Micro-droplet dynamic culture and real-time detection device

Abstract

The invention discloses a micro-droplet dynamic culture and real-time detection device which comprises a dynamic culture module and a real-time detection module, and the dynamic culture module comprises a first culture pipeline, a first pump body and a second pump body; the first end of the first culture pipeline is connected with the first pump body, and the second end is connected with the second pump body; the first culture pipeline contains micro-droplets, and the micro-droplets are driven by the first pump body and the second pump body to circularly move in the first culture pipeline; the real-time detection module is connected with the dynamic culture module and comprises a detection device, and the detection device is used for detecting the micro-droplets from the dynamic culture module. By controlling the pushing and pumping actions of the first pump body and the second pump body, liquid drops are driven to circularly move in the first culture pipeline for dynamic culture, high-flux long-time liquid drop circular dynamic culture is realized, meanwhile, the dynamic culture module is connected with the real-time detection module, and during circular reciprocating culture, the real-time detection module is connected with the real-time detection module. And real-time detection of the micro-droplets is realized through the detection equipment.

Description

Technical Field

[0001] This application relates to a device for dynamic culture and real-time detection of microdroplets, and more particularly to a device for dynamic culture and real-time detection of microdroplets. Background Technology

[0002] With increasing demands for experimental efficiency and data scale across various fields, high-throughput droplet microfluidics has become a research hotspot due to its ability to generate massive numbers of independent microdroplet reaction units. This technology enables large-scale parallel experiments, significantly improving experimental efficiency while avoiding cross-contamination and reducing reagent consumption. However, related technologies employ static cultivation methods for droplets, allowing only final detection and hindering real-time monitoring throughout the entire process. Furthermore, static cultivation can lead to slippage and fusion of numerous droplets over long periods, altering their morphology and spacing. These changes increase the difficulty of droplet detection. Therefore, the sheer number of droplets makes long-term operation a challenge, and static cultivation cannot support extended high-throughput cyclic cultivation. Thus, a device capable of real-time detection while simultaneously achieving high-throughput cyclic cultivation is urgently needed. Summary of the Invention

[0003] The problem to be solved by the embodiments of this application is to provide a device for dynamic culture and real-time detection of microdroplets, so as to avoid the problem that static culture cannot achieve high-throughput culture and cannot perform real-time detection, thereby improving the culture throughput of microdroplets and performing real-time detection.

[0004] To address the aforementioned problems, according to one aspect of the embodiments of this application, a device for dynamic cultivation and real-time detection of microdroplets is provided, comprising:

[0005] A dynamic culture module includes a first culture tube, a first pump body, and a second pump body; a first end of the first culture tube is connected to the first pump body, and a second end is connected to the second pump body; wherein, the first culture tube contains microdroplets, and the microdroplets circulate within the first culture tube under the drive of the first pump body and the second pump body;

[0006] A real-time detection module is connected to the dynamic culture module. The real-time detection module includes a detection device for detecting microdroplets from the dynamic culture module.

[0007] Optionally, the dynamic culture module further includes a multi-way valve, which is connected to the first culture pipeline, the second pump body, and the real-time detection module.

[0008] Furthermore, the dynamic culture module also includes a first quick connector and a second quick connector. The first end of the first culture pipeline is connected to the first pump body through the first quick connector, and the second end is connected to the multi-way valve through the second quick connector.

[0009] Optionally, the dynamic culture module further includes a microdroplet sensor, which is disposed at the ends of the first and second ends of the first culture tube, and the microdroplet transmitter is used to monitor the movement of microdroplets in the first culture tube.

[0010] Optionally, the real-time detection module further includes an identification device, one end of which is connected to the dynamic culture module and the other end of which is connected to the detection device.

[0011] Furthermore, the real-time detection module also includes a filtering device, one end of which is connected to the identification device and the other end of which is connected to the detection device.

[0012] Furthermore, the real-time detection module also includes a collection bottle, which is connected to the screening device.

[0013] Optionally, the real-time detection module further includes a second culture tube, which is connected to the detection device.

[0014] Furthermore, the real-time detection module also includes a third pump body, which is connected to the second culture tubing.

[0015] Furthermore, the real-time detection module also includes a third quick connector and a fourth quick connector. The first end of the second culture tube is connected to the detection device through the third quick connector, and the second end is connected to the third pump body through the fourth quick connector.

[0016] The technical solution provided in this application brings at least the following beneficial effects:

[0017] This application incorporates a dynamic culture module where microdroplets are cultured in a first culture tube. The droplets are driven to circulate within the first culture tube by the pumping action of a first and second pump, achieving dynamic cyclic culture of the microdroplets. This increases the culture throughput and supports long-term, high-throughput cyclic culture. The application also includes a real-time detection module to monitor the droplets cultured in the dynamic culture module in real time. This detection device enables real-time monitoring of the entire droplet culture process, obtaining continuous, high-temporal-resolution data trajectories, thereby accurately grasping the real-time status of the culture process. The droplet dynamic culture and real-time detection device provided in this application allows for simultaneous high-throughput cyclic culture and real-time detection of microdroplets. Attached Figure Description

[0018] Figure 1 This is a schematic diagram of the structure of a microdroplet dynamic culture and real-time detection device according to an embodiment of this application.

[0019] Explanation of reference numerals in the attached drawings: 1-Dynamic culture module; 11-First culture tubing; 12-First pump body; 13-Second pump body; 14-Multi-port valve; 15-First quick connector; 16-Second quick connector; 2-Real-time detection module; 21-Detection device; 22-Identification device; 23-Screening device; 24-Collection bottle; 25-Second culture tubing; 26-Third pump body; 27-Third quick connector; 28-Fourth quick connector. Detailed Implementation

[0020] To make the above-mentioned objectives, features and advantages of this application more apparent and understandable, this application will be further described in detail below with reference to the accompanying drawings and specific embodiments.

[0021] The embodiments of this application will now be described with reference to the accompanying drawings. However, it should be understood that these descriptions are exemplary only and are not intended to limit the scope of this application. In the following detailed description, numerous specific details are set forth to provide a thorough understanding of the embodiments of this application for ease of explanation. However, it will be apparent that one or more embodiments may be implemented without these specific details. Furthermore, descriptions of well-known structures and technologies are omitted in the following description to avoid unnecessarily obscuring the concepts of this application.

[0022] It should be noted that the directional terms mentioned in the embodiments, such as "up," "down," "front," "back," "left," and "right," are only for reference to the directions in the accompanying drawings. When the orientation of an element / component changes, "up" in the corresponding structure may change to "down." Therefore, the above description is for the purpose of clearly describing relative positions and should not be construed as absolute. The above description is also not intended to limit the scope of protection of this application. Throughout the accompanying drawings, the same elements are represented by the same or similar reference numerals. Conventional structures or constructions will be omitted where they may cause confusion in understanding this application. The shapes and dimensions of the components in the drawings do not reflect actual size and proportion, but are only schematic representations of the embodiments of this application. Furthermore, in the claims, any reference symbols placed between parentheses should not be construed as limiting the claims.

[0023] This application achieves bidirectional movement of droplets between the culture and detection areas by setting up a cyclic culture zone and a real-time droplet detection zone, and by utilizing the coordinated pushing action of the first and third oil pumps. This effectively supports high-throughput, long-term cyclic culture of droplets, prevents droplet settling, and enables real-time online identification, detection, and screening of droplet states. It elevates traditional endpoint detection to dynamic monitoring and intervention-enabled full-process management, significantly improving experimental efficiency and data quality.

[0024] Figure 1 This is a schematic diagram of the structure of a microdroplet dynamic culture and real-time detection device according to an embodiment of this application.

[0025] In the embodiments of this application, reference is made to Figure 1 1-Dynamic culture module; 11-First culture tubing; 12-First pump body; 13-Second pump body; 14-Multi-port valve; 15-First quick connector; 16-Second quick connector; 2-Real-time detection module; 21-Detection device; 22-Identification device; 23-Screening device; 24-Collection bottle; 25-Second culture tubing; 26-Third pump body; 27-Third quick connector; 28-Fourth quick connector.

[0026] Microdroplets typically refer to tiny liquid particles with a diameter of less than 200 micrometers. In microfluidics, they are dispersed as a phase within a continuous phase such as an oil phase, forming independent units. Each droplet can be considered a miniature reactor or culture vessel. The tiny droplet size, down to the picometer, allows for the parallel generation, manipulation, and analysis of millions of independent reaction units in a single experiment, greatly increasing throughput. The physical isolation of the droplets by the oil phase effectively prevents cross-contamination between samples and avoids reagent adsorption loss on the channel walls. Due to their small size and large specific surface area, the efficiency of internal mixing and external exchange of substances within the droplets is high.

[0027] It should be noted that while static culture is commonly used in microdroplet cultivation, it can easily create concentration gradients of nutrients and metabolic waste within the droplets or culture chamber, leading to microenvironment heterogeneity and affecting the uniformity of cells or reactants. Furthermore, in high-throughput droplet cultivation, a large number of droplets may slip and merge over long periods, and their morphology and spacing may change, increasing the difficulty of droplet detection. The sheer number of droplets makes long-term operation a challenge, thus static culture cannot support long-term high-throughput cyclic culture. Moreover, real-time detection of microdroplets requires them to pass through detection points or devices for data processing. Static culture typically only begins operation at the end of the culture process, sequentially passing through detection points or devices for final detection, making it impossible to monitor the entire droplet process (from the start of cultivation to collection).

[0028] Therefore, in the embodiments of this application, such as Figure 1As shown, this embodiment provides a microdroplet dynamic culture and real-time detection device, including: a dynamic culture module 1, which includes a first culture tube 11, a first pump body 12, and a second pump body 13; a first end of the first culture tube 11 is connected to the first pump body 12, and a second end is connected to the second pump body 13; wherein, the first culture tube 11 contains microdroplets, and the microdroplets circulate within the first culture tube 11 under the drive of the first pump body 12 and the second pump body 13; and a real-time detection module 2, which is connected to the dynamic culture module 1 and includes a detection device 21 for detecting microdroplets from the dynamic culture module 1.

[0029] Specifically, the microdroplet dynamic culture and real-time detection device 21 may include a dynamic culture module 1 and a real-time detection module 2. The dynamic culture module 1 may consist of a first culture tube 11, a first pump body 12, and a second pump body 13. The first culture tube 11 can accommodate microdroplets, creating a closed dynamic culture space for microdroplets that allows for circulation and provides a stable environment. The first culture tube 11 can be a flexible tube; the longer the tube, the more microdroplets it can accommodate. The specific length can be set according to actual needs and is not limited here. To save space, the first culture tube 11 can be coiled to form a coil.

[0030] The microdroplets can be dynamically circulated in the first culture tube 11 by the first pump body 12 and the second pump body 13. Specifically, the first pump body 12 and the second pump body 13 can generate directional fluid dynamics; through precise and coordinated pushing and pulling actions, a stable, unidirectional fluid flow is formed in the culture tube, providing power for the circulatory movement of the microdroplets; by adjusting the output of the two pumps, the movement speed and reciprocating stroke of the microdroplets in the tube can be precisely controlled. One pump pushes, and the other pulls, forming a unidirectional resultant force. The two pumps alternately push / pull, causing the droplets to move back and forth within a fixed section of the tube. For example, when the first pump body 12 performs an extraction action and the second pump body 13 performs an ejection action, the microdroplets move towards the first pump body 12; when the second pump body 13 performs an extraction action and the first pump body 12 performs an ejection action, the microdroplets move towards the second pump body 13. The first pump body 12 and the second pump body 13 thus cycle through extraction and pushing, thereby achieving the circulatory reciprocating movement of the microdroplets and realizing the purpose of dynamic culture.

[0031] The first pump body 12 and the second pump body 13 work together from both ends of the pipeline to push and pull, driving the culture medium containing microdroplets to circulate in the first culture pipeline 11. This dynamic method, compared with the traditional static culture in which the droplets are placed still in the well plate, can improve the mixing uniformity of the substances inside the droplets and can achieve long-term, controllable continuous culture.

[0032] It should be noted that the length of the first culture tube 11 can be greater than the length of the tube occupied by the microdroplets. That is, in the entire fluid pipeline of the device, the microdroplets, as discrete dispersed phases, are enveloped and separated by the oil phase; apart from the tube occupied by the microdroplets themselves, the remaining space in the entire device's pipeline is filled with the oil phase. During cyclical movement, the microdroplets will not exceed the range of the first culture tube 11. Therefore, the length of the first culture tube 11 should meet the length range required for the microdroplets' cyclical movement. In the entire pipeline of the device, apart from the microdroplets, the rest is oil phase. When the pump performs an extraction action, the oil phase is drawn into the pump body, and the microdroplets move towards that side; when the pump performs an ejection action, the oil phase is ejected from the pump body, and the microdroplets move away from that side.

[0033] It should be noted that, in the embodiments of this application, the first pump body 12 and the second pump body 13 can be connected to the control system. The control system can preset the pumping interval of the first pump body 12 and the second pump body 13 according to parameters such as the length of the first culture tube 11, the flow speed of the microdroplets, and the total amount of microdroplets. The first pump body 12 and the second pump body 13 pump and push according to the preset time interval, driving the microdroplets to reciprocate in the first culture tube 11.

[0034] The real-time detection module 2 is connected to the dynamic culture module 1, allowing microdroplets in the first culture tube 11 to enter the real-time detection module 2. The real-time detection module 2 can be equipped with a detection device 21, which can perform online analysis of microdroplets in motion or briefly stationary. This detection is real-time and non-invasive, requiring no interruption of the culture process or opening the system for sampling, and can continuously monitor changes in key parameters such as cell density, pH value, fluorescence signal, or metabolite concentration within the droplets. Integrating the detection module into the dynamic culture flow path upgrades the traditional endpoint detection mode to a process monitoring mode, enabling real-time acquisition of culture data, detection of abnormalities in the culture state, and feedback for adjustment of culture conditions.

[0035] It should be noted that the detection device 21 can be an optical, fluorescence, or image sensor, etc. Absorbance detection, fluorescence detection, etc., can be performed using the detection device 21. For example, during fluorescence detection, the microchannels on the chip can be used as detection points. When microdroplets encapsulating cells pass through these points under the drive of the pump, the fluorescence microscope above can automatically focus and perform multi-channel fluorescence imaging. The software can analyze the fluorescence intensity in real time, thereby plotting dynamic curves of cell proliferation or reporter gene expression. The entire process requires no downtime for sampling, achieving integrated culture-observation.

[0036] Furthermore, the dynamic culture module 1 also includes a multi-way valve 14, which is connected to the first culture pipeline 11, the second pump body 13 and the real-time detection module 2.

[0037] Specifically, the multi-port valve 14 is a fluid control element that switches the connection state between three or more fluid interfaces by changing the position of its internal valve core. The valve core is driven manually, electrically, or pneumatically (e.g., by rotating the valve core or sliding the valve stem), thereby preset and changing the on / off combinations of different pipe ports to achieve flow path reorientation. For example, one input can be flexibly switched to one of multiple outputs, or multiple inputs can be directed to a single output. In high-precision systems such as microfluidics, the multi-port valve 14 (especially the electromagnetically driven type) can achieve automated, high-precision flow path program control.

[0038] In the embodiments of this application, a multi-way valve 14 can be provided in the dynamic culture module 1. The multi-way valve 14 can be located at the intersection of the first culture pipeline 11, the second pump body 13, and the real-time detection module 2, connecting the first culture pipeline 11, the second pump body 13, and the real-time detection module 2. This can guide the movement of microdroplets between different functional modules. For example, when microdroplets need to circulate between the first culture tubing 11, the passage between the multi-way valve 14 and the first culture tubing 11 and the second pump body 13 can be opened, and the passage between the multi-way valve 14 and the real-time detection module 2 can be closed. At this time, by pushing the first pump body 12 and the second pump body 13, the microdroplets can circulate between the first culture tubing 11 without moving towards the real-time detection module 2. When it is necessary to deliver the microdroplets to the real-time detection module 2 for detection, the passage between the multi-way valve 14 and the first culture tubing 11 and the real-time detection module 2 can be opened, and the passage between the multi-way valve 14 and the second pump body 13 can be closed. At this time, by pushing out the first pump body 12, the microdroplets can move towards the real-time detection module 2 without flowing towards the second pump body 13.

[0039] It should be noted that the specific number of passages of the multi-way valve 14 can be set according to actual needs. For example, it can be set as a three-way valve, that is, connecting the first culture pipeline 11, the second pump body 13 and the real-time detection module 2. If it is necessary to connect other pipelines or systems, it can also be set as a four-way valve, a five-way valve, etc. There is no limitation here.

[0040] Furthermore, the dynamic culture module 1 also includes a first quick connector 15 and a second quick connector 16. The first end of the first culture pipeline 11 is connected to the first pump body 12 through the first quick connector 15, and the second end is connected to the multi-way valve 14 through the second quick connector 16.

[0041] Specifically, the first quick connector 15 and the second quick connector 16 can be connected to the two ends of the first culture tubing 11, respectively. The first pump body 12 can be connected to the first quick connector 15 via a tubing. The first quick connector 15 and the second quick connector 16 allow for quick, sealed, and reliable connection and disconnection between the pump body and the culture tubing. When the pump body needs to be calibrated, maintained, or replaced, there is no need to disassemble the complex fluid tubing, thus avoiding the risk of contamination, leakage, or damage that may be introduced by frequent disassembly, and improving the reliability and maintenance efficiency of the system. The multi-way valve 14 can be connected to the second quick connector 16 via a tubing. Through different flow path settings of the multi-way valve 14, microdroplets can be guided to move flexibly along different paths. The second quick connector 16 makes the connection between the first culture tubing 11 and the multi-way valve 14 more convenient, and the downstream functional modules can be adjusted or expanded according to experimental needs.

[0042] Furthermore, the dynamic culture module 1 also includes a microdroplet sensor, which is disposed at the ends of the first and second ends of the first culture tube 11. The microdroplet sensor is used to monitor the movement of microdroplets in the first culture tube 11.

[0043] Specifically, the microdroplets need to move within the culture tubing. To monitor whether the microdroplets reach or approach a preset boundary position during their movement, microdroplet sensors can be deployed at the first and second ends of the first culture tubing 11 to monitor and provide feedback on the droplet state, accurately determining the real-time position and movement of the microdroplets within the circulation tubing. When a droplet passes through the end of the tubing or the preset boundary position, the sensor (usually based on optical or capacitive principles) can instantly capture the signal and determine whether the droplet has reached the preset boundary. If it has, the control system can control the microdroplet to move in the opposite direction according to a pre-set program, thereby achieving the reciprocating motion of the microdroplets within the culture tubing.

[0044] By acquiring the precise time it takes for the droplet to pass through both ends, the system can calculate the droplet's actual speed and cycle time, and perform closed-loop tracking of its specific position in the pipeline. The microdroplet data monitored by the microdroplet sensor can be sent to the control system, which can then feed this data back to the control units of the first pump body 12 and the second pump body 13. This allows for dynamic adjustment of the pump's pushing and pulling rhythm and force, thereby achieving precise programmed control of the droplet's speed, direction, and residence area throughout the pipeline, ensuring the stability and reliability of the cultivation process.

[0045] It should be noted that the microdroplet sensor can also monitor whether the movement speed of the microdroplets is normal, as well as whether there are abnormalities such as droplet loss, merging, or blockage, and send a notification to the control system so that the control system can issue a warning and notify personnel to intervene.

[0046] Furthermore, the real-time detection module 2 also includes an identification device 22, one end of which is connected to the dynamic culture module 1 and the other end of which is connected to the detection device 21.

[0047] Specifically, the identification device 22 can be an optical detection device, an image recognition device, an electrical detection device, or an acoustic detection device. For optical identification, a specific wavelength of light (such as a laser) can be used to illuminate the droplet as it flows through the detection window, and the generated fluorescence, reflected light, or absorbed light signals can be detected. For image recognition, a high-speed camera or microscope can be used to continuously photograph the droplet, and image algorithms can be used to analyze its size, shape, color, internal structure, or specific markers (such as fluorescent spots). For electrical identification, the changes in impedance, capacitance, or conductivity caused by the droplet passing through a microelectrode can be measured to reflect its size, content concentration, or dielectric properties. For acoustic detection, the modulation effect of the droplet on surface acoustic waves or ultrasonic waves can be analyzed, or the droplet can be manipulated using acoustic radiation force. For example, a high-speed CMOS camera can be installed on the outside of the corresponding position in the pipeline as an image recognition sensor, along with a bright field light source. When the droplet flows through the transparent pipeline section, the camera captures continuous frames, and the built-in image processing algorithm analyzes the diameter and roundness of the droplet in real time as a basis for initial screening. Alternatively, the identification device 22 can be replaced with a photomultiplier tube, which performs preliminary identification by detecting the intensity of the scattered light pulse or basic fluorescence signal generated when the droplet passes through.

[0048] The identification device 22 receives microdroplets from the dynamic culture module 1 and controls the path of the microdroplets into the detection device 21 based on the judgment result. It can perform real-time identification and screening of circulating microdroplets, for example, automatically identifying the target droplets to be detected based on preset optical characteristics (such as specific fluorescent label intensity), physical characteristics (such as droplet size), or time logic (such as after a specific culture cycle). This can improve detection efficiency and reagent utilization, reduce data redundancy, and achieve real-time, event-triggered detection. For example, it can trigger high-resolution imaging or spectral analysis only when cell droplets expressing a specific reporter gene are identified.

[0049] Furthermore, the real-time detection module 2 also includes a screening device 23, one end of which is connected to the identification device 22 and the other end of which is connected to the detection device 21.

[0050] Specifically, the screening device 23 is connected to the identification device 22. After the identification device 22 identifies the microdroplets, the screening device 23 receives the judgment command from the identification device 22 and performs physical separation and routing operations on the microdroplets. According to the working principle, the screening device 23 can be divided into two main categories: active and passive. Active screening includes: dielectric electrophoresis screening, which applies voltage to the microelectrode to generate a non-uniform electric field and uses the difference in dielectric properties of the droplets or contents to generate a controllable force to deflect them; pneumatic microvalve / hydraulic switching screening, which uses air pressure or hydraulic pressure to instantly push the membrane valve on the chip or switch the flow path to change the flow direction of the target droplets; and piezo-acoustic sorting, which uses the piezoelectric effect to generate sound waves to precisely control the droplets containing individual cells to fall into the designated collection container. Passive screening includes: microchannel structure screening, which utilizes the specific geometry of microchannels (such as contraction-expansion, branching, and barrier arrays) to achieve separation based on the physical properties of droplets, such as size and deformability; and interfacial tension screening, which utilizes the difference in interfacial tension between different fluids to control the entry of droplets into a specific phase at the confluence of channels. In this application, the screening device 23 can be selected according to actual needs, and no restrictions are imposed here.

[0051] The screening device 23 precisely sorts and guides the target droplets that require in-depth detection to the detection device 21, while sending other droplets back to the main culture loop or guiding them to other collection paths. For example, droplets that do not meet the detection requirements can be introduced into the collection bottle 24 for collection.

[0052] The screening device 23 allows the system to perform high-throughput, automated targeted screening and enrichment of massive amounts of microdroplets based on the recognition results. For example, in cell culture, it can automatically select cell droplets with optimal growth status or expressing specific proteins for subsequent detection or amplification; in enzyme evolution screening, it can rapidly isolate mutants with the highest activity. This can improve the efficiency and purposefulness of experiments, enabling the entire platform to autonomously complete the complete intelligent workflow of culture-recognition-sorting-detection.

[0053] Furthermore, the real-time detection module 2 also includes a collection bottle 24, which is connected to the screening device 23.

[0054] Specifically, the collection bottle 24 is connected to the screening device 23. The collection bottle 24 receives and stores microdroplets that have been determined by the screening device 23 and do not require further offline analysis or have retention value (i.e., droplets that do not meet the requirements), so that qualified droplets can enter the detection device 21.

[0055] Furthermore, the real-time detection module 2 also includes a second culture tube 25, which is connected to the detection device 21.

[0056] Specifically, a second culture pipeline 25 can be set in the real-time detection module 2. After the microdroplets are detected by the detection device 21, they can be transported to the second culture pipeline 25 for cultivation. Upon entering the real-time detection module 2, the microdroplets are first identified by the identification device 22, and then filtered out by the screening device 23. These unqualified droplets can be collected. Qualified target droplets can enter the second culture pipeline 25 for further cultivation and, through the cooperation of the third pump 26 and the first pump 12, return to the detection device 21 for further detection. Qualified droplets can be collected. The second culture pipeline 25 is connected to the detection device 21, guiding the microdroplets to be analyzed from the main circulation culture path to the detection area.

[0057] The first culture line 11 can provide long-term and stable cyclic culture conditions for microdroplets, while the second culture line 25 can serve as a dedicated interface to ensure that detection activities (such as laser irradiation and image acquisition) do not interfere with the continuity and stability of the main culture process.

[0058] It should be noted that after the droplets pass through the screening device 23 for the first time, some unqualified droplets may be discarded or the target droplets may be collected. At this time, the third pump 26 stops working and pauses. The first pump 12 pushes the droplets into the collection bottle 24, which may cause the droplets in front to stay in the detection device 21 for a long time, and the time difference of the entire droplets passing through the detection device 21 will increase. Therefore, after the droplets enter the real-time detection module 2 from the dynamic culture module 1, they can be tested for the first time when they enter the detection device 21. All qualified droplets that have passed through the identification device 22 can be tested when they return to the detection device 21 from the second culture pipeline 25.

[0059] Furthermore, the real-time detection module 2 also includes a third pump body 26, which is connected to the second culture tubing 25.

[0060] Specifically, the third pump body 26 can cooperate with the first pump body 12 to push and draw, and cooperate with the multi-way valve 14 to control the closure of the passage of the second pump body 13, realizing the reciprocating movement of the droplet between the dynamic culture module 1 and the real-time detection module 2. For example, when it is necessary for the droplet to enter the real-time detection module 2 from the dynamic culture module 1, the passage connecting the multi-way valve 14 to the first culture pipeline 11 and the real-time detection module 2 can be opened, the passage connecting the second pump body 13 can be closed, the first pump body 12 can be pushed out, and the third pump body 26 can draw it out. At this time, the droplet is driven to move. Since the passage connecting the multi-way valve 14 to the second pump body 13 is closed, the droplet can flow to the real-time detection module 2; when it is necessary for the droplet to enter the dynamic culture module 1 from the real-time detection module 2, the third pump body 26 can be pushed out, and the first pump body 12 can draw it out. At this time, the droplet is driven to move. Since the passage connecting the multi-way valve 14 to the second pump body 13 is closed, the droplet can flow to the dynamic culture module 1.

[0061] By precisely controlling the fluid in the second culture tube 25 through the third pump body 26, it can be ensured that the microdroplets pass smoothly through the detection area at an ideal speed and posture, reducing the interference of fluid disturbance on optical, electrochemical and other detection signals, improving the resolution, reproducibility and accuracy of detection data, and enabling real-time detection data to be reliably fed back to the regulation of the culture process.

[0062] Furthermore, the real-time detection module 2 also includes a third quick connector 27 and a fourth quick connector 28. The first end of the second culture tube 25 is connected to the detection device 21 through the third quick connector 27, and the second end is connected to the third pump body 26 through the fourth quick connector 28.

[0063] Specifically, the third quick connector 27 and the fourth quick connector 28 can be connected to the two ends of the second culture tubing 25, respectively. The detection device 21 can be connected to the first quick connector 15 through the tubing, enabling plug-and-play functionality of the detection unit. This allows different types of detection devices 21 (such as optical microscope and spectrometer probes) to be flexibly, quickly, and securely connected to the system, facilitating the replacement or upgrading of detection methods according to experimental needs without altering the core flow path. The fourth quick connector 28 connects the second culture tubing 25 to the third pump body 26, making the third pump body 26 detachable. This facilitates the calibration, maintenance, or replacement of the third pump body 26, ensuring long-term stability of the driving accuracy.

[0064] The third quick connector 27 and the fourth quick connector 28 decouple the three core components of the detection pipeline, pump body and detection device 21 from the physical connection, which improves the convenience of assembly, debugging and maintenance of the real-time detection module 2. When the detection channel needs to be cleaned or disinfected or a certain part fails, the connector can be quickly disconnected for local treatment, which improves the reliability and availability of the system and reduces the complexity and cost of long-term operation.

[0065] It should be noted that in this embodiment, the device can be divided into a dynamic culture module 1 and a real-time detection module 2. During dynamic culture, the multi-way valve 14 can be opened to connect the first culture pipeline 11 and the second pump body 13. By controlling the first pump body 12 and the second pump body 13 to cooperate in pumping and pushing, the microdroplets can be made to reciprocate in the first culture pipeline 11 for dynamic culture. When testing is required, the passage connecting the multi-way valve 14 to the second pump body 13 can be closed, and the passage connecting to the real-time detection module 2 can be opened. The first pump body 12 and the third pump body 26 can then be deployed to extract the droplets, which will then flow to the real-time detection module 2. The micro-droplets are first identified by the identification device 22, and then the screening device 23 filters out droplets that do not meet the requirements for collection. The target droplets that meet the requirements can first flow through the detection device 21 and enter the second culture pipeline 25. At this time, the third pump body 26 can be deployed, and the first pump body 12 can be deployed to extract the droplets, causing them to flow towards the dynamic culture module 1. When they flow into the detection device 21, they can be tested. The target droplets that meet the requirements after testing can be collected after passing through the screening device 23, and the remaining droplets are returned to the dynamic culture module 1 to continue dynamic culture. When testing is required, the above steps can be repeated.

[0066] In other words, in this embodiment, the droplet movement is divided into two stages within a complete working cycle of the device: forward movement and reverse movement. During forward movement, the droplet is pushed from the dynamic culture module 1 into the real-time detection module 2 and flows through the identification device 22. At the identification device 22, the size, shape, or basic fluorescence of the droplet can be initially identified and judged by rapid imaging or optical signals. Subsequently, based on the identification results, droplets that do not meet the preset standards (such as abnormal size, empty droplets, etc.) can be actively sorted by the sorting mechanism when flowing through the screening device 23 and rejected into the collection bottle 24, thereby achieving initial screening. Qualified droplets continue to move forward and enter the second culture pipeline 25. After all droplets have completed forward movement, the system switches the oil pump action and starts reverse movement. At this point, the droplets are pushed back from the real-time detection module 2 to the dynamic culture module 1 and flow through the detection device 21. At the detection device 21, the droplets can be detected more precisely (such as biochemical indicators such as fluorescence intensity and growth rate). Subsequently, when the droplets flow through the screening device 23, the sorting mechanism can be activated again based on the detection results to sort the target droplets that meet specific conditions (such as cell droplets that highly express the target protein) to the outlet of the collection bottle 24, thereby achieving precise sorting of the target droplets.

[0067] When droplets no longer require real-time detection and screening, or when they return to the first culture tube 11 after completing one round of detection and sorting, the dynamic culture module 1 enters an independent cyclic culture mode. In this mode, the first pump 12 and the second pump 13 work together, using a push-pull method to provide stable and uniform power for the closed-loop cyclic movement of droplets within the first culture tube 11. This cyclical movement effectively avoids problems such as sedimentation, aggregation, or uneven nutrient gradients caused by prolonged static placement of droplets, and can achieve the culture of 2000 microdroplets with a size of 2-3 μL for 7 to 10 days.

[0068] This application comprises a dynamic culture module 1 and a real-time detection module 2. The dynamic culture module 1 consists of a first pump body 12, a second pump body 13, and a first culture pipeline 11. The first pump body 12 and the second pump body 13 act as power sources, using a push-pull mechanism to circulate the droplets within the first culture pipeline 11, achieving dynamic culture of microdroplets and increasing the culture throughput. This supports long-term, high-throughput cyclic culture. The real-time droplet detection area includes droplet identification, screening, and detection. Droplets sequentially enter the second culture pipeline 25 through the identification device 22. Unqualified droplets are discarded (collected) through the screening channel and detected during reverse movement. After detection, the droplets return to the cyclic culture area for continued culture. The droplet dynamic culture and real-time detection device provided in this application can achieve high-throughput cyclic culture and perform real-time detection of microdroplets. By separating the culture area and the detection area, it can support both long-term culture and on-demand droplet detection and screening.

[0069] It should be noted that, in this document, relational terms such as "first" and "second" are used merely to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one..." does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes the element.

[0070] It should be noted that when a component is described as "fixed to" another component, it can be directly on the other component or may have a component in between. When a component is considered "connected to" another component, it can be directly connected to the other component or may have a component in between. When a component is considered "set on" another component, it can be directly set on the other component or may have a component in between. The terms "vertical," "horizontal," "left," "right," and similar expressions used in this document are for illustrative purposes only.

[0071] Although preferred embodiments of this application have been described, those skilled in the art, upon learning the basic inventive concept, can make other changes and modifications to these embodiments. Therefore, the appended claims are intended to be interpreted as including the preferred embodiments as well as all modifications and variations falling within the scope of this application.

[0072] Obviously, those skilled in the art can make various modifications and variations to this application without departing from the spirit and scope of this application. Therefore, if such modifications and variations fall within the scope of the claims of this application and their equivalents, this application also intends to include such modifications and variations.

Claims

1. A device for dynamic cultivation and real-time detection of microdroplets, characterized in that, The device includes: A dynamic culture module includes a first culture tube, a first pump body, and a second pump body; a first end of the first culture tube is connected to the first pump body, and a second end is connected to the second pump body; wherein, the first culture tube contains microdroplets, and the microdroplets circulate within the first culture tube under the drive of the first pump body and the second pump body; A real-time detection module is connected to the dynamic culture module. The real-time detection module includes a detection device for detecting microdroplets from the dynamic culture module.

2. The microdroplet dynamic culture and real-time detection device according to claim 1, characterized in that, The dynamic culture module also includes a multi-way valve, which is connected to the first culture pipeline, the second pump body, and the real-time detection module.

3. The microdroplet dynamic culture and real-time detection device according to claim 2, characterized in that, The dynamic culture module further includes a first quick connector and a second quick connector. The first end of the first culture pipeline is connected to the first pump body through the first quick connector, and the second end is connected to the multi-way valve through the second quick connector.

4. The microdroplet dynamic culture and real-time detection device according to claim 1, characterized in that, The dynamic culture module also includes a microdroplet sensor, which is disposed at the ends of the first and second ends of the first culture tube. The microdroplet transmitter is used to monitor the movement of microdroplets in the first culture tube.

5. The microdroplet dynamic culture and real-time detection device according to claim 1, characterized in that, The real-time detection module also includes an identification device, one end of which is connected to the dynamic culture module and the other end of which is connected to the detection device.

6. The microdroplet dynamic culture and real-time detection device according to claim 5, characterized in that, The real-time detection module also includes a filtering device, one end of which is connected to the identification device and the other end of which is connected to the detection device.

7. The microdroplet dynamic culture and real-time detection device according to claim 6, characterized in that, The real-time detection module also includes a collection bottle, which is connected to the screening device.

8. The microdroplet dynamic culture and real-time detection device according to claim 1, characterized in that, The real-time detection module also includes a second culture tube, which is connected to the detection device.

9. The microdroplet dynamic culture and real-time detection device according to claim 8, characterized in that, The real-time detection module also includes a third pump body, which is connected to the second culture pipeline.

10. The microdroplet dynamic culture and real-time detection device according to claim 9, characterized in that, The real-time detection module also includes a third quick connector and a fourth quick connector. The first end of the second culture tube is connected to the detection device through the third quick connector, and the second end is connected to the third pump body through the fourth quick connector.

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