Cell replenishment method and cell culture integrated device

Abstract

The application discloses a cell liquid supplementing method and a cell culture integrated device, and relates to the technical field of cell engineering. The cell liquid supplementing method comprises the following steps: in the process of cell culture through a cell culture container made of transparent material, a color image of the cell culture container is acquired in response to a color acquisition instruction; color values of the color image are clustered to obtain a color value clustering result; a current dominant color value of a cell solution in the cell culture container is determined according to the color value clustering result; whether the culture medium needs to be supplemented into the cell culture container at present is determined through a liquid supplementing prediction model obtained through pre-training based on the dominant color value; and if it is determined that the culture medium needs to be supplemented into the cell culture container at present, the culture medium is added into the cell culture container. By using the cell liquid supplementing method, the suitable liquid supplementing time can be determined according to the dominant color change in the cell growth process, and the accuracy of determining the liquid supplementing time is improved.

Description

Technical Field

[0001] This application relates to the field of cell engineering technology, and in particular to a cell rehydration method and an integrated cell culture device. Background Technology

[0002] During cell culture, fresh culture medium needs to be added to the cell solution to maintain the nutritional environment required for cell growth; this process is called cell replenishment. In existing incubators or culture flasks, cell culture is typically performed by manually observing color changes in the cell solution and replenishing the medium accordingly. For example, replenishment is done manually when the cell solution turns yellow. Alternatively, a timed replenishment method can be used, where replenishment is performed at regular intervals.

[0003] However, both of the aforementioned traditional fluid resuscitation methods have drawbacks. Regarding the method of determining the timing of resuscitation through manual observation, different samples have different cell growth rates, and the accuracy of human observation is insufficient. Using this method, it is easy to miss the optimal resuscitation time, which can lead to slow cell growth, failure to reach the required cell number, or even cell growth arrest or cell death. As for the timed resuscitation method, it cannot be based on the specific growth stage of the cells, easily getting stuck in the initial retardation phase or the cell death phase, also easily missing the optimal resuscitation time. Summary of the Invention

[0004] This application provides a cell rehydration method and an integrated cell culture device to address or at least partially address the deficiencies or shortcomings in related technologies.

[0005] In a first aspect, this application provides a cell rehydration method, the cell rehydration method comprising: During cell culture using a transparent cell culture container, a color image of the cell culture container is acquired in response to a color acquisition command. Cluster the color values ​​of the color image to obtain color value clustering results; Based on the color value clustering results, the current dominant hue color value of the cell solution in the cell culture container is determined; Based on the primary color value, a pre-trained fluid replenishment prediction model is used to determine whether culture medium needs to be added to the cell culture container. If it is determined that the culture medium needs to be added to the cell culture container, then the culture medium is added to the cell culture container.

[0006] Secondly, this application provides an integrated cell culture device, the integrated cell culture device comprising: Culture medium box, which has space for placing culture medium containers for storing culture medium; A cell culture chamber includes a coating device and a culture device. The coating device has a space for placing a cell source container and a coating container. The culture device has a space for placing a cell culture container. The space for placing the cell culture container is equipped with a matching color image sensor and a reflective component. The reflective component is located in the light source emission direction of the matching color image sensor. The valve system includes a first valve device for controlling the pipeline between the culture medium container and the coating container, a second valve device for controlling the pipeline between the cell source container and the coating container, and a third valve device for controlling the pipeline between the coating container and the cell culture container. When the culture medium container, the coating container, the cell source container, and the cell culture container are all connected by pipelines, a closed environment is formed. A control module is electrically connected to the culture medium tank, the cell culture tank, and the valve system, respectively. The control module contains a pre-trained fluid replenishment prediction model and is used to implement the following steps: During cell culture via the cell culture container, in response to a color acquisition command, the color image sensor is controlled to acquire a color image of the cell culture container. Cluster the color values ​​of the color image to obtain color value clustering results; Based on the color value clustering results, the current dominant hue color value of the cell solution in the cell culture container is determined; Based on the primary color value, the replenishment prediction model determines whether culture medium needs to be added to the cell culture vessel. If it is determined that the culture medium needs to be added to the cell culture container, then the first valve device and the third valve device are opened.

[0007] According to the specific embodiments provided in this application, the following technical effects are disclosed: In this embodiment of the application, during cell culture using a transparent cell culture container, a color image of the cell culture container is first acquired in response to a color acquisition command, thereby obtaining the color data of the cell solution in the transparent cell culture container. Then, the color values ​​of the acquired color image are clustered to identify the hue distribution characteristics of the cell solution. Subsequently, based on the color value clustering results, the current dominant hue value of the cell solution can be determined. The relationship between the dominant hue value of the cell solution and the replenishment requirement is analyzed using a replenishment prediction model to determine whether replenishment is currently needed. If it is determined that replenishment is currently needed, the culture medium is added to the cell culture container to achieve cell replenishment. This application embodiment monitors the color changes of cell solutions using color image data of cell solutions that can accurately represent color. It accurately anchors the current dominant color of the cell solution through clustering of image color values, eliminating the influence of local color in the cell solution. It also uses a fluid replenishment prediction model to clarify the fluid replenishment requirement corresponding to the current dominant color of the cell solution. Compared with traditional methods, color clustering based on cell solution color images improves the accuracy of determining the dominant color of the cell solution. By associating the dominant color of the cell solution with the fluid replenishment requirement through the fluid replenishment prediction model, it can determine the appropriate time for fluid replenishment based on the changes in the dominant color during the specific growth process of cells, thereby improving the accuracy of determining the timing of fluid replenishment and avoiding missing the optimal time for fluid replenishment. Attached Figure Description

[0008] Figure 1 A schematic flowchart of a cell rehydration method provided in this application embodiment; Figure 2 A structural block diagram of an integrated cell culture device provided in an embodiment of this application; Figure 3 A structural block diagram of another integrated cell culture device provided in the embodiments of this application; Figure 4 A schematic diagram of the physical structure of the internal coating and culture platform of a cell culture incubator provided in an embodiment of this application; Figure 5 A schematic diagram of the physical assembly of a culture platform in an integrated cell culture device provided in this application embodiment; Figure 6 A schematic diagram of a protective cover for a cell culture container in an integrated cell culture device provided in this application embodiment; Figure 7 This is a schematic diagram of the physical structure of an integrated cell culture device provided in an embodiment of this application; Figure 8 This is a schematic diagram of a cell culture system provided in an embodiment of this application. Detailed Implementation

[0009] Before detailing the specific implementation methods of this application, we will first further introduce the prior art.

[0010] In existing technologies, determining the timing of fluid resuscitation typically employs two methods mentioned in the background section: one is to determine the timing of resuscitation by manually observing changes in the color of the cell solution (referred to as prior art 1), and the other is to resuscitate at set times (referred to as prior art 2). Neither of these methods can accurately determine the timing of resuscitation, easily leading to missing the optimal resuscitation time. Resuscitation that is done too early or too late can easily cause slow cell growth or even growth arrest.

[0011] Compared to Existing Technology 1, Existing Technology 2 is more widely used in industrialized cell culture because it can be designed as an automated mechanism. Therefore, the following will focus on the shortcomings of Existing Technology 2. The cell growth cycle is generally divided into four phases: initial lag phase, logarithmic phase, stationary phase, and death phase. In the initial lag phase, cells need to synthesize enzymes and metabolites to adapt to the environment; at this time, the cell number does not increase significantly, and the growth curve is flat. In the logarithmic phase, after adapting to the environment, cells are metabolically active, the division rate is at its maximum, the cell number increases logarithmically, and the growth curve rises sharply. In the stationary phase, due to nutrient consumption, accumulation of metabolites, and space constraints, the number of newly added cells and the number of dead cells tend to balance, and the growth curve tends to flatten. In the death phase, nutrients are depleted, harmful metabolites accumulate in large quantities, the cell death rate exceeds the division rate, the cell number decreases significantly, and the growth curve shows a downward trend. Due to sample differences, the time experienced in each phase varies. Therefore, the timed fluid replenishment method of Existing Technology 2 is prone to getting stuck in the initial lag phase or the death phase when fluid replenishment is performed.

[0012] Furthermore, for fast-growing cells, existing technology 2 results in prolonged overall cell culture time due to imprecise timing of fluid replenishment. For slow-growing cells, existing technology 2 is even less able to achieve precise timing of fluid replenishment compared to existing technology 1, easily causing cells to stop growing or even undergo apoptosis, leading to culture failure.

[0013] Currently, to accurately determine the timing of fluid replacement, cell density is typically determined during cell culture by opening the cell culture bag and taking a sample (i.e., opening the cell culture container and removing a portion of the cell solution). Based on the cell density, it is then determined whether fluid replacement should be initiated; if the cell density is high, fluid replacement is performed to ensure each cell has sufficient nutrients for growth. However, this method of determining fluid replacement timing by opening the bag for sampling (referred to as prior art 3) is highly susceptible to cell contamination. This is especially problematic in medical cell engineering, where cultured cells are often used for treatment, vaccine development, and other applications directly related to human life and safety. Contamination of cultured cells can lead to a series of serious consequences.

[0014] To further avoid cell contamination during bag opening and sampling, a method is currently used to construct a sterile environment and perform bag opening and sampling in a sterile environment to determine the timing of fluid resuscitation (referred to as prior art 4).

[0015] However, existing technology 4 still requires manual sampling and fluid replenishment. Therefore, constructing a sterile environment requires additional facilities for manual operations in the sampling and fluid replenishment process. These facilities require at least one sterile operating box (a device equipped with operating gloves, where operators must insert their hands into the gloves to perform the operations) or one sterile operating room (a room where operators must enter to perform the operations). Currently, the facilities used are mostly a combination of a sterile operating box and a sterile operating room. Both the sterile operating box and the cell culture device are located in the sterile operating room. Operators enter the sterile operating room to perform cell culture using the cell culture device and to perform manual sampling and fluid replenishment in the sterile operating box. However, such sterile conditions are a very demanding laboratory-level sterile environment, which is difficult to achieve. In addition, the sterile environment of a sterile operating box combined with a sterile operating room consumes a lot of energy.

[0016] Furthermore, to avoid cross-contamination between different sample cells (e.g., cells extracted from two different organisms), a sterile operating chamber or even a sterile operating room typically only culturees one sample of cells. For example, in the medical field, cells are cultured from the patient's own body and then reinfused, requiring strict isolation from contamination. Therefore, the difficulty in achieving a sterile environment and the stringent requirements for isolation between different sample cells make it difficult to industrialize existing technology 4.

[0017] For example, in response to the needs of hospitals or medical companies to culture medical cells, hospitals or medical companies do not have the capability to create such a rigorous and large-scale sterile culture environment for a single sample of cells. Among these, sterile operating boxes + cell culture devices, or sterile rooms, are all large-scale sterile culture environments.

[0018] Based on the problems existing in the prior art 1-4, this application provides a cell rehydration method and an integrated cell culture device.

[0019] The specific implementation methods of the cell rehydration method and integrated cell culture equipment provided in this application will be described in detail below.

[0020] In the embodiments of this application, such as Figure 1 As shown, this cell rehydration method includes the following steps S11 to S15. Wherein: Step S11: During cell culture using a transparent cell culture container, a color image of the cell culture container is acquired in response to a color acquisition command.

[0021] The cell source required for cell culture can include any type such as immune cells or stem cells. In the embodiments of this application, it can be determined whether the coating container needs to be pretreated before cell culture, such as through a coating cleaning process, depending on the type of cell source. For example, for immune cells, the coating container can be cleaned before cell culture, and then the primary cells can be transferred to the coating container for culture. After a certain period of culture, the cells can be transferred to a cell culture container for further cell culture. As another example, for stem cells, cell culture can be performed directly without pretreatment.

[0022] Alternatively, the cell culture container can be a bag-shaped container, i.e., a cell culture bag.

[0023] In this embodiment, color images of the cell culture container need to be acquired during cell culture. Therefore, to ensure that the color images accurately reflect the color characteristics of the cell solution in the cell culture container, the cell culture container can be made of a transparent material. Optionally, the transparent material may include FEP (Fluorinated ethylene propylene copolymer), which has high light transmittance (up to 95% or more, suitable for non-contact detection) and high air permeability (oxygen transmittance 3200 ml / (m²)). 2 The carbon dioxide transmission rate is approximately 6776 ml / (m·day·atm). 2 It has a permeability of approximately 0.5 ml / (m·day·atm), ensuring the necessary gas and osmotic pressure environment for cell growth; high water resistance (water vapor permeability is 0.5 ml / (m·day·at ... 2 It has the characteristics of low light transmittance (around 1000 ml / (m·day·atm), preventing cell solution evaporation and thus eliminating the need for additional humidity control of the cell solution; low precipitation (due to its inert material and stable chemical properties); and non-toxicity. In contrast, EVA (Ethylene Vinyl Acetate) material used in traditional cell culture bags has a light transmittance of approximately 80% and an oxygen transmittance of approximately 2600 ml / (m·day·atm·day). 2 ·day·atm), water vapor transmission rate 5ml / (m 2 (day·atm), chemical inertness is generally low, and safety level is lower than that of FEP material. Compared with EVA material, FEP material cell culture containers are more suitable for cell culture, and their high light transmittance makes it easier to collect accurate cell solution color data.

[0024] In this embodiment of the application, during cell culture using a cell culture container made of transparent material, if a color acquisition command is detected, the color acquisition command is responded to, and a color image of the cell culture container is acquired.

[0025] The color acquisition command can be triggered by the control module in the cell culture integrated device. The triggering time of the color acquisition command can be preset, and optionally, a mechanism for manually triggering the color acquisition command can be added to adapt to personalized needs.

[0026] In one optional implementation, the color acquisition instruction can be a periodic instruction preset based on multiple time intervals. The color acquisition instruction can be triggered and executed once every time interval according to the order of the multiple time intervals, and the multiple time intervals can be the same time interval or at least partially different time intervals.

[0027] For example, a color acquisition command can be triggered once every other time interval, arranged in a sequence of multiple identical time intervals. For instance, the color acquisition command could be set to trigger every 2 hours during cell culture, allowing a color image of the cell culture vessel to be acquired every 2 hours, thereby periodically determining whether fluid replenishment is needed based on the color image.

[0028] For example, color acquisition commands can be triggered at regular intervals, arranged in a sequence of at least partially distinct time intervals. Based on the first two of the four phases of the cell growth cycle described above, for instance, the color acquisition commands could be set to trigger every 5 hours during the first two days of cell culture (corresponding to the initial lag phase, the duration of which depends on the specific cell species), and then every 2 hours thereafter (corresponding to the logarithmic phase). It should be noted that cell harvesting, after cell culture is complete, is typically performed during the logarithmic phase.

[0029] In one optional implementation, a color image sensor can be used to acquire a color image of the cell culture container, thereby acquiring color data of the cell solution. In order to accurately acquire the color image, the color image sensor and the cell culture container can be set close to the cell culture container. For example, the color image sensor can be set at the placement position of the cell culture container, so that when the cell culture container is placed at this placement position, the color image sensor can be close to the bottom of the cell culture container to acquire the color image.

[0030] Optionally, the color image sensor may include a linear color image sensor, such as a CIS sensor (Contact Image Sensor). In a linear color image sensor, the sensing units are arranged linearly, and a line of pixel data can be acquired through line scanning. The pixel value can represent color, such as RGB (Red, Green, Blue) values. Compared to area-array color image sensors such as CCD (Charge Coupled Device) sensors and CMOS (Complementary Metal Oxide Semiconductor) sensors, linear color image sensors occupy less space, a significant advantage in cell culture. Because cells require aeration during culture, such as oxygen and carbon dioxide, the hardware used to place the cell culture container (e.g., the culture platform in the integrated cell culture device mentioned below) needs to ensure good gas interaction between the cell culture container and the gaseous environment. The smaller size of a linear color image sensor allows for more space on the hardware used to place the cell culture container to ensure aeration.

[0031] In one optional implementation, the step of acquiring a color image of the cell culture vessel using a linear color image sensor may specifically include: During the relative motion between the cell solution in the cell culture container and the linear color image sensor, the cell culture container is continuously photographed multiple times by the linear color image sensor to obtain multiple sets of linear color pixel data. A color image is obtained by stitching together multiple linear color pixel data.

[0032] In this embodiment, since the linear color image sensor can only acquire one line of color pixel data at a time, i.e., one linear color pixel data, the data volume of one line of color pixel data is small, which can easily lead to inaccurate determination of the dominant color value of the cell solution and reduce the reliability of the results. Therefore, the cell culture container can be continuously photographed multiple times by the linear color image sensor. Moreover, this continuous multiple photography is carried out while the cell solution in the cell culture container is in relative motion with the linear color image sensor. The relative motion can create a flowing acquisition window, thereby acquiring multiple linear color pixel data of the dynamic cell solution. Stitching multiple linear color pixel data can obtain a two-dimensional color image, that is, obtain the two-dimensional spatial distribution information of the color of the cell solution, avoiding the misjudgment or inaccuracy caused by single-line sampling. In this embodiment, multiple linear color pixel data objectively reflect the overall color of the cell solution, without the need to collect color pixel data at every position of the cell solution, thus effectively balancing the computational efficiency and accuracy of color determination.

[0033] The number of times the linear color image sensor can continuously capture images of the cell culture container can be preset parameters, such as 4, 5, 6 or 7 times, etc.

[0034] In an optional embodiment, the cell solution in the cell culture container moves relative to the linear color image sensor, including: the cell solution in the cell culture container moves while the linear color image sensor remains stationary (i.e., a relative motion mode of [stationary sensor + moving solution]), or the linear color image sensor moves while the cell solution in the cell culture container remains stationary (i.e., a relative motion mode of [stationary solution + moving sensor]).

[0035] Compared to sensors such as RGB color sensors that can only collect single-point color data, linear array color image sensors can collect multiple lines of color data and then stitch them together to form a two-dimensional color image. This allows them to collect enough color data of the cell solution to ensure the accuracy of determining the timing of fluid replacement, while also saving more space than area array color image sensors.

[0036] Step S12: Cluster the color values ​​of the color image to obtain the color value clustering results.

[0037] In this step, clustering operations can be used to process the color values ​​(i.e., pixel values) of a color image, thereby classifying similar color values ​​in the color image.

[0038] In an optional implementation, clustering can be performed using a clustering algorithm. For example, the K-Means clustering algorithm can be used. The K-Means algorithm first divides the dataset (color values ​​of a color image in this embodiment) into K clusters, where K is a preset hyperparameter. In this embodiment, K can optionally be set to an integer within the range [4,6], such as 4 or 5. Then, K data points are randomly selected from the dataset as the initial K cluster centers. The distance (usually Euclidean distance) between each data point and each cluster center is calculated, and the data is assigned to the nearest cluster center. The cluster center mean is recalculated iteratively until the termination conditions are met. The termination conditions include no (or a minimum number) data points being reassigned, the cluster centers no longer changing, and the intra-cluster squared error being minimized to a local minimum.

[0039] For example, the color values ​​mentioned above can be RGB values ​​or RAW (Raw Image Format) data, etc.

[0040] Step S13: Determine the current dominant color value of the cell solution in the cell culture container based on the color value clustering results.

[0041] Color value clustering results can characterize the current hue distribution of the cell solution in the cell culture container. In this step, the dominant hue color value of the cell solution in the cell culture container can be determined based on the color value clustering results.

[0042] Following the previous example, assuming K is 5, the color value clustering result contains 5 clusters, and each cluster contains the color values ​​belonging to that cluster.

[0043] In an alternative implementation, step S13 can be achieved by the following steps S131 to S132.

[0044] Step S131: Determine the target category with the most color values ​​among all categories (clusters) obtained from the color value clustering results.

[0045] For example, assuming K is 5, and the number of color values ​​contained in the 5 clusters are 12, 100, 50, 2, and 5 respectively, then the cluster corresponding to 100 is the target category.

[0046] Step S132: Determine the current dominant hue color value of the cell solution in the cell culture container based on the cluster center (centroid) of the target category.

[0047] In this embodiment, the current dominant hue color value of the cell solution in the cell culture container can be characterized by the centroid of the cluster with the largest data proportion.

[0048] If at least two clusters have the same number of color values ​​as the first: In one optional implementation, the cell solution can be shaken, and then the color acquisition command can be triggered again to reacquire the color image. Color value clustering can be performed again until a unique target category is found. Then, the cluster center of the target category is determined as the current dominant hue color value of the cell solution in the cell culture container. In another optional implementation, the centroids of all clusters that are tied for first place can be weighted and summed (using normalized weights) or weighted and averaged (using unnormalized weights) to obtain the current dominant hue color value of the cell solution in the cell culture container. For example, the weight corresponding to the centroid of each cluster participating in the weighted summation or weighted average can be the proportion of the number of color values ​​contained in that cluster to the total number of color values ​​contained in all clusters that are tied for first place.

[0049] In an optional implementation, step S132 can be achieved by the following steps: The cluster centers of the target category are converted to the HSV (Hue-Saturation-Value) color space to obtain the HSV color values ​​corresponding to the cluster centers of the target category. The HSV color value corresponding to the cluster center of the target category is determined as the current dominant color value of the cell solution in the cell culture container.

[0050] In this embodiment, the color value obtained based on hardware (e.g., a sensor) is typically the sensor's native color space (e.g., directly outputting RAW data), or converted from the sensor's native color space to the RGB (red, green, blue) color space (e.g., directly outputting RGB data).

[0051] The HSV color space is a color representation method based on human intuitive perception of color, which is more consistent with the colors observed by the human eye. Therefore, in this embodiment, the centroid of the cluster with the largest data proportion is first converted to the HSV color space, and then the HSV color value corresponding to the centroid of the cluster with the largest data proportion is determined as the current dominant color value of the cell solution in the cell culture container. In this way, through automated hardware acquisition and color space conversion, the same effect as the color of the cell solution observed by the human eye can be achieved without relying on manual observation.

[0052] Step S14: Based on the primary color value, determine whether culture medium needs to be added to the cell culture container using a pre-trained replenishment prediction model.

[0053] In the embodiments of this application, a fluid replenishment prediction model can be pre-trained and deployed. The fluid replenishment prediction model can be used to determine whether it is necessary to replenish culture medium into the cell culture container based on the primary color value, that is, to determine whether the time for fluid replenishment has been reached based on the primary color value.

[0054] In one optional implementation, the fluid replenishment prediction model may include a cell density prediction model for predicting cell density. Optionally, the cell density prediction model may correspond to the currently cultured cell species. In this embodiment, for example, since different cell species exhibit different color change patterns during culture—for instance, some cell species may have a darker dominant color when reaching the required fluid replenishment density, while others may have a lighter dominant color—a cell density prediction model can be pre-trained for each cell species. This allows for selective application based on the currently cultured cell species during the specific cell culture process.

[0055] Based on the cell density prediction model provided in this embodiment, step S14 can optionally be implemented by the following steps S141 to S143.

[0056] Step S141: Based on the primary color value, determine the current cell density of the cell solution in the cell culture container using a cell density prediction model.

[0057] In this step, the cell density prediction model can output the current cell density of the cell solution in the cell culture container based on the dominant hue color value.

[0058] Step S142: If the cell density exceeds the cell density threshold, determine that culture medium needs to be added to the cell culture container.

[0059] If the cell density exceeds the cell density threshold, it indicates that the cell culture environment has entered a stage of resource shortage and high metabolic pressure. Therefore, it can be determined that the time for replenishment has arrived and culture medium needs to be added to the cell culture container.

[0060] In one example, the cell density threshold can correspond to the cell species currently being cultured. This is because different cell species grow at different rates. For example, cells that metabolize quickly, consume a lot of glucose, and produce acid quickly need to be replenished with fluid at a lower cell density, while cells that metabolize slowly and are more tolerant can tolerate a higher cell density. Therefore, the cell density threshold can be set separately for different cell species.

[0061] In another example, the cell density threshold can correspond to the current culture stage of the cell culture. That is, different cell density thresholds can be set for different cell culture stages. This is because the cell growth and reproduction rates differ at different culture stages. For example, in the logarithmic growth phase, the cell number increases logarithmically. Therefore, during the logarithmic growth phase, the cell density threshold can be sequentially increased for multiple logarithmic growth stages to prolong the logarithmic growth trend as much as possible, allowing the cells to quickly reach the required number. Optionally, the duration of each culture stage can be pre-set based on experience, such as 2 days for the first culture stage, 3 days for the second, and so on. Then, the current culture stage can be determined. For example, if the cell has been cultured to day 3, it can be determined that the cell is currently in the second culture stage. Alternatively, optionally, based on collected cell growth data (such as cell density, current cell culture duration, dominant hue value, etc.), a pre-trained cell growth stage prediction model can be used to predict the current culture stage of the cell. Then, the corresponding cell density threshold can be determined based on the current culture stage.

[0062] In another example, the aforementioned cell density threshold can correspond simultaneously to both the currently cultured cell type and the current culture stage. A baseline cell density threshold can be set for each cell type (different types may correspond to different baseline cell density thresholds). Based on the currently cultured cell type and the current culture stage, the baseline cell density threshold is increased or decreased to obtain the current cell density threshold. For example, if the initial lag phase is in use, the baseline cell density threshold is used; if the logarithmic phase is in use, the threshold is used as a starting point, and the value of 'a' is increased by 'a' for each logarithmic growth stage. The specific value of 'a' can be determined based on the cell type.

[0063] Step S143: If the cell density does not exceed the cell density threshold, it is determined that there is no need to add culture medium to the cell culture container at present.

[0064] If the cell density does not exceed the cell density threshold, it indicates that the cell culture environment is currently in a stage where resources are sufficient and metabolic pressure is adequate. Therefore, it can be determined that the time for replenishing fluid has not yet been reached, and there is no need to replenish the culture medium in the cell culture container.

[0065] Cell density is a crucial parameter in cell culture, directly impacting cell growth, functional expression, experimental results reliability, and final yield. It serves as a core parameter connecting cell biological characteristics with culture process control. This embodiment utilizes a cell density prediction model to directly predict the current cell density of the cell solution in the cell culture container. This allows for obtaining this vital parameter without opening the bag or installing additional cell density detection devices. Furthermore, by comparing the current cell density with the cell density threshold corresponding to the currently cultured cell type, it's possible to determine whether replenishment is timely. Thus, without additional operations or devices, it achieves both accurate determination of replenishment timing and acquisition of cell density.

[0066] Optionally, step S141 in the above embodiments can be implemented by the following steps: inputting cell solution data, including at least the primary hue color value, into the cell density prediction model, and outputting the current cell density of the cell solution in the cell culture container through the cell density prediction model.

[0067] In this embodiment, the input data for the cell density prediction model is cell solution data, which includes at least the dominant hue color value of the cell solution. That is, the dominant hue color value of the cell solution is the core basis for predicting cell density. Optionally, more cell solution-related input data can be added to the cell density prediction model to improve the accuracy of cell density prediction. For example, the cell solution data may also include the current cell solution volume and / or the current cell culture duration.

[0068] In an optional implementation, the cell density prediction model includes a Bayesian ridge regression model, which naturally supports incrementality, interpretability, and uncertainty.

[0069] Cell density prediction models can be obtained through the following training process, including: Obtain the sample set required for training. Each sample in the sample set includes cell solution data and corresponding cell density label data. The cell density label data is also known as the label. The cell solution data may include the dominant hue color value of the cell solution at time t (which may be the HSV value). Optionally, it may also include the cell solution volume and / or cell culture time corresponding to time t. Optionally, the cell density label data may be the actual cell density determined by sampling using a flow cytometer. Sample preprocessing is used to clean the sample set and remove duplicate or erroneous samples. The sample set is divided into a training set and a test set. Optionally, the sample set can be divided into a training set and a test set in a 7:3 ratio. The initial cell density prediction model is trained using a training set. The training objective is to maximize the posterior estimate. After each prediction result is output by the cell density prediction model, the standard deviation of the prediction is determined. If the standard deviation exceeds the first standard deviation threshold (e.g., 0.5), it indicates that the prediction result of the cell density prediction model is unreliable and needs to be trained and learned again. After training, the cell density prediction model is tested using a test set. If the test results indicate that the prediction performance of the cell density prediction model is not good, training can continue. When continuing training, samples with small prediction standard deviations (e.g., below the second standard deviation threshold) can be selected for learning. For example, 3-5 samples with small prediction standard deviations can be selected because samples with small prediction standard deviations are samples that the cell density prediction model can predict relatively accurately. These samples have a more significant effect on improving model performance. Using such samples for incremental model training can improve the model training efficiency.

[0070] The aforementioned cell growth stage prediction model and the subsequently mentioned binary classification model can also be trained in a similar manner. It is important to note that when training the cell growth stage prediction model, each sample in its sample set includes cell growth data and corresponding culture stage annotation data. Similarly, when training the binary classification model, each sample in its sample set includes cell growth data and corresponding classification result annotation data.

[0071] In another alternative implementation, the fluid replenishment prediction model may include a fluid replenishment demand classification model. Accordingly, step S14 may determine the classification result of whether culture medium needs to be replenished into the cell culture container based on the primary color value and the fluid replenishment demand classification model.

[0072] In this embodiment, the fluid replenishment requirement classification model can be, for example, a binary classification model. The output space of the fluid replenishment requirement classification model can include two categories: one category represents the category requiring fluid replenishment when culture medium needs to be added to the cell culture vessel, and the other category represents the category not requiring fluid replenishment when culture medium does not need to be added to the cell culture vessel. The input data for the fluid replenishment requirement classification model is cell solution data, which includes at least the dominant hue color value of the cell solution. Optionally, the cell solution data may also include the current cell solution volume and / or the current cell culture duration. In this embodiment, cell solution data, including at least the dominant hue color value, can be input into the fluid replenishment requirement classification model, and the model outputs either a category requiring fluid replenishment or a category not requiring fluid replenishment.

[0073] Step S15: If it is determined that culture medium needs to be added to the cell culture container, then the culture medium is added to the cell culture container.

[0074] If the replenishment prediction model determines that culture medium needs to be added to the cell culture vessel, i.e., that the replenishment time has been determined, then the culture medium can be added to the cell culture vessel, thereby achieving replenishment at the accurate time. The amount of culture medium replenished each time can be preset based on the cell growth cycle or calculated using other methods; this application does not specifically limit this.

[0075] If it is determined that there is no need to replenish the culture medium in the cell culture vessel at present, no action is required.

[0076] In this embodiment of the application, during cell culture using a transparent cell culture container, a color image of the cell culture container is first acquired in response to a color acquisition command, thereby obtaining the color data of the cell solution in the transparent cell culture container. Then, the color values ​​of the acquired color image are clustered to identify the hue distribution characteristics of the cell solution. Subsequently, based on the color value clustering results, the current dominant hue value of the cell solution can be determined. The relationship between the dominant hue value of the cell solution and the replenishment requirement is analyzed using a replenishment prediction model to determine whether replenishment is currently needed. If it is determined that replenishment is currently needed, the culture medium is added to the cell culture container to achieve cell replenishment. This application embodiment monitors the color changes of cell solutions using color image data of cell solutions that can accurately represent color. It accurately anchors the current dominant color of the cell solution through clustering of image color values, eliminating the influence of local color in the cell solution. It also uses a fluid replenishment prediction model to clarify the fluid replenishment requirement corresponding to the current dominant color of the cell solution. Compared with traditional methods, color clustering based on cell solution color images improves the accuracy of determining the dominant color of the cell solution. By associating the dominant color of the cell solution with the fluid replenishment requirement through the fluid replenishment prediction model, it can determine the appropriate time for fluid replenishment based on the changes in the dominant color during the specific growth process of cells, thereby improving the accuracy of determining the timing of fluid replenishment and avoiding missing the optimal time for fluid replenishment.

[0077] Compared to prior art 1, the embodiments of this application can accurately determine the timing of fluid replenishment and also automate the process, thus enabling its application in industrial cell culture. Compared to prior art 2, the embodiments of this application can shorten the overall culture time for fast-growing cells and increase the probability of successful culture for slow-growing cells.

[0078] In addition, the inventors also conceived of an implementation method for accurately determining the timing of fluid replacement, namely, detecting cell density using a density sensor and then determining whether fluid replacement is needed based on a comparison between the cell density and a cell density threshold. However, density sensors can only perform localized measurements of the cell solution. Therefore, cell homogeneity greatly affects the accuracy of the density detection results. Since cells are difficult to homogenize, especially with some cell clusters, the density detection results can have significant deviations. In contrast, the embodiments of this application can determine whether fluid replacement is needed based on the color characteristics of the cell solution. The color of the cell solution is determined by pH, which is unrelated to cell homogeneity. As cells proliferate, they release lactic acid, causing changes in the solution's pH. Therefore, compared to the aforementioned fluid replacement method based on density sensors, the embodiments of this application can avoid the problem of inaccurate determination of fluid replacement timing due to insufficient cell homogeneity.

[0079] In an optional implementation, this application embodiment may also provide an abnormal alarm mechanism, as follows: when the current cell culture time exceeds the preset culture time, if it is not determined from the start time of cell culture to the current time that culture medium needs to be added to the cell culture container, an abnormal cell growth alarm will be issued.

[0080] During cell culture, if cells grow normally, at least one instance of fluid replenishment should be identified within a set timeframe (e.g., 3 days). However, if no fluid replenishment is identified within this timeframe, it indicates a problem in the cell growth process. Therefore, an abnormal alarm can be issued based on the cell growth status. Optionally, the cell growth abnormality alarm can be triggered in one or more of the following ways: displaying a cell growth abnormality alarm message, outputting a cell growth abnormality alarm audio, or outputting a cell growth abnormality alarm light signal. For example, the cell growth abnormality alarm audio may include a first voice audio, a first alarm tone audio, etc. The cell growth abnormality alarm light signal may, for example, be a light signal of a certain wavelength (e.g., blue light), and may be emitted via an LED indicator.

[0081] After an alarm is triggered indicating abnormal cell growth, the operator can troubleshoot the problem. If the troubleshooting reveals that the alarm was false, the alarm can be deactivated and the cell culture process can continue. If it is determined that the culture cannot continue, the cell culture program can be terminated, thus avoiding waste.

[0082] In this embodiment, by means of a detection mechanism to determine whether fluid replenishment is needed, cell growth can be indirectly monitored by checking whether there has been a situation within a certain time limit in which fluid replenishment is needed. This eliminates the need for additional monitoring devices to monitor cell growth and reduces the complexity of cell culture equipment.

[0083] Figure 2This is a structural block diagram of an integrated cell culture device provided in an embodiment of this application. Figure 3 A structural block diagram of another integrated cell culture device provided in the embodiments of this application is shown below. Figure 2 and Figure 3 As shown, the integrated cell culture device includes a culture medium tank 10, a temperature pretreatment module 20, a display and control module 30, a cell culture chamber 40, a valve system 50, and a fluid power system 60.

[0084] The culture medium tank 10 is used to store culture media. It has space for placing culture medium containers, allowing the containers to be filled with the media before being placed inside for storage. The culture medium tank 10 provides the necessary temperature for storing the media (culture media typically require low-temperature storage, the specific temperature depending on the type of media), preventing media deterioration. The culture media used in cell culture can include at least two types: activation media and amplification media. Activation media can be used to awaken dormant, injured, or low-temperature stressed cells, restoring their normal metabolic and physiological activity. Amplification media can be used for rapid cell proliferation, obtaining a large number of highly viable cells in a short time. The type of culture medium used in cell culture depends on the cell type and culture requirements.

[0085] Optionally, the culture medium tank 10 may be equipped with a weighing module (e.g., Figure 2 or Figure 3 The weighing module 1 is used to weigh the culture medium. Optionally, if cell culture requires multiple culture media (e.g., activation medium and amplification medium) and / or a certain culture medium needs to be produced in multiple portions (e.g., amplification medium requires two or three bags), for example, refer to... Figure 3 In the culture medium containers 1, 2, 3 and 4, one of the culture medium containers can be used to hold activation culture medium, and the other three culture medium containers can be used to hold amplification culture medium. The weighing module in the culture medium box 10 can weigh the total weight of all the culture medium, or it can weigh each culture medium separately.

[0086] The temperature pretreatment module 20 is used to pretreat the culture medium (typically including preheating) to eliminate the temperature difference between the culture medium and the cells, thus eliminating the temperature adaptation phase for cell growth. It should be noted that the temperature pretreatment module 20 is an optional module in the integrated cell culture device. It can be replaced by other equivalent modules or other means. For example, the temperature of the culture medium can be controlled externally by adjusting the room temperature during dispensing, or the temperature can be allowed to rise after dispensing before being used for cell culture. The temperature pretreatment method for the culture medium is not limited to these methods. For example, the temperature pretreatment module 20 can be a heating plate.

[0087] The cell culture incubator 40 includes a coating device 41 and a culture device 42.

[0088] Figure 4 A schematic diagram of the physical structure of a cell culture chamber according to an embodiment of this application is provided. The coating device 41 is provided with a placement space 411 for a cell source container, a placement space 412 for a coating container, and a placement space 413 for a waste liquid container. This application does not limit the specific locations of the placement spaces for the cell source container, the coating container, and the waste liquid container. Figure 4 The image only illustrates one possible placement layout. The coating device 41 can provide placement space for the cell source container, coating container, and waste liquid container via a coating platform. The coating platform is equipped with a control motor for swinging, vibrating, or tilting the liquid inside the coating container. The cell source container stores the initial cell source required for cell culture, the coating container performs the cell coating process, and the waste liquid container stores waste liquid, such as culture medium liquid used for cleaning the container.

[0089] The culture apparatus 42 is provided with one or more cell culture container placement spaces, and one of the cell culture container placement spaces in the culture apparatus 42 can be used to place one cell culture container. At least one of the one or more cell culture container placement spaces (e.g. Figure 2 or Figure 3 The cell culture container placement space provided by the culture platform 1 is equipped with a matching color image sensor and a reflective component, with the reflective component located in the light source emission direction of the matching color image sensor.

[0090] The culture device 42 can provide space for cell culture containers via a culture platform. Optionally, one culture platform can provide space for one cell culture container. A control motor can be configured for each culture platform to rock, vibrate, or tilt the liquid inside the cell culture container. Optionally, all culture platforms can be equipped with one control motor, which can control all culture platforms to rock, vibrate, or tilt simultaneously. Alternatively, each culture platform can be equipped with its own control motor, which can individually control the corresponding culture platform to rock, vibrate, or tilt. For example, as shown... Figure 3 and Figure 4 As shown, the culture apparatus 42 may include three culture platforms, each providing space for a cell culture container. Optionally, at least one culture platform is equipped with a weighing module for weighing the cell culture containers placed on the corresponding platform. For example... Figure 2 The cultivation platform 1 is equipped with a weighing module 2; for example, Figure 3On the culture platforms 1, 2, and 3, there are respectively weighing modules 2, 3, and 4; for another example, it is also possible that only the culture platform 1 has the weighing module 2, and the culture platforms 2 and 3 do not have weighing modules.

[0091] Optionally, the hardware configurations corresponding to each culture platform can be the same or different. For example, each culture platform can be correspondingly provided with a weighing module and equipped with a corresponding peristaltic pump and pinch valve, or it can be that only one culture platform has a weighing module and is equipped with a peristaltic pump, while other culture platforms do not have a weighing module and are not equipped with a peristaltic pump. The embodiments of the present application do not limit this.

[0092] Figure 5 It is a schematic diagram of the physical assembly of the culture platform in the cell culture integrated device, as Figure 4 shown, the culture platform can include a bottom plate 4201 and a protective cover 4202 that are配套 to the cell culture container. When the protective cover 4202 is assembled to the配套 bottom plate 4201, a placement space for the cell culture container can be formed. Since each time cell culture is carried out, the cell culture container needs to be placed on the bottom plate 4201 and the protective cover 4202 is assembled, and after the cell culture is completed, the cell culture container needs to be taken out. Therefore, the assembly method of the protective cover 4202 and the bottom plate 4201 needs to be a repeatedly detachable and installable assembly method, so as to facilitate the taking and placing of the cell culture container. Optionally, the assembly method of the protective cover 4202 and the bottom plate 4201 includes one or more of the following methods: snap fit, magnetic attraction, plug connection, and slide rail connection, etc. It can be understood that the assembly method of the protective cover 4202 and the bottom plate 4201 is not limited to the above example methods.

[0093] As Figure 5 shown, a color image sensor 4201a can be provided on the bottom plate 4201. Figure 6 It is a schematic diagram of the protective cover of the cell culture container in a cell culture integrated device, as Figure 6 shown, a reflective component 4202a can be provided on the protective cover 4202 that is配套 to the bottom plate 4201. Among them, the color image sensor 4201a can be used to obtain a color image of the cell culture container. The principle is to emit light to the cell culture container and then receive the reflected light signal. The reflective component 4202a can be used to reflect the light passing through the transparent cell culture container onto the sensing unit of the color image sensor 4201a, so that the reflected light signal can be received by the color image sensor 4201a. Among them, the light reflected by the reflective component 4202a will pass through the transparent cell culture container again and then be received by the color image sensor 4201a.

[0094] Optionally, as Figure 5As shown, the base plate 4201 can be provided with multiple first vent holes 4201b, and the protective cover 4202 can be provided with multiple second vent holes 4202b to ensure the air permeability requirements of cells during culture. The area of ​​the second vent holes can be smaller than the area of ​​the first vent holes, allowing the base plate 4201 to have better support for the cell culture container.

[0095] The valve system 50 includes multiple valve devices. The location of the multiple valve devices depends on the pipeline path to be controlled. They are not limited to the inside of the culture medium tank 10, the temperature pretreatment module 20, or the cell culture tank 40. They can also be located outside the culture medium tank 10, the temperature pretreatment module 20, or the cell culture tank 40, as long as they are installed on the integrated cell culture equipment.

[0096] Valve system 50 includes a plurality of valve devices, each valve device including one or more valves, optionally, the valves being pinch valves, the plurality of valve devices including a first valve device for controlling the piping between the culture medium container and the coating container (e.g. Figure 2 Pinch valve 1 in the middle, or Figure 3 Pinch valve 1, pinch valve 7, and pinch valve 8; and a second valve device for controlling the tubing between the cell source container and the coating container (e.g., ...). Figure 2 or Figure 3 The clamp valve 2), and a third valve device for controlling the tubing between the coating container and the cell culture container (e.g., the clamp valve 2), and a third valve device for controlling the tubing between the coating container and the cell culture container. Figure 2 Pinch valves 3 and 4 in the middle, or Figure 3 (Pinch valves 3, 4, 5, and 6 in the middle).

[0097] It should be noted that the first valve device for controlling the piping (via the temperature pretreatment module 20) between the culture medium container and the coating container can control the outflow of different types of culture media through different valves, and can also control the outflow of the same type of culture media through different or the same valves. For example, as... Figure 3 As shown, culture medium container 1 can store one type of culture medium (e.g., activation culture medium), culture medium container 2 can store another type of culture medium (e.g., an amplification culture medium), and culture medium containers 3 and 4 can store the same third type of culture medium (e.g., another amplification culture medium). The culture medium in culture medium container 1 can be controlled by clamp valve 1, the culture medium in culture medium container 2 can be controlled by clamp valve 7, and the same type of culture medium in culture medium containers 3 and 4 can be controlled by clamp valve 8.

[0098] The culture medium container, coating container, cell source container, and cell culture container, when connected by tubing between the culture medium container and the coating container, between the cell source container and the coating container, and between the coating container and the cell culture container, form a closed environment. After being connected under sterile conditions, each tubing and container creates a closed, sterile environment. The valve system 50 controls the on / off state of the tubing between the containers, thereby enabling liquid exchange between them. Each tubing and container can be a disposable consumable.

[0099] The liquid propulsion system 60 includes a plurality of liquid propulsion devices, each of which may include one or more liquid propulsion pumps. Optionally, the liquid propulsion pumps may be peristaltic pumps. The plurality of liquid propulsion devices include a first liquid propulsion device for controlling the liquid flow between the culture medium container and the coating container (e.g., ...). Figure 2 The peristaltic pump 1) and the second hydrodynamic device (e.g., a peristaltic pump) used to control the liquid flow between the waste liquid container and the coated container. Figure 2 The peristaltic pump 2 in the middle, and a third hydrodynamic device (e.g., a peristaltic pump 2) for controlling the liquid flow between the coating container and the cell culture container. Figure 2 The peristaltic pump 3 in the system. The liquid power unit can be used to control the flow direction and velocity of the liquid in the pipeline. It should be noted that the liquid power system 60 is an optional module in the integrated cell culture equipment. The flow direction and velocity of the liquid in the pipeline can be controlled by controlling pipeline vibration, utilizing gravity, etc.

[0100] The display control module 30 includes a control module and a display device. The display device can be used to display culture information and control information during the cell culture process. Culture information includes cell type, culture time, cell density, weighing data (or data that can be converted to volume for display), color images, etc. Control information includes valve on / off status, fluid dynamics device status, the tilt angle and speed of the coating platform, and the tilt angle and speed of the culture platform, etc. Optionally, the display device may include a display screen. The control module is electrically connected to the culture tank 10, temperature pretreatment module 20, cell culture tank 40, valve system 50, fluid dynamics system 60, and display device. The control module is equipped with a pre-trained fluid replenishment prediction model and is used to implement the following steps S21 to S25, including: Step S21: During cell culture using a cell culture container, in response to a color acquisition command, the color image sensor is controlled to acquire a color image of the cell culture container.

[0101] The specific implementation method of this step can be referred to step S11 in the embodiment of the cell rehydration method.

[0102] To ensure that the acquired color images accurately reflect the color of the cell solution in the cell culture container, users can be prompted to use transparent cell culture containers. For example, this prompt can be stated in the instruction manual of the integrated cell culture device, or it can be included in the device operation demonstration video, etc. Additionally, transparent cell culture containers compatible with the integrated cell culture device can be provided to the user.

[0103] In this step, the control module can trigger a color acquisition command, call the color image sensor, and control the color image sensor to acquire a color image.

[0104] Optionally, the color image sensor includes a linear array color image sensor. Correspondingly, the control module can also be used to implement the following steps: controlling the relative movement between the cell solution in the cell culture container and the linear array color image sensor, and controlling the linear array color image sensor to acquire color images of the cell culture container during the relative movement. It should be noted that if there are multiple cell culture containers, and each cell culture container has a corresponding linear array color image sensor in its placement space, then the linear array color image sensor can only acquire color images for its corresponding cell culture container.

[0105] Optionally, the position of the linear color image sensor can be fixed, and the control module can control the cell solution in the cell culture container to move relative to the linear color image sensor, thereby realizing a relative motion mode of "the sensor is stationary and the solution moves".

[0106] In another relative motion mode, where the solution remains stationary while the sensor moves, a complex motion control structure and a relatively large cell culture container are required to enable the linear color image sensor to move. This is because if the linear color image sensor moves, the reflector also needs to move to ensure effective sensor acquisition. However, if the reflector is fixed, the cell culture container needs to be limited to a certain height (e.g., 8 mm) to ensure effective sensor acquisition. Furthermore, since cells require good aeration during culture, and the linear color image sensor and its motion control structure are entities that can block gas transmission, a relatively large cell culture container is also required to increase the aeration area to meet the cell's aeration needs.

[0107] By adopting a relative motion method of "fixed sensor + moving solution", the linear array color image sensor does not need to move and occupies a small area. The movement of the cell solution can be achieved simply by tilting the cell culture container, without the need for additional devices (shaking the cell solution during cell culture is a routine operation, so the tilt control device of the cell culture container is a basic hardware structure that can be directly used to control the cell culture container to tilt). In this way, more space can be freed up on the hardware device used to place the cell culture container (such as the base plate 4201 and protective cover 4202 of the culture platform) to set up vents, thereby ensuring the cell's aeration needs.

[0108] Therefore, the relative motion method of "fixed sensor + moving solution" can simultaneously ensure cell aeration and sensor acquisition effect, without limiting the size of cell culture container, which is extremely beneficial for industrial applications.

[0109] Step S22: Cluster the color values ​​of the color image to obtain the color value clustering results.

[0110] The specific implementation method of this step can be referred to step S12 in the embodiment of the cell rehydration method.

[0111] Step S23: Determine the current dominant color value of the cell solution in the cell culture container based on the color value clustering results.

[0112] The specific implementation method of this step can be referred to step S13 in the embodiment of the cell rehydration method.

[0113] Step S24: Based on the primary color value, determine whether culture medium needs to be added to the cell culture vessel using a replenishment prediction model.

[0114] The specific implementation method of this step can be referred to step S14 in the embodiment of the cell rehydration method.

[0115] In this step, the control module can call the deployed replenishment prediction model and determine whether culture medium needs to be replenished into the cell culture container based on the primary color value.

[0116] Step S25: If it is determined that culture medium needs to be added to the cell culture container, then the first valve device and the third valve device are opened.

[0117] In this step, if the control module determines that liquid replenishment is needed, it can control the first valve device and the third valve device to open, thereby opening the pipeline between the culture medium container and the cell culture container, so that the culture medium can flow from the culture medium container to the cell culture container, thus achieving liquid replenishment.

[0118] Since different types of culture media can be used, when opening the first valve device, only the valve between the culture medium container and the coating container containing the culture medium required for the current culture stage can be opened (e.g., Figure 3 (shown as pinch valve 1, pinch valve 7, or pinch valve 8), instead of all valves in the first valve assembly.

[0119] If there are multiple cell culture containers (e.g.) Figure 3 As shown), and it is determined that cell culture container 1 needs replenishment, then when opening the third valve device, only the valve between the coated container and cell culture container 1 needs to be opened (e.g., Figure 3 Pinch valves 3 and 4 in the middle), instead of opening all valves in the third valve assembly (such as...). Figure 3 Pinch valves 5 and 6 in the middle can be left unopened. (Refer to...) Figure 3 If it is determined that cell culture container 2 requires replenishment of fluid, the valve between the coated container and cell culture container 2 can be opened (e.g., ...). Figure 3 (Pinch valves 3 and 5 in the middle). See reference. Figure 3 If it is determined that cell culture container 3 needs replenishment, the valve between the coated container and cell culture container 3 can be opened (e.g., ...). Figure 3 (Pinch valve 3 and pinch valve 6 in the middle).

[0120] To ensure the accuracy of the data (i.e., color images) acquired by the color image sensor, the integrated cell culture equipment may include at least a light-shielding device for protecting the cell culture incubator 40 from light, such as a light-shielding door or curtain. This light-shielding device should at least protect the cell culture incubator 40 from light, thereby ensuring the data acquisition effect of the color image sensor. Furthermore, since most culture media also require light protection, this light-shielding device can also be used to protect the culture medium tank 10 from light, or a light-shielding device can be provided in the integrated cell culture equipment specifically for protecting the culture medium from light in the culture medium tank 10.

[0121] For example, Figure 7This is a schematic diagram of the physical structure of an integrated cell culture device provided in an embodiment of this application. The integrated cell culture device may include a light-shielding outer shell 70 and a light-shielding door 80. Further optionally, the culture medium tank 10 in the integrated cell culture device may include a transparent culture medium tank door 12 for observing the remaining amount and state of the culture medium and ensuring the ambient temperature of the culture medium. The cell culture chamber 40 in the integrated cell culture device may include a transparent culture chamber door 42 for observing the cell culture situation and ensuring the ambient temperature and gas concentration required for cell growth. Both the culture medium tank 10 and the cell culture chamber 40 can be surrounded by the light-shielding outer shell 70. The light-shielding door 80 can be located outside the culture medium tank door and the culture chamber door, serving as the first door when the entire integrated cell culture device is opened. In this way, during the cell culture process, the light-shielding door can be opened, and the culture medium can be observed through the transparent culture medium tank door 12 without affecting the ambient temperature of the culture medium, and the cell solution can be observed through the transparent culture chamber door 42 without affecting the ambient temperature and gas concentration of the cell environment.

[0122] In this embodiment, the tubing and containers, when connected under sterile conditions, form a closed, sterile cell culture environment. During cell culture using transparent containers, color changes in the cell solution are monitored using color image data of the cell solution, which accurately represents color. Clustering of image color values ​​accurately anchors the current dominant hue of the cell solution, eliminating the influence of localized colors. Furthermore, a rehydration prediction model is used to determine the rehydration requirement corresponding to the current dominant hue of the cell solution. Compared to the prior art 3, which determines rehydration timing by opening the bag and sampling to check cell density, this embodiment, through a cell rehydration method configured in the control module, not only accurately determines the rehydration timing but also enables automatic rehydration in a closed, sterile environment, avoiding the risk of cell contamination caused by opening the bag for sampling. Compared to the prior art 4, the embodiments of this application can achieve closed aseptic environment deployment, automated cell culture, and automated aseptic fluid replenishment through small-scale integrated cell culture equipment. There is no need for additional large-scale facilities such as aseptic operation boxes or aseptic operation rooms, which reduces the difficulty of achieving an aseptic environment and energy consumption. The culture needs of one sample of cells can be completed by one integrated cell culture equipment. Multiple integrated cell culture equipment can simultaneously meet the culture needs of multiple sample cells without mutual contamination. This enables the industrial application of small-scale bacterial culture needs, that is, to provide cell culture services in batches, safely, and at low cost.

[0123] In one optional implementation, this application embodiment can provide a cell growth abnormality alarm mechanism. Specifically, if the current cell culture time exceeds a preset culture time, and no need to replenish culture medium in the cell culture container has been determined between the start of cell culture and the current time, a cell growth abnormality alarm will be issued. For details, please refer to the relevant content in the cell replenishment method embodiment, which will not be repeated here.

[0124] In one optional implementation, the present application embodiment can provide a pipeline abnormality alarm mechanism, which may optionally include one or more of the following mechanisms: pipeline installation abnormality alarm mechanism, pipeline blockage abnormality alarm mechanism, and pipeline leakage abnormality alarm mechanism.

[0125] The tubing installation anomaly alarm mechanism uses data collected by photoelectric sensors installed inside the cell culture integrated equipment to monitor tubing position and determine whether the tubing is installed correctly. If the signal received at a certain location is not the signal that a correctly installed tubing should have, an alarm can be triggered. Optionally, the alarm can be triggered in one or more of the following ways: displaying an alarm message, outputting an audio alarm, or outputting a light alarm. For example, the audio alarm may include a second voice message, a second alarm tone, etc. The light alarm may be a light signal of a certain wavelength (e.g., red light), which can be emitted by an LED (Light Emitting Diode) indicator.

[0126] The pipeline blockage alarm mechanism determines whether a pipeline is blocked by monitoring changes in the weighing data of the containers. If the weighing data of a container remains unchanged or changes very little within a set period, a pipeline blockage alarm can be triggered. For example, if, within a certain time limit after the start of fluid replenishment, the weighing data of the culture medium and / or cell solution remains unchanged or changes very little (e.g., the weight change is below a preset weight threshold), it can be determined that the pipeline between the culture medium container and the cell culture container is blocked. Optionally, the pipeline blockage alarm can be triggered in one or more of the following ways: displaying a pipeline blockage alarm message, outputting a pipeline blockage alarm audio, or outputting a pipeline blockage alarm light signal. For example, the pipeline blockage alarm audio may include a third voice audio, a third alarm tone audio, etc. The output pipeline blockage alarm light signal can be, for example, a light signal of a certain wavelength (e.g., a green light signal), which can be emitted through an LED indicator.

[0127] The pipeline leakage alarm mechanism can determine whether there is a leak by monitoring changes in the weighing data of the containers. If the difference between the decrease in the weight of the culture medium in the culture medium container and the increase in the weight of the cell solution in the cell culture container exceeds a preset value, a pipeline leakage alarm can be triggered. Optionally, the pipeline leakage alarm can be triggered in one or more of the following ways: displaying a pipeline leakage alarm message, outputting a pipeline leakage alarm audio, or outputting a pipeline leakage alarm light signal. For example, the pipeline leakage alarm audio may include a fourth voice audio, a fourth alarm tone audio, etc. The output pipeline leakage alarm light signal can be, for example, a light signal of a certain wavelength (e.g., a yellow light signal), which can be emitted through an LED indicator.

[0128] In an optional implementation, this application embodiment can provide a container abnormality alarm mechanism. This mechanism can determine whether the container is correctly placed through weighing and / or position detection. If the weighing is too high or too low, or if the container is detected to be misaligned, a container abnormality alarm can be triggered. Optionally, the container abnormality alarm can be triggered in one or more of the following ways: displaying a container abnormality alarm message, outputting a container abnormality alarm audio, or outputting a container abnormality alarm light signal. For example, the container abnormality alarm audio may include a fifth voice audio, a fifth alarm tone audio, etc. The container abnormality alarm light signal can be, for example, a light signal of a certain wavelength (e.g., a violet light signal), and can be emitted through an LED indicator.

[0129] In one optional implementation, the present application embodiments may provide an environmental anomaly alarm mechanism, which may optionally include one or more of the following mechanisms: an alarm mechanism for abnormal ambient temperature in a culture medium chamber, an alarm mechanism for abnormal ambient temperature in a cell culture chamber, and an alarm mechanism for abnormal gas concentration in a cell culture chamber.

[0130] The culture medium incubator ambient temperature abnormality alarm mechanism can determine whether the ambient temperature of the culture medium incubator is abnormal by collecting temperature data from a first temperature sensor installed in the incubator. If the temperature data exceeds a first preset temperature range, the ambient temperature abnormality alarm mechanism is activated. Optionally, the alarm notification method can include one or more of the following: displaying an alarm notification message, outputting an audio alarm notification, or outputting an optical alarm notification. For example, the alarm notification sound can include a sixth voice audio signal or a sixth alarm tone audio signal.

[0131] The cell culture incubator environmental temperature abnormality alarm mechanism can determine whether the cell culture incubator's environmental temperature is abnormal by collecting temperature data from a second temperature sensor installed in the cell culture incubator. If the cell culture incubator temperature data exceeds a second preset temperature range, an environmental temperature abnormality alarm mechanism is activated. Optionally, the alarm notification method can include one or more of the following: displaying an environmental temperature abnormality alarm message, outputting an audio alarm notification, or outputting an optical alarm notification. For example, the environmental temperature abnormality alarm sound can include a seventh voice audio signal or a seventh alarm tone audio signal.

[0132] The cell culture incubator gas concentration abnormality alarm mechanism can determine whether the gas concentration in the cell culture incubator is abnormal by collecting gas concentration data from a gas concentration sensor (e.g., a carbon dioxide concentration sensor) installed in the cell culture incubator. If the gas concentration data exceeds a preset gas concentration range, the cell culture incubator gas concentration abnormality alarm mechanism is activated. Optionally, the alarm notification method can include one or more of the following: displaying a cell culture incubator gas concentration abnormality alarm message, outputting a cell culture incubator gas concentration abnormality alarm audio, or outputting a cell culture incubator gas concentration abnormality alarm light signal. For example, the cell culture incubator gas concentration abnormality alarm audio can include an eighth voice audio, an eighth alarm audio, etc.

[0133] In one optional implementation, this application embodiment can provide an incubator door opening alarm mechanism. This mechanism can be achieved by installing proximity sensors at locations such as the handle and door frame of the incubator door. The proximity sensors collect proximity signals to determine whether the incubator door is open. If a proximity signal is detected at the incubator door during the entire culture process, it is determined that the incubator door is open, and an alarm is triggered. Optionally, the incubator door opening alarm notification can include one or more of the following methods: displaying an alarm message, outputting an audio message, or outputting a light signal. For example, the alarm sound may include a ninth voice audio signal, a ninth alarm tone audio signal, etc.

[0134] The aforementioned types of optical signals can be emitted by LEDs and distinguished by different colors and / or flashing frequencies.

[0135] Figure 8 This is a schematic diagram of a cell culture system provided in an embodiment of this application. The following will be based on... Figure 3 and Figure 8This example provides a concrete example of the process from tubing assembly and installation to cell culture replenishment, designed to achieve a culture target of harvesting more than 6 L of immune cells. Figure 3 Weighing modules 3 and 4 can be disabled or not set up. Accordingly, each cell culture vessel needs to be replenished with liquid in stages, not simultaneously. It should be noted that some terms used in the following examples may differ from those used in the preceding text. However, the relationships and differences between the two are easily understood by those skilled in the art. For example, a coating bag is an optional specific type of coating container, and a CIS sensor is an optional specific type of linear color image sensor, etc.

[0136] Before cell culture, a culture environment needs to be set up, as follows: First, the containers and tubing are assembled to create a closed, sterile environment. This can be done under a laminar flow hood in the laboratory. Primary immune cells are added to the cell bag, coating solution is added to the coating bag, and culture medium is added to the culture medium bag, such as... Figure 8 As shown, the culture medium includes activation medium and amplification medium. There is one bag of activation medium, controlled by clamp valve 7. There are three bags of amplification medium, one controlled by clamp valve 1 and the other two by clamp valve 8. The culture medium bags, coating bags, waste liquid bags, cell bags, and culture bags are then connected via tubing (usually flexible tubing). The culture medium bags, cell bags, and waste liquid bags are all connected to the coating bag, which in turn is connected to the culture bags. The culture bags are made of transparent FEP material.

[0137] Then, the entire connected closed piping system is installed into the integrated cell culture device. Culture medium bags are placed on weighing module 1 in culture medium tank 10, and the total weight of all culture media is measured. The actual amount of culture media reduced is used to determine the actual replenishment volume. The temperature in culture medium tank 10 can be controlled between 2 and 8 degrees Celsius. Clamp valves 1, 7, and 8 can be inserted into the tubing of each culture medium bag to control which culture medium is added. The culture medium tubing exits culture medium tank 10 and sequentially passes through temperature pretreatment module 20 (e.g., ...). Figure 8The culture medium (shown as a heating plate) and peristaltic pump 1 enter the cell culture chamber 40. The temperature pretreatment module 20 can heat the culture medium from 2-8 degrees Celsius to about 37 degrees Celsius, and the peristaltic pump 1 provides power for the outflow of the culture medium. The cell culture chamber 40 provides cells with a suitable temperature and carbon dioxide concentration for growth. The temperature and carbon dioxide concentration can be controlled by the control module in the display control module 30. Since the culture bag is made of FEP material with high air permeability but good water resistance, humidity control is not required inside the cell culture chamber 40. The coating bag is placed on the coating platform in the coating device 41. The coating platform can include a coating base plate and a coating protective cover that can be interlocked to fix and protect the coating bag. After the coating bag is placed on the coating base plate, the coating protective cover can be fastened on. The cell bag can be hung on the bracket of the coating platform. The cell bag tubing is clamped into the clamp valve 2 to control the flow of primary cells from the cell bag into the coating bag. The waste liquid bag is clamped into the peristaltic pump 2 to pump the waste liquid in the coating bag into the waste liquid bag. The coating platform has swaying and vibration functions to reduce cell residue inside the coating bag and maximize cell entry into the culture bag. A clamp valve 3 is inserted into the outlet of the coating bag tubing to control the outflow of liquid from the coating bag. Each culture bag is placed on the culture bag base plate of each culture platform, one-to-one. The culture bag holes are secured with fixing posts on the culture bag base plate. Then, a culture bag protective cover is placed on top of the culture bag and fixed by pins inserted into the corresponding holes on the culture bag base plate. After the protective cover is in place, the reflector on the protective cover is directly above the CIS sensor on the culture bag base plate at a fixed distance. Each culture bag tubing is clamped into the corresponding clamp valve among clamp valves 4-6 to control which culture bag the liquid from the coating bag flows into. Culture platform 1 is equipped with a weighing module 2 to determine whether the liquid in culture bag 1 has increased or decreased in weight. The tubing of culture bag 1 passes through a peristaltic pump 3 to provide power for the inflow or outflow of liquid from culture bag 1. Cells can be cultured in culture bag 1 first, and then transferred to culture bags 2 and 3 after reaching a certain volume. Therefore, cell transfer can be achieved simply by setting a peristaltic pump for culture platform 1.

[0138] At this point, the cultivation environment has been set up.

[0139] After the culture environment is set up, cell culture can begin. The specific process is as follows: Step 1: Control the peristaltic pump 2 to rotate forward, so as to control the transfer of waste liquid in the coated bag into the waste liquid bag; Step 2: Control the opening of the clamp valve 7 corresponding to the culture medium bag 2 (activation culture medium), and control the peristaltic pump 1 to rotate forward to add the activation culture medium into the coating bag, thereby cleaning the coating bag; Step 3: Control the peristaltic pump 2 to rotate forward so that the liquid in the coated bag is completely pumped into the waste liquid bag; Step 4: Control the peristaltic pump 2 to reverse so as to add a small amount of gas into the coated bag to avoid a negative pressure environment inside; Step 5: Control the opening of clamp valve 2, control the vibration of the coating platform, and simultaneously control the coating platform to tilt to one side so that the cells in the cell bag can flow fully into the coating bag; Step 6: Allow the cells to grow in the coated bag for 3 days; Step 7: Control the opening of clamp valve 7 and control the peristaltic pump 1 to rotate forward to add 30 mL of activation medium from culture medium bag 2 to the coating bag; Step 8: Allow the cells to continue growing inside the coated bag for 2 days; Step 9: Control the tilt of the coating platform, control the opening of clamp valve 3 and clamp valve 4, and control the peristaltic pump 3 to reverse so as to transfer the cell solution in the coating bag into the culture bag 1; Step 10: Control the opening of the clamp valve 7 and control the peristaltic pump 1 to rotate forward to add 60 mL of activation medium from the culture medium bag 2 to the coating bag; Step 11: Control the opening of clamp valve 3 and clamp valve 4, and control the peristaltic pump 3 to reverse so as to add 60mL of activation culture medium from the coated bag to culture bag 1. At this time, the volume of cell solution in culture bag 1 is 120mL. Step 12: Control the tilt of culture platform 1 to ensure that the cells in culture bag 1 are in full contact with the culture medium, so that the cells can grow at the appropriate liquid volume; Step 13: Control the opening of the clamp valve 1 corresponding to the culture medium bag 1 (amplification culture medium), and control the peristaltic pump 1 to rotate forward to add 20mL of amplification culture medium to the coating bag, so that the cells remaining in the coating bag can continue to grow. When replenishing the medium later, the culture medium can flush the cells remaining in the coating bag into the culture bag through the coating bag, so as to make full use of the cells. Step 14: Maintain the current static culture state of cells in culture bag 1 for a maximum of 3 days. During this period, trigger a color acquisition command every 2 hours to control the CIS sensor of culture platform 1 to acquire a color image of the cell solution in culture bag 1. After each color image is acquired, cluster the color values ​​of the color image to obtain the color value clustering result. Based on the color value clustering result, determine the current primary color value of the cell solution in culture bag 1. Based on the primary color value, determine the current cell density of the cell solution in culture bag 1 through a pre-selected cell density prediction model corresponding to the currently cultured cell type. If the current cell density exceeds the cell density threshold corresponding to the currently cultured cell type, it is determined that fluid replenishment is required, and then proceed to the next step to replenish fluid. If the current cell density does not exceed the above cell density threshold, it is determined that fluid replenishment is not required, and no fluid replenishment operation is performed. Wait for the next color acquisition command. Step 15: Once it is determined in the previous step that replenishment is needed, call the preset replenishment volume parameter for the current culture stage, such as 180mL, and control the opening of clamp valve 1, clamp valve 3, and clamp valve 4, control peristaltic pump 1 to rotate forward and peristaltic pump 3 to rotate in reverse, so as to add 180mL of amplification culture medium from culture medium bag 1 to culture bag 1. It should be noted that the culture medium needs to pass through the coating bag from the culture medium bag to the culture bag. Step 16: After the fluid replenishment is completed, control culture platform 1 (or control all culture platforms) to shake for 30 seconds to mix the cell solution in culture bag 1. Step 17: The cells continue to grow in culture bag 1. Continue to monitor cell growth, and use the same control logic as in step 14. Step 18: During the current static cell culture in culture bag 1, it is determined that replenishment of medium is needed. Following the control logic of step 14, add 400 mL of amplification medium from culture bag 1 to culture bag 1. At this time, the volume of cell solution in culture bag 1 is 700 mL. Step 19: Control culture platform 1 (or all culture platforms) to shake for 30 seconds to mix the cell solution in culture bag 1; Step 20: The cells continue to grow in culture bag 1. Continue to monitor cell growth, and use the same control logic as in step 14. Step 21: During the current static cell culture in culture bag 1, it is determined that replenishment of medium is needed. Following the control logic in step 14, add 800 mL of amplification medium from culture bag 1 to culture bag 1. At this time, the total volume of cell solution in culture bag 1 is 1500 mL. Step 22: Control culture platform 1 (or all culture platforms) to shake for 30 seconds to mix the cell solution in culture bag 1; Step 23: At this time, the growth space in culture bag 1 is insufficient, so it can be transferred to other culture bags in batches. Control the opening of clamp valve 4 and clamp valve 5, control the peristaltic pump 3 to rotate forward, and transfer the cell solution in culture bag 1 to culture bag 2, transferring 575mL. Step 24: Control culture platform 1 and culture platform 2 (or all culture platforms) to shake for 2 minutes to mix the cell solutions in culture bag 1 and culture bag 2; Step 25: Control the opening of clamp valve 4 and clamp valve 6, control the peristaltic pump 3 to rotate forward, and transfer the cell solution in culture bag 1 into culture bag 3, transferring 575mL; Step 26: Control culture platform 1 and culture platform 3 (or all culture platforms) to shake for 2 minutes to mix the cell solutions in culture bag 1 and culture bag 3. At this time, the volume of cell solution in culture bag 1 is 350mL, the volume of cell solution in culture bag 2 is 575mL, and the volume of cell solution in culture bag 3 is 575mL. Step 27: Shake all culture platforms for 2 minutes to mix the cell solutions in each culture bag; Step 28: Control the opening of clamp valve 1, clamp valve 3 and clamp valve 6, control the peristaltic pump 1 to rotate forward, and add 450mL of amplification culture medium from culture bag 2 to culture bag 3. At this time, the volume of cell solution in culture bag 3 is 1025mL. Step 29: Control the opening of clamp valve 1, clamp valve 3 and clamp valve 5, control the peristaltic pump 1 to rotate forward, and add 140mL of amplification medium from culture medium bag 2 to culture bag 2. At this time, the amplification medium in culture medium bag 1 has been used up, and it is necessary to add amplification medium from other culture medium bags to culture bag 2. Step 30: Control the opening of clamp valves 8, 3 and 5, control the peristaltic pump 1 to rotate forward, and add 300mL of amplification culture medium from culture bag 3 and culture bag 4 to culture bag 2. At this time, the volume of cell solution in culture bag 2 is 1015mL. Step 31: Control the opening of clamp valves 8, 3 and 4, control peristaltic pump 1 to rotate forward and peristaltic pump 3 to rotate in reverse, and add 600mL of amplification culture medium from culture medium bags 3 and 4 to culture bag 1. At this time, the volume of cell solution in culture bag 1 is 950mL. Step 32: Shake all culture platforms for 1 minute to mix the cell solutions in each culture bag; Step 33: The cells continue to grow in their respective culture bags. The cell growth is monitored, and the control logic is the same as in step 14. It is determined whether additional fluid is needed for each culture bag. Step 34: During the current static cell culture in culture bag 3, if it is determined that fluid replenishment is needed, control the opening of clamp valve 8, clamp valve 3 and clamp valve 6, control the peristaltic pump 1 to rotate forward, and add 575 mL of amplification medium from culture bag 3 and culture bag 4 into culture bag 3. At this time, the volume of cell solution in culture bag 3 is 1600 mL. Step 35: During the current static cell culture in culture bag 2, it is determined that replenishment is needed. Control the opening of clamp valves 8, 3 and 5, and control the peristaltic pump 1 to rotate forward. Add 575 mL of amplification medium from culture bags 3 and 4 to culture bag 2. At this time, the volume of cell solution in culture bag 2 is 1590 mL. Step 36: During the current static cell culture in culture bag 1, if it is determined that fluid replenishment is needed, control the opening of clamp valves 8, 3 and 4, control the peristaltic pump 1 to rotate forward and the peristaltic pump 3 to rotate in reverse, and add 650 mL of amplification medium from culture bag 3 and culture bag 4 to culture bag 1. At this time, the volume of cell solution in culture bag 1 is 1600 mL. Step 37: Shake all culture platforms for 1 minute to mix the cell solutions in each culture bag; Step 38: The cells continue to grow in their respective culture bags. The cell growth is monitored, and the control logic is the same as in step 14. It is determined whether liquid replenishment is needed for each culture bag. Step 39: During the current static cell culture in culture bag 3, it is determined that replenishment of medium is needed. Following the control logic of step 34, add 423 mL of amplification medium from culture bag 3 and culture bag 4 to culture bag 3. At this time, the volume of cell solution in culture bag 3 is 2023 mL. Step 40: During the current static cell culture in culture bag 2, it is determined that replenishment is needed. Following the control logic of step 35, add 433 mL of amplification medium from culture bag 3 and culture bag 4 to culture bag 2. At this time, the volume of cell solution in culture bag 2 is 2023 mL. Step 41: During the current static cell culture process in culture bag 1, it is determined that replenishment is needed. Following the control logic in step 36, add 423 mL of amplification medium from culture bag 3 and culture bag 4 to culture bag 1. At this time, the volume of cell solution in culture bag 1 is 2023 mL. Step 42: The cells continue to grow in their respective culture bags. The cell growth is monitored, and the control logic is the same as in step 14. It is determined whether each culture bag needs additional fluid. When it is determined which culture bag needs additional fluid, the cells are harvested for that culture bag.

[0140] By following the steps above, cell culture and automated cell replenishment can be achieved in a closed, sterile environment, and the required number of immune cells can be successfully harvested.

[0141] It should be noted that the step of mixing the cell solution in the above example can be added as needed, and this application does not limit it.

[0142] The display and control module 30 in the integrated cell culture equipment can be used to edit the cell culture program (e.g., adjust the trigger interval of color acquisition commands, adjust the replenishment volume, adjust the platform swing duration, etc.), issue control commands to control the actions of various peristaltic pumps, clamp valves, and base plate control motors (used to control swinging, vibration, or tilting) throughout the cell culture process, control the culture environment such as temperature and carbon dioxide concentration, and display and save culture environment data during the cell culture process, such as the temperature, humidity, carbon dioxide concentration, weight of culture bags, weight of culture medium, and temperature of culture medium in the culture chamber throughout the entire culture process. The display and control module 30 can also upload the culture environment data to the server in real time, allowing users to view the cell culture status on a terminal connected to the server.

[0143] In one exemplary embodiment, a computer program product is provided, including a computer program that, when executed by a processor, implements the steps in the above-described method embodiments.

[0144] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0145] This document uses specific examples to illustrate the principles and implementation methods of this application. The descriptions of the above embodiments are only for the purpose of helping to understand the methods and core ideas of this application. Furthermore, those skilled in the art will recognize that, based on the ideas of this application, there will be changes in the specific implementation methods and application scope. Therefore, the content of this specification should not be construed as a limitation of this application.

Claims

1. A method for cell rehydration, characterized in that, The cell rehydration method includes: During cell culture using a transparent cell culture container, a color image of the cell culture container is acquired in response to a color acquisition command. Cluster the color values ​​of the color image to obtain color value clustering results; Based on the color value clustering results, the current dominant hue color value of the cell solution in the cell culture container is determined; Based on the primary color value, a pre-trained fluid replenishment prediction model is used to determine whether culture medium needs to be added to the cell culture container. If it is determined that the culture medium needs to be added to the cell culture container, then the culture medium is added to the cell culture container.

2. The cell rehydration method according to claim 1, characterized in that, The fluid replenishment prediction model includes a cell density prediction model. The process of determining whether culture medium needs to be replenished into the cell culture vessel based on the dominant color value, obtained through a pre-trained fluid replenishment prediction model, includes: Based on the primary color value, the current cell density of the cell solution in the cell culture container is determined by the cell density prediction model. If the cell density exceeds the cell density threshold, it is determined that culture medium needs to be added to the cell culture container. If the cell density does not exceed the cell density threshold, it is determined that there is no need to replenish the culture medium in the cell culture container at present.

3. The cell rehydration method according to claim 2, characterized in that, The cell density threshold corresponds to the cell type currently being cultured and / or the culture stage at which the current cell culture duration is located.

4. The cell rehydration method according to claim 1, characterized in that, The step of determining the current dominant hue color value of the cell solution in the cell culture container based on the color value clustering results includes: From the color value clustering results, determine the target category that contains the most color values ​​among all the categories obtained from the clustering; The cluster centers of the target category are converted to the HSV color space to obtain the HSV color values ​​corresponding to the cluster centers of the target category; The HSV color value corresponding to the cluster center of the target category is determined as the current dominant color value of the cell solution in the cell culture container.

5. The cell rehydration method according to claim 1, characterized in that, The cell rehydration method also includes: If the current cell culture time exceeds the preset culture time, and it is not determined from the start time of cell culture to the current time that the culture medium needs to be added to the cell culture container, an alarm for abnormal cell growth will be issued.

6. The cell rehydration method according to claim 1, characterized in that, The color acquisition instruction is a periodic instruction based on multiple time intervals. The color acquisition instruction is triggered and executed once every time interval according to the order of the multiple time intervals. The multiple time intervals are the same time intervals or at least some different time intervals.

7. An integrated cell culture device, characterized in that, The integrated cell culture device includes: Culture medium box, which has space for placing culture medium containers for storing culture medium; A cell culture chamber includes a coating device and a culture device. The coating device has a space for placing a cell source container and a coating container. The culture device has a space for placing a cell culture container. The space for placing the cell culture container is equipped with a matching color image sensor and a reflective component. The reflective component is located in the light source emission direction of the matching color image sensor. The valve system includes a first valve device for controlling the pipeline between the culture medium container and the coating container, a second valve device for controlling the pipeline between the cell source container and the coating container, and a third valve device for controlling the pipeline between the coating container and the cell culture container. When the culture medium container, the coating container, the cell source container, and the cell culture container are all connected by pipelines, a closed environment is formed. A control module is electrically connected to the culture medium tank, the cell culture tank, and the valve system, respectively. The control module contains a pre-trained fluid replenishment prediction model and is used to implement the following steps: During cell culture via the cell culture container, in response to a color acquisition command, the color image sensor is controlled to acquire a color image of the cell culture container. Cluster the color values ​​of the color image to obtain color value clustering results; Based on the color value clustering results, the current dominant hue color value of the cell solution in the cell culture container is determined; Based on the primary color value, the replenishment prediction model determines whether culture medium needs to be added to the cell culture vessel. If it is determined that the culture medium needs to be added to the cell culture container, then the first valve device and the third valve device are opened.

8. The integrated cell culture device according to claim 7, characterized in that, The integrated cell culture device includes at least a light-shielding device for shielding the cell culture chamber from light.

9. The integrated cell culture device according to claim 7, characterized in that, The culture device includes a base plate and a protective cover for the cell culture container. When the protective cover is assembled with the base plate, it forms a space for placing the cell culture container. The base plate is equipped with the color image sensor, and the protective cover is equipped with the reflective component. Both the base plate and the protective cover are provided with ventilation holes.

10. The integrated cell culture device according to claim 7, characterized in that, The color image sensor includes a linear color image sensor; The control module is also used to perform the following steps: controlling the cell solution in the cell culture container to move relative to the linear color image sensor, and controlling the linear color image sensor to acquire a color image of the cell culture container during the relative movement.

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