-shaped bifurcation where the flow-limiting channel, the cell culture chamber, and the flow storage tank are connected, and the diversion pile is in the shape of a triangular cone; the diversion pile is used to limit the flow into the cell culture chamber flow channel by diversion when the liquid flow rate is too fast.

[0021] More preferably, the diversion pile is provided with a cell culture chamber flow channel and a reservoir flow channel, the two ends of the cell culture chamber flow channel are respectively connected to the front-section flow limiting flow channel and the cell culture chamber, and the two ends of the reservoir flow channel are respectively connected to the front-section flow limiting flow channel and the reservoir; the inlet height of the cell culture chamber flow channel is lower than the inlet height of the reservoir flow channel; when the liquid passes through the diversion pile, when the flow rate is normal, since the flow channel of the reservoir flow channel is higher than the cell culture chamber flow channel, the liquid will flow from the cell culture chamber flow channel into the cell culture chamber; and when the liquid flow rate is too large, the liquid level exceeds the bottom surface of the flow channel through the reservoir flow channel, and the liquid exceeding the height difference will flow from the reservoir flow channel into the reservoir, and the flow rate flowing into the cell culture chamber flow channel is controlled by the diversion effect.

[0022] Preferably, the liquid exchange device comprises: a first micro pump, a first hose, and a culture fluid pool, wherein both ends of the first hose are connected to the culture fluid injection port and the culture fluid pool respectively, and the first micro pump is arranged on the first hose;

[0023] The gas injection device includes: a second micro pump, a second hose, and a gas storage tank. The two ends of the second hose are respectively connected to the gas injection port and the gas storage tank. The second micro pump is arranged on the second hose.

[0024] Preferably, the multi-sensor data fusion subsystem further comprises: a PC, a WiFi receiver, a data fusion subsystem, a data analysis subsystem, and a negative feedback closed-loop control subsystem, wherein the WiFi receiver, the data fusion subsystem, the data analysis subsystem, and the negative feedback closed-loop control subsystem are arranged in the PC;

[0025] The constant temperature subsystem further includes: a voltage-stabilized power supply and a power line, wherein the voltage-stabilized power supply is electrically connected to the circuit board via the power line.

[0026] Preferably, the operation method of the cell culture system comprises the following steps:

[0027] Step 1: Inject cleaning fluid into the microfluidic chip through the liquid exchange device for cleaning, and then suck out the cleaning fluid through the liquid exchange device after cleaning is completed;

[0028] Step 2: Inject gas into the microfluidic chip through the gas injection device, and then suck out the remaining cleaning liquid through the liquid exchange device;

[0029] Step 3: After the microfluidic chip is completely dried, the cell suspension is injected into the cell culture chamber of the microfluidic chip through the liquid exchange device. Subsequently, the cell culture fluid is injected into the cell culture chamber of the microfluidic chip through the liquid exchange device as needed. During the injection process of the cell suspension and cell culture fluid, the microfluidic chip ensures that the flow rate entering multiple cell culture chambers remains consistent through the differential balancing flow channel, the flow limiting flow channel, the flow reservoir and the structures contained therein;

[0030] Step 4: Maintain the temperature of the cell culture chamber through the constant temperature subsystem, and monitor and analyze the cell parameters in multiple cell culture chambers through the multi-sensor data fusion subsystem.

[0031] The beneficial effects of the present invention are:

[0032] 1. The present invention establishes a miniaturized and intelligent cell culture system through multi-sensor data fusion and flow-controllable microfluidic chip, realizes the monitoring and control of the cell culture environment, and can directly realize cell culture and observation in the microfluidic chip.

[0033] 2. The differential balancing flow channel, flow limiting flow channel, flow reservoir and the structures contained therein of the present invention can ensure precise control of the flow rates of multiple cell culture chambers.

[0034] 3. The constant temperature subsystem of the present invention can accurately control the temperature of the cell culture chamber for a long time through the negative feedback closed-loop control subsystem, thereby ensuring the stability of the cell culture environment.

[0035] 4. The multi-sensor data fusion subsystem used in the present invention can realize simultaneous monitoring and in-depth analysis of multiple cell parameters. The machine learning algorithm in the system can quickly process and analyze these massive and complex data, explore the potential associations and patterns between the data, and establish a mapping model between the physiological state of the cell and multiple parameters through a deep learning algorithm. This allows for a comprehensive and accurate understanding of the physiological state of the cell, identifying key factors affecting cell growth and experimental results, and further establishing a prediction model based on a deep neural network based on these factors. This allows for prediction of the culture results of different cell populations before the experiment, adjusting the culture conditions in advance, and reducing the result deviations caused by differences in cell populations. At the same time, the system of the present invention can monitor the cell state in real time and adjust the culture parameters in a timely manner based on model feedback to ensure the consistency of the experimental conditions, thereby improving the repeatability and reliability of the experimental results. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 It is a schematic structural diagram of a cell culture system of the cell culture system of the present invention with multi-sensor data fusion and flow controllable microfluidic chip;

[0037] Figure 2 1 is a top view of a flow-controllable microfluidic chip when N=5 in an embodiment of the present invention;

[0038] Figure 3 1 is a schematic diagram of a partial structure of a microfluidic chip in an embodiment of the present invention;

[0039] Figure 4 For the present invention Figure 3 Schematic diagram of the AA cross section.

[0040] In the figure, 1. microfluidic chip; 100. culture medium injection port; 110. front section flow channel; 111. injection port connecting flow channel; 112. horizontal flow channel; 120. differential balance flow channel; 121. differential balance diaphragm; 130. flow limiting flow channel; 131. front section flow limiting flow channel; 132. cantilever beam; 133. diversion pile; 134. cell culture chamber flow channel; 135. reservoir flow channel; 140. cell culture chamber; 141. circuit board; 142. fluorescence sensor; 143. temperature sensor; 144. carbon dioxide sensor; 145. oxygen sensor; 146. serpentine resistance wire; 147. gas injection port ; 148. Wire through-hole; 150. Reservoir; 2. Liquid exchange device; 210. First micropump; 220. First hose; 230. Culture medium tank; 240. First sealing rubber ring; 3. Gas injection device; 310. Second micropump; 320. Second hose; 330. Gas storage tank; 340. Second sealing rubber ring; 4. Multi-sensor data fusion subsystem; 410. PC computer; 411. WiFi receiver; 412. Data fusion subsystem; 413. Data analysis subsystem; 414. Negative feedback closed-loop control subsystem; 5. Constant temperature subsystem; 510. Voltage-stabilized power supply; 520. Power cord. DETAILED DESCRIPTION

[0041] The following will provide a clear and complete description of the relevant technologies in the present invention in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of them. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.

[0042] refer to Figures 1 to 4As shown, the cell culture system based on multi-sensor data fusion and flow-controllable microfluidic chip in this embodiment has the characteristics of miniaturization, portability and intelligence compared to the existing cell culture system based on microfluidic chip, realizes the monitoring and control of the cell culture environment, can realize cell culture and observation directly in the microfluidic chip, can ensure the precise control of the flow of multiple cell culture chambers, can accurately control the temperature of the cell culture chamber for a long time, and ensure the stability of the cell culture environment. The multi-sensor data fusion subsystem adopted can help to realize the simultaneous monitoring and in-depth analysis of multiple cell parameters. The machine learning algorithm in the system can quickly process and analyze these massive and complex data, explore the potential correlations and patterns between the data, and establish a mapping model between the physiological state of cells and multiple parameters through deep learning algorithms, so as to fully and accurately understand the physiological state of cells, find out the key factors affecting cell growth and experimental results, and further establish a prediction model based on deep neural network based on these factors. The culture results of different cell populations are predicted before the experiment, the culture conditions are adjusted in advance, and the result deviation caused by differences in cell populations is reduced. At the same time, during the experiment, the system can monitor the cell status in real time and adjust the culture parameters in time according to the model feedback to ensure the consistency of the experimental conditions, thereby improving the repeatability and reliability of the experimental results.

[0043] As attached Figure 1 As shown, the cell culture system based on multi-sensor data fusion and flow-controllable microfluidic chip of this specific embodiment can be used to simulate the human physiological environment to quickly screen drugs, evaluate the effects and toxicity of drugs on different cells, shorten the R&D cycle, and reduce costs; it can be used to formulate personalized treatment plans based on the culture response of the patient's individual cells; it can be used to observe basic life activities such as cell growth, proliferation, differentiation, and apoptosis, and monitor changes in cell behavior in real time; it can be used to study the growth characteristics, metabolic laws, and group behavior of microbial cells; and the study of direct contact and signal exchange between cells helps to reveal the mechanisms of immune regulation and disease occurrence.

[0044] The overall structure of this embodiment includes: a microfluidic chip 1, a liquid exchange device 2, a gas injection device 3, a multi-sensor data fusion subsystem 4 and a constant temperature subsystem 5;

[0045] This specific embodiment establishes a miniaturized and intelligent cell culture system, realizes the monitoring and control of the cell culture environment, and can directly realize cell culture and observation in the microfluidic chip 1.

[0046] As attached Figure 2As shown, the microfluidic chip 1 comprises N culture fluid injection ports 100, 2N front-end flow channels 110, differential balance flow channels 120, flow limiting flow channels 130, cell culture chambers 140, and a reservoir 150. One culture fluid injection port 100 is connected to two front-end flow channels 110, and N is between 5 and 20. The microfluidic chip 1 is made of multi-layer PDMS material. Figure 3 shown.

[0047] The culture fluid inlet 100 is cylindrical with a diameter of 1 to 1.5 mm. The front flow channel 110 includes an inlet connecting channel 111 and a horizontal channel 112, which are used to transfer culture fluid from the culture fluid inlet 100 to the differential balancing channel 120. The inlet connecting channel 111 is 300 μm wide and forms a 22.5-degree angle with the horizontal, connecting the culture fluid inlet 100 and the horizontal channel 112. The horizontal channel 112 is 300 μm wide and connects the inlet connecting channel 111 and the differential balancing channel 120. The differential balancing channel 120 is 500 μm wide and connects the front channel 110 with the flow-limiting channel 130. Its cross-sectional area is larger than that of the front channel 110 and the flow-limiting channel 130. A differential balancing membrane 121 is provided between adjacent differential balancing channels 120 to regulate the flow rate between them, ensuring that the flow rates in adjacent differential balancing channels 120 are the same, while also transferring the culture medium to the flow-limiting channel 130. The differential balancing membrane 121 is approximately 200 μm thick and is injected with nanoparticles to increase its elasticity. The flow-limiting channel 130 connects the differential balancing channel 120, the cell culture chamber 140, and the flow reservoir 150, and is used to limit the flow of culture fluid entering the cell culture chamber 140 when the flow velocity is too high; the flow-limiting channel 130 is "Y"-shaped, and includes a front-section flow-limiting channel 131, a cantilever beam 132, a diversion pile 133, a cell culture chamber flow channel 134, and a flow reservoir flow channel 135. The front flow limiting channel 131 has a width of 300 μm, connecting the differential balance channel 120, the cell culture chamber channel 134 and the reservoir channel 135, and the cross-sectional area of ​​the channel is the same as that of the front flow channel; the cantilever beam 132 is arranged in the front flow limiting channel 131, the cantilever beam is 200 μm long and 50 μm wide, fixed on the side wall of the channel, and is not connected to the upper and lower planes of the channel. It is used to reduce the liquid flow rate by elastic deformation when the liquid flow rate is too fast; the diversion pile 133 is located between the cell culture chamber channel 134 and the reservoir channel 135, and is used to reduce the liquid flow rate by elastic deformation when the liquid flow rate is too fast. The diversion effect limits the flow into the cell culture chamber flow channel 134; the reservoir flow channel 135 is 300um wide and is located at one of the bifurcations of the "Y"-shaped flow limiting flow channel, connecting the front-section flow limiting flow channel 131 and the reservoir 150. The cross-sectional area of ​​the flow channel is smaller than that of the front-section flow limiting flow channel 131, and the bottom surface of the flow channel is 1 / 3 higher than that of the front-section flow limiting flow channel; the cell culture chamber flow channel 134 is 300um wide and is located at the other bifurcation of the "Y"-shaped flow limiting flow channel, connecting the front-section flow limiting flow channel 131 and the cell culture chamber flow channel 134. The cross-sectional area of ​​the flow channel is the same as that of the front-section flow limiting flow channel 131.The cell culture chamber 140 is rectangular in shape, with rounded corners on the bottom and top, measuring 5 mm long and 2 mm wide, with a 0.1 mm corner radius. The top houses a circuit board 141, a fluorescence sensor 142, a temperature sensor 143, a carbon dioxide sensor 144, an oxygen sensor 145, a serpentine resistance wire 146, a gas inlet 147, and a wire through hole 148. The circuit board 141 secures and powers the fluorescence sensor 142, temperature sensor 143, carbon dioxide sensor 144, oxygen sensor 145, and serpentine resistance wire 146. The fluorescence sensor 142 collects multi-dimensional data, including cell metabolic activity, gene expression, and protein secretion. The carbon dioxide sensor 144 monitors the carbon dioxide concentration in the cell culture chamber 140 in real time. The oxygen sensor 145 monitors the oxygen concentration in the cell culture chamber 140 in real time. The temperature sensor 143 monitors the temperature in the cell culture chamber 140 in real time. Serpentine resistance wire 146 is used to heat and control the temperature within cell culture chamber 140. Gas injection port 147 is cylindrical with a diameter of 1-1.5 mm and is used to connect to a gas injection device and introduce gas into cell culture chamber 140. Wire hole 148 is cylindrical with a diameter of 0.25 mm and is used to lead out the power cord. Reservoir 150 is a rounded rectangular shape with a length of 5 mm and a width of 1 mm, with a corner radius of 0.1 mm.

[0048] In the actual use of the microfluidic chip 1, when the flow velocities in two adjacent differentially balanced flow channels 120 are different, the differentially balanced diaphragm 121 is subjected to pressure, and the pressure direction is from the flow channel with a large flow rate to the flow channel with a small flow rate. Since the differentially balanced diaphragm 121 is injected with nanoparticles to increase elasticity, the differentially balanced diaphragm 121 will deform under the action of pressure, resulting in an increase in the width of the flow channel with a large flow rate. According to the continuity equation Q=vA (where Q is the flow rate, v is the flow rate, and A is the cross-sectional area of ​​the flow channel), the instantaneous flow channel cross-sectional area A increases when the flow channel width increases, the flow rate decreases, and the flow channel with a small flow rate decreases. The channel width is reduced, the cross-sectional area is reduced, and the flow rate is increased; the flow rate of the two adjacent differential balancing channels 120 when reaching the flow limiting channel 130 can be kept consistent; the cantilever beam 132 in the subsequent flow limiting channel 130 will not deform or work when the flow is normal, but when the flow is too large, it can reduce the flow rate and thus the flow through elastic deformation; at the same time, the liquid passes through the diversion pile 133 and is located between the cell culture chamber flow channel 134 and the reservoir flow channel 135. When the flow is normal, the bottom surface of the flow channel of the reservoir flow channel 135 is 1 / 3 higher than that of the front flow limiting channel 131, as shown in the attached figure. Figure 4As shown, all liquid flows from cell culture chamber flow channel 134 into cell culture chamber 140. However, if the liquid flow rate is too high, the liquid level exceeds the bottom surface of the flow channel through reservoir flow channel 135, and some liquid flows from reservoir flow channel 135 into reservoir 150, thereby controlling the flow rate into cell culture chamber flow channel 134 through the diversion effect. The differential balancing flow channel 120, flow limiting flow channel 130, reservoir 150, and the structure included in the present invention can ensure precise control of the flow rate of multiple cell culture chambers.

[0049] The fluid exchange device 2 includes a first micropump 210, a first hose 220, a culture fluid reservoir 230, and a first sealing rubber ring 240. These are used to inject cell suspension and culture fluid into the culture fluid inlet 100 of the microfluidic chip 1, as well as to inject cleaning fluid into the culture fluid inlet 100 of the microfluidic chip 1 for cleaning and aspirating the cleaning fluid. The first micropump 210 is used to transfer the cell suspension and culture fluid from the culture fluid reservoir 230 into the microfluidic chip 1 via the first hose 220, as well as to inject and aspirate the cleaning fluid. The first hose 220, used to transfer the cell suspension, culture fluid, and cleaning fluid, has an inner diameter of 0.25-0.5 mm. The culture fluid reservoir 230 is connected to the microfluidic chip 1 via the first hose 220 and the first micropump 210, storing the initial cell suspension and subsequent cell culture fluid and cleaning fluid. The first sealing rubber ring 240, with a diameter of 1.5 mm, seals the connection between the first hose 220 and the microfluidic chip 1 to prevent the ingress of external impurities.

[0050] The gas injection device 3 includes a second micropump 310, a second hose 320, a gas storage tank 330, and a second sealing rubber ring 340. These are used to inject gas into the gas injection port 147 within the cell culture chamber 140 of the microfluidic chip 1. The gas is a mixture of 3% oxygen, 5% carbon dioxide, and 92% nitrogen. This gas can be used to quickly dry the microfluidic chip 1 during the cleaning phase and provide the necessary culture environment during the cell culture phase. The second micropump 310 is used to transfer gas to the cell culture chamber 140. The second hose 320, used for gas transfer, has an inner diameter of 0.25-0.5 mm. The gas storage tank 330 is connected to the cell culture chamber 140 via the second micropump 310 and the second hose 320 to store the gas. The second sealing rubber ring 340, with a diameter of 1.5 mm, is used to seal the connection between the second hose 320 and the cell culture chamber 140 to prevent the ingress of external impurities.

[0051] The multi-sensor data fusion subsystem 4 includes multiple fluorescence sensors 142, temperature sensors 143, carbon dioxide sensors 144, and oxygen sensors 145 within the cell culture chambers 140 of the microfluidic chip 1, as well as a PC 410. The PC 410 communicates and transmits data with the multiple fluorescence sensors 142, temperature sensors 143, carbon dioxide sensors 144, and oxygen sensors 145 via a WiFi communication protocol. The PC includes a WiFi receiver 411, a data fusion subsystem 412, and a data analysis subsystem 413. The data fusion subsystem 412, located within the PC 410, is responsible for collecting and labeling the monitoring data from the multiple fluorescence sensors 142, temperature sensors 143, carbon dioxide sensors 144, and oxygen sensors 145, as well as for noise filtering and classification. The data analysis subsystem 413, located within the PC 410, uses the monitoring data processed by the data fusion subsystem 412 to train a deep neural network, discover potential correlations and patterns between the data, and establish a predictive model based on the deep neural network.

[0052] The multi-sensor data fusion subsystem 4 of this embodiment is capable of simultaneously monitoring and deeply analyzing multiple cell parameters. The machine learning algorithm in the system can rapidly process and analyze this massive and complex data, exploring potential connections and patterns between the data. Through deep learning algorithms, a mapping model between the physiological state of cells and multiple parameters is established, thereby comprehensively and accurately understanding the physiological state of cells, identifying key factors affecting cell growth and experimental results, and further establishing a prediction model based on deep neural networks based on these factors. This allows for the prediction of culture results for different cell populations before the experiment, allowing for pre-adjustment of culture conditions and reducing bias in results due to differences in cell populations. Furthermore, the multi-sensor data fusion subsystem 4 of this embodiment can monitor cell status in real time and promptly adjust culture parameters based on model feedback to ensure consistency in cell culture, thereby improving the repeatability and reliability of experimental results.

[0053] The constant temperature subsystem 5 includes a negative feedback closed-loop control subsystem 414, a temperature sensor 143, a regulated power supply 510, and a power cord 520. The regulated power supply 510 is used to power the fluorescence sensor 142, temperature sensor 143, carbon dioxide sensor 144, oxygen sensor 145, and serpentine resistance wire 146 on the circuit board 141 of the multiple microfluidic chips 1. The power cord 520 is introduced into the cell culture chamber 140 of the microfluidic chip 1 through the wire through-hole 148. The power cord 520 enters the microfluidic chip 1 through the wire through-hole 148 on the microfluidic chip 1 and is used to connect the regulated power supply 510 to the fluorescence sensor 142, temperature sensor 143, carbon dioxide sensor 144, oxygen sensor 145, and serpentine resistance wire 146 on the circuit board 141. The negative feedback closed-loop control subsystem 414 is located in the PC 410 , and uses a PI structure as the system controller, a temperature sensor 143 of the cell culture chamber 140 as the system signal input, and a serpentine resistance wire 146 as the system actuator to form a complete negative feedback closed-loop control subsystem 414 . After the temperature is set in the PC 410 , the negative feedback closed-loop control subsystem 414 is used to maintain a constant temperature in the cell culture chamber 140 .

[0054] The constant temperature subsystem 5 of this specific embodiment can accurately control the temperature of the cell culture chamber 140 for a long time through the negative feedback closed-loop control subsystem 414. When the temperature is greater than the preset temperature, the heating of the serpentine resistance wire 146 is stopped; when the temperature is less than the preset temperature, the serpentine resistance wire 146 starts heating; thereby keeping the temperature of the cell culture chamber 140 constant and ensuring the stability of the cell culture environment.

[0055] This embodiment further relates to a method for implementing cell culture based on multi-sensor data fusion and flow-controllable microfluidic chip, which is applicable to the cell culture system of this embodiment, and the method comprises:

[0056] Step 1: Inject cleaning liquid into the microfluidic chip 1 through the liquid exchange device 2 and clean it. After cleaning, the cleaning liquid is sucked out through the liquid exchange device 2;

[0057] Step 2: Inject gas into the microfluidic chip 1 through the gas injection device 3, and then suck out the remaining cleaning liquid through the liquid exchange device 2.

[0058] Step 3: After the microfluidic chip 1 is completely dried, the cell suspension is injected into the cell culture chamber 140 of the microfluidic chip 1 through the liquid exchange device 2. Subsequently, the cell culture fluid is injected into the cell culture chamber 140 of the microfluidic chip 1 through the liquid exchange device 2 as needed. During the injection process of the cell suspension and cell culture fluid, the microfluidic chip 1 ensures that the flow rate entering multiple cell culture chambers 140 remains consistent through the differential balancing flow channel 120, the flow limiting flow channel 130, the reservoir 150 and the structures contained therein.

[0059] Step 4: The temperature of the cell culture chamber 140 is kept constant at the set value by the constant temperature subsystem 5. At the same time, the multi-sensor data fusion subsystem 4 is used to simultaneously monitor and deeply analyze the cell parameters in multiple cell culture chambers 140 to establish a prediction model.

[0060] In summary, the present invention establishes a miniaturized, intelligent cell culture system through multi-sensor data fusion and a flow-controllable microfluidic chip. This system enables monitoring and control of the cell culture environment, enabling cell culture and observation directly within the microfluidic chip. Furthermore, the system can monitor the cell status of multiple different cell culture chambers in real time and adjust culture parameters based on model feedback, ensuring the consistency of cell populations across multiple cell culture chambers, thereby improving the repeatability and reliability of experimental results. Therefore, the present invention has broad application prospects in the field of cell culture.

[0061] It should be emphasized that the above are only preferred embodiments of the present invention and do not limit the present invention in any form. Any simple modifications, equivalent changes and modifications made to the above embodiments based on the technical essence of the present invention are still within the scope of the technical solution of the present invention.

Claims

1. A cell culture system based on multi-sensor data fusion and flow controllable microfluidic chip, characterized in that: include: A microfluidic chip (1) is used to monitor and / or control cell culture parameters of a cell culture chamber (140) disposed within the microfluidic chip (1); the cell culture parameters include: chamber flow, carbon dioxide concentration, oxygen concentration, temperature, metabolic activity, gene expression, and protein secretion; A liquid exchange device (2) is used to inject a cleaning liquid into the microfluidic chip (1) for cleaning and to suck out the cleaning liquid during the cleaning phase, and is also used to inject a cell suspension and a culture liquid into the microfluidic chip (1) during the culture phase; A gas injection device (3) is used to dry the microfluidic chip (1) during the cleaning phase and to deliver culture gas to the cell culture chamber (140) for cell culture during the culture phase; the culture gas includes oxygen and carbon dioxide; a multi-sensor data fusion subsystem (4) for monitoring and analyzing cell parameters of a plurality of cell culture chambers (140), and for predicting culture results of different cell populations and adjusting culture conditions of each cell culture chamber (140); a constant temperature subsystem (5), for controlling the temperature of the cell culture chamber (140); The microfluidic chip (1) is provided with a culture fluid injection port (100) and a cell culture chamber (140), and a cell culture fluid flow channel along the direction from the culture fluid injection port (100) to the cell culture chamber (140) is in the shape of a plurality of bifurcated fluid channels; the cell culture chamber (140) is provided with a circuit board (141), a fluorescence sensor (142), a temperature sensor (143), a carbon dioxide sensor (144), an oxygen sensor (145), a serpentine resistance wire (146), and a gas injection port (147); the fluorescence sensor (142), the temperature sensor (143), the carbon dioxide sensor (144), the oxygen sensor (145), and the serpentine resistance wire (146) are respectively electrically connected to the circuit board (141); The pipeline of the liquid exchange device (2) is connected to the culture fluid injection port (100), the pipeline of the gas injection device (3) is connected to the gas injection port (147), and the multi-sensor data fusion subsystem (4) and the constant temperature subsystem (5) are respectively electrically connected to the circuit board (141); The cell culture fluid flow channel from the culture fluid injection port (100) to the cell culture chamber (140) comprises: the culture fluid injection port (100) is respectively connected to the inlets of two front section flow channels (110), the outlets of the two front section flow channels (110) are respectively connected to the inlet of the differential balancing flow channel (120), the outlet of the differential balancing flow channel (120) is respectively connected to the inlet of the flow limiting flow channel (130), and the outlet of the flow limiting flow channel (130) is respectively connected to the cell culture chamber (140) and the reservoir (150); The multi-sensor data fusion subsystem (4) comprises: a PC (410), a WiFi receiver (411), a data fusion subsystem (412), a data analysis subsystem (413), and a negative feedback closed-loop control subsystem (414); the WiFi receiver (411), the data fusion subsystem (412), the data analysis subsystem (413), and the negative feedback closed-loop control subsystem (414) are arranged in the PC (410); The constant temperature subsystem (5) further comprises: a voltage-stabilized power supply (510) and a power line (520), wherein the voltage-stabilized power supply (510) is electrically connected to the circuit board (141) via the power line (520).

2. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 1, characterized in that: The microfluidic chip (1) is provided with N culture solution injection ports (100), where 1≤N≤100, and N is an integer.

3. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 1, characterized in that: The culture fluid injection port (100) is connected to the two front flow channels (110) at a connection point that is forked in a "Y" shape; The flow-through and flow-limiting flow channels (130) are connected to the cell culture chamber (140) and the flow reservoir (150) at their connection points, forming a "Y"-shaped bifurcation.

4. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 3, characterized in that: The two front flow channels (110) and the two differential balancing flow channels (120) are arranged side by side along the fluid flow direction; a differential balancing diaphragm (121) is arranged between the two differential balancing flow channels (120); the differential balancing diaphragm (121) is made of elastic nanomaterial; and the differential balancing diaphragm (121) separates a fluid pipeline into two parallel differential balancing flow channels (120) along the fluid flow direction.

5. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 3, characterized in that: The communication mode of the flow-through and flow-limiting channel (130) connecting the cell culture chamber (140) and the flow reservoir (150) includes: the flow-through and flow-limiting channel (130) is provided with: a front-section flow-limiting channel (131), a cantilever beam (132), and a diversion pile (133) in sequence along the fluid flow direction; the cantilever beam (132) is made of elastic deformable material; the cantilever beam (132) is arranged on the left and right sides of the inner wall of the front-section flow-limiting channel (131); and fluid passage openings are reserved on the upper and lower sides of the cantilever beam (132); The diversion pile (133) is arranged at a "Y"-shaped bifurcation where the flow-through and flow-limiting flow channel (130), the cell culture chamber (140), and the flow reservoir (150) are connected. The diversion pile (133) is in the shape of a triangular pyramid.

6. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 5, characterized in that: The diversion pile (133) is provided with a cell culture chamber flow channel (134) and a flow reservoir flow channel (135); the two ends of the cell culture chamber flow channel (134) are respectively connected to the front-section flow limiting flow channel (131) and the cell culture chamber (140); the two ends of the flow reservoir flow channel (135) are respectively connected to the front-section flow limiting flow channel (131) and the flow reservoir (150); the inlet height of the cell culture chamber flow channel (134) is lower than the inlet height of the flow reservoir flow channel (135).

7. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 1, characterized in that: The liquid exchange device (2) comprises: a first micro pump (210), a first hose (220), and a culture fluid pool (230); the two ends of the first hose (220) are respectively connected to the culture fluid injection port (100) and the culture fluid pool (230); the first micro pump (210) is arranged on the first hose (220); The gas injection device (3) comprises: a second micro pump (310), a second hose (320), and a gas storage tank (330); the two ends of the second hose (320) are respectively connected to the gas injection port (147) and the gas storage tank (330); and the second micro pump (310) is arranged on the second hose (320).

8. The cell culture system of multi-sensor data fusion and flow controllable microfluidic chip according to claim 1, characterized in that: The operating method of the cell culture system comprises the following steps: Step 1: injecting a cleaning liquid into the microfluidic chip (1) through the liquid exchange device (2) for cleaning, and then sucking out the cleaning liquid through the liquid exchange device (2) after the cleaning is completed; Step 2: injecting gas into the microfluidic chip (1) through the gas injection device (3), and then sucking out the remaining cleaning liquid through the liquid exchange device (2); Step 3: After the microfluidic chip (1) is completely dried, the cell suspension is injected into the cell culture chamber (140) of the microfluidic chip (1) through the liquid exchange device (2); Step 4: The temperature of the cell culture chamber (140) is maintained by the constant temperature subsystem (5), and the cell parameters in the multiple cell culture chambers (140) are monitored and analyzed by the multi-sensor data fusion subsystem (4).

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