Single-droplet printing system and method

Abstract

The invention discloses a single-droplet printing system and method. The single-droplet printing system comprises a single-droplet printing chip, an image recognition device and a droplet printing device. The single-droplet printing chip comprises a sample bin, a first sheath fluid input module, a second sheath fluid input module, an image recognition area and a droplet output interface; the sample bin is communicated with the first sheath fluid input module; the first sheath fluid input module is communicated with the second sheath fluid input module; the first sheath fluid input module comprises an image recognition area; image recognition devices are arranged on the periphery of the image recognition area and used for recognizing any liquid drop corresponding to the single liquid drop sample passing through the target pipeline; the second sheath liquid input module is used for wrapping any liquid drop in the single-liquid-drop sample and the corresponding first sheath liquid to form a target liquid drop; and the liquid drop printing device is used for receiving the target liquid drops output by the liquid drop output interface, and the target liquid drops are dropped into the corresponding positions of the target collection containers.

Description

TECHNICAL FIELD

[0001] Embodiments of the present application relate to the technical field of bio-manufacturing, and in particular to a single droplet printing system and method. BACKGROUND

[0002] Single droplet separation technology is a technology that can separate single organisms from microbial populations, and has wide application prospects. At present, single droplet separation technology has become an indispensable research tool in many fields. For example, in the fields of microbiology, biomedicine, ecology, environmental science, etc., single clones obtained by separation can find more detailed cell diversity, understand the behavior and reaction mechanism of cells, etc.

[0003] Although single droplet separation technology has made significant progress, there are still many problems in its practical application, especially in droplet printing. In the current laboratory single droplet separation process, the traditional droplet separation technology cannot achieve precise control of droplets. For example, the current droplet printing system cannot adapt to the printing of small-volume single droplets, thereby reducing the printing precision and affecting the continuity of droplet printing. SUMMARY

[0004] The problem to be solved by the embodiments of the present application is to provide a single droplet printing system and method to improve the precise control of droplets and improve the efficiency of single droplet printing.

[0005] To solve the above problems, according to a first aspect of embodiments of the present application, a single droplet printing system is provided, the system comprising: a single droplet printing chip, an image recognition device, and a droplet printing device;

[0006] The single droplet printing chip comprises a sample bin, a first sheath liquid input module, a second sheath liquid input module, an image recognition area, and a droplet output interface;

[0007] The sample bin is used to store single droplet samples;

[0008] The sample bin and the first sheath liquid input module are in communication;

[0009] The first sheath liquid input module and the second sheath liquid input module are in communication;

[0010] The first sheath liquid input module comprises an image recognition area;

[0011] The image recognition area is a target pipeline close to the interface position corresponding to the communication between the first sheath liquid input module and the second sheath liquid input module, and the image recognition device is arranged around the target pipeline, and the image recognition device is used to recognize each droplet corresponding to the single droplet sample passing through the target pipeline;

[0012] The second sheath liquid input module is configured to wrap any one of the single droplet samples and the corresponding first sheath liquid to form a target droplet.

[0013] The droplet printing device is configured to receive the target droplet output by the droplet output interface and drop into a corresponding position of a target collection container.

[0014] Optionally, the image recognition device and the droplet printing device are in communication connection.

[0015] The image recognition device is configured to, when detecting that any one of the single droplet samples passes through the image recognition area, calculate a target time for any one of the single droplet samples to move to the droplet output interface according to a second sheath liquid flow rate and a preset distance of the single droplet sample from the droplet output interface, and transmit the target time to the droplet printing device, so that the droplet printing device performs a single droplet printing countdown operation according to the target time.

[0016] Optionally, the first sheath liquid input module comprises a first sheath liquid storage and a second pipeline.

[0017] The second pipeline comprises the target pipeline.

[0018] The second pipeline is provided with at least two communication interfaces.

[0019] The first sheath liquid storage is configured to store the first sheath liquid.

[0020] Optionally, the second pipeline comprises a first communication interface.

[0021] The first pipeline connected to the sample storage passes through the first communication interface and communicates with the second pipeline, and a T-shaped structure is formed between the first pipeline and the second pipeline.

[0022] Optionally, the system further comprises a power device.

[0023] The power device is configured to drive the first sheath liquid to move along the second pipeline.

[0024] The power device is further configured to drive the single droplet sample in the sample storage to flow into the second pipeline through the first communication interface along the first pipeline connected to the sample storage, the single droplet sample serving as a dispersed phase, and the first sheath liquid serving as a continuous phase, so that the single droplet sample continues to move along the second pipeline under the driving of the first sheath liquid.

[0025] Optionally, the second sheath liquid input module comprises a third pipeline and a second sheath liquid storage.

[0026] The second sheath liquid storage is configured to store the second sheath liquid.

[0027] The second sheath liquid is moved along the third pipeline by the power device.

[0028] Optionally, the first sheath liquid input module and the second sheath liquid input module are in communication;

[0029] The second pipeline comprises a second communication interface;

[0030] The third pipeline comprises a third communication interface;

[0031] The second pipeline in the first sheath liquid input module is in communication with the third communication interface in the third pipeline in the second sheath liquid input module through the second communication interface;

[0032] A T-shaped structure is formed between the third pipeline and the second pipeline.

[0033] Optionally, the image recognition area is arranged between the first communication interface and the second communication interface, and is close to the second communication interface.

[0034] Optionally, one end of the third pipeline is connected with the droplet output interface, and the other end is connected with the second sheath liquid storage.

[0035] Under the driving of the first sheath liquid, the single-droplet sample and the first sheath liquid continue to move along the second pipeline into the third pipeline, and the second sheath liquid wraps any one of the single-droplet sample and the corresponding first sheath liquid to form a target droplet.

[0036] The droplet output interface is used for dropping the target droplet into a corresponding position of a target collection container through a droplet printing device.

[0037] In a second aspect of the embodiment of the present application, a single-droplet printing method is provided, which is applied to a single-droplet printing chip in the single-droplet printing system of any one of the preceding first aspect, and the method comprises:

[0038] The single-droplet sample after the preprocessing is obtained;

[0039] The single-droplet sample is arranged in the first pipeline corresponding to the sample storage according to a preset distance.

[0040] The single droplet sample is driven to move to an image recognition area according to a preset interval and a preset flow rate by the first sheath liquid movement, so that when the image recognition device detects that any one of the single droplet sample passes through the image recognition area, the image recognition detection device calculates a target time for any one of the droplets to move to a droplet output interface according to a second sheath liquid flow rate and a preset distance of the droplet from the droplet output interface, and the image recognition detection device controls the droplet printing device to perform a single droplet printing countdown operation based on the target time.

[0041] The second sheath liquid drives any one of the droplets in the single droplet sample mixed with the first sheath liquid to move along the third pipeline to the droplet output interface through the second communication interface of the second pipeline and the third communication interface of the third pipeline, and the second sheath liquid wraps any one of the droplets and the corresponding first sheath liquid to form a target droplet.

[0042] When the countdown operation corresponding to the countdown time ends, the target droplet flows to the droplet output interface, and the droplet printing device is triggered, so that the droplet printing device controls the target droplet to drop into a corresponding position of a target collection container.

[0043] The technical scheme provided by the application at least brings the following beneficial effects:

[0044] In the application, each droplet in the single droplet sample is double-wrapped by the first sheath liquid module and the second sheath liquid module in the single droplet printing chip, that is, the size of the droplet is accurately controlled through the chip channel design, so that the droplet can be perfectly printed to a multi-well plate or a culture dish by self-gravity during droplet printing, the droplet containing a single clone is wrapped by two sheath liquids, the volume of the individual is enlarged, the picoliter-level droplet collected can be enlarged to microliter level, and accurate control of the droplet is realized; the image recognition area is arranged at a position close to the third communication interface of the second sheath liquid module, and the image recognition device is arranged in the image recognition area, that is, there is a droplet recognition area before the double-wrapped droplet is formed, the delay countdown is triggered after the droplet passes through the recognition area, the movement of the waste spoon is triggered after the countdown ends, and the double droplet containing the positive droplet drops down, so that the picoliter-level single droplet after sorting can be printed, and the subsequent expansion culture is more beneficial.

[0045] The above description is only a summary of the technical scheme of the application, in order to more clearly understand the technical means of the application, the application can be implemented according to the content of the specification, and in order to make the above and other purposes, characteristics and advantages of the application more obvious and easy to understand, the following specific embodiments of the application are described. BRIEF DESCRIPTION OF DRAWINGS

[0046] Figure 1is a schematic structural diagram of a single droplet printing system and method provided by an embodiment of the present application;

[0047] Figure 2 is a schematic structural diagram of a single droplet printing chip in a single droplet printing system and method provided by an embodiment of the present application;

[0048] Figure 3 is a flow chart of steps of a single droplet printing method provided by an embodiment of the present application;

[0049] Figure 4 is a schematic diagram of the principle of a single droplet printing chip in a single droplet printing system and method provided by an embodiment of the present application.

[0050] Legend: 1-single droplet printing chip; 2-image recognition device; 3-droplet printing device; 4-power device; 11-sample bin; 12-first sheath liquid input module; 121-first sheath liquid bin; 122-second pipeline; 1221-first communication interface; 1222-second communication interface; 13-second sheath liquid input module; 131-third pipeline; 132-second sheath liquid bin; 1311-third communication interface; 14-image recognition area; 15-droplet output interface; 16-first pipeline; 17-chip packaging module. DETAILED DESCRIPTION

[0051] Exemplary embodiments of the present application will be described herein below with reference to the accompanying drawings. While exemplary embodiments of the present application are shown in the drawings, it is understood that the present application can be embodied in various forms and should not be limited by the embodiments set forth herein. Rather, these embodiments are provided so that this application will be thorough and complete, and will fully convey the scope of the application to those skilled in the art.

[0052] Hereinafter, embodiments of the present application will be described with reference to the accompanying drawings. However, it should be understood that these descriptions are merely exemplary and are not intended to limit the scope of the present application. In the following detailed description, numerous specific details are set forth in order to provide a thorough understanding of the embodiments of the present application. However, it will be apparent to one skilled in the art that one or more embodiments can be practiced without these specific details. In addition, in the following description, descriptions of well-known structures and techniques have been omitted to avoid unnecessarily obscuring the concept of the present application.

[0053] It should be noted that the directional terms mentioned in the embodiments, such as "upper", "lower", "front", "back", "left", "right" and the like, are only the directions of the drawings, and when the positions of the elements / components are changed, "upper" in the corresponding structure may change to "lower", therefore the above description should not be understood as absolute meaning in order to clearly describe the relative positions, and the above description is not intended to limit the protection scope of the present application. Throughout the drawings, the same elements are denoted by the same or similar reference numerals. When it is possible to cause confusion in understanding the present application, the conventional structures or configurations will be omitted. The shapes and sizes of the components in the drawings do not reflect the actual sizes and proportions, but only illustrate the contents of the embodiments of the present application. In addition, in the claims, any reference symbol located between parentheses should not be construed as a limitation on the claims.

[0054] The single droplet printing system and method provided by the present application is based on the field of droplet microfluidics, innovatively combines visual droplet recognition technology and double-wrapped droplet technology, generates high-throughput picoliter-level single droplets, and sorts the droplets. With the supplement and spacing of the sheath liquid, the droplets flow to the T-shaped channel, and then form double-wrapped droplets through the transverse shearing of the third phase. The double-wrapped droplets are output from the chip to the pipeline, and finally connected to the printing end to perform gravity-based drop printing. There is a droplet recognition area before the double-wrapped droplets are formed, which triggers a delayed countdown after the droplet passes through, and then triggers the movement of the waste spoon to make the double droplets containing the target individual droplets drop down. The length of time depends on the time from droplet recognition to flow to the printing end. In this way, the sorted picoliter-level single droplets can be printed, which is more conducive to subsequent expansion culture. By step-by-step amplification of the channel size, the droplets containing the target individual are wrapped by two kinds of sheath liquid, the volume of the individual is amplified, and the collected picoliter-level droplets can be amplified to microliter-level, realizing precise control of the droplets.

[0055] It should be noted that in the embodiments of the present application, the high-throughput picoliter-level single droplet generation and sorting of the droplets are positive droplets, and the target individual can be a target droplet after secondary wrapping and amplification of the positive droplets after sorting.

[0056] Figure 1 is a system structure schematic diagram of a single droplet printing system and method provided by an embodiment of the present application; Figure 2 is a single droplet printing chip structure schematic diagram in an exemplary single droplet printing system and method provided by an embodiment of the present application.

[0057] In the embodiments of the present application, reference is made to Figures 1-21-single droplet printing chip; 2-image recognition device; 3-droplet printing device; 4-power device; 11-sample bin; 12-first sheath liquid input module; 121-first sheath liquid bin; 122-second pipeline; 1221-first communication interface; 1222-second communication interface; 13-second sheath liquid input module; 131-third pipeline; 132-second sheath liquid bin; 1311-third communication interface; 14-image recognition area; 15-droplet output interface; 16-first pipeline; 17-single droplet chip substrate.

[0058] It should be noted that, due to the current droplet printing system cannot adapt to the small volume of single droplet printing, resulting in the problem of unable to control the droplet accurately, reduce the efficiency of single droplet printing.

[0059] Therefore, in the first embodiment of the present application, a single droplet printing system is provided, which comprises a single droplet printing chip, an image recognition device and a droplet printing device.

[0060] The single droplet printing chip comprises a sample bin, a first sheath liquid input module, a second sheath liquid input module, an image recognition area and a droplet output interface.

[0061] The sample bin is used for storing single droplet samples.

[0062] The sample bin and the first sheath liquid input module are in communication.

[0063] The first sheath liquid input module and the second sheath liquid input module are in communication.

[0064] The first sheath liquid input module comprises an image recognition area.

[0065] The image recognition area is a target pipeline close to the corresponding interface position of the communication between the first sheath liquid input module and the second sheath liquid input module, and the image recognition device is arranged around the target pipeline, and the image recognition device is used for identifying each droplet corresponding to the single droplet sample passing through the target pipeline.

[0066] The second sheath liquid input module is used for wrapping any one droplet in the single droplet sample and the corresponding first sheath liquid to form a target droplet.

[0067] The droplet printing device is used for receiving the target droplet output by the droplet output interface, and dropping into the corresponding position of the target collection container.

[0068] It should be noted that, referring to Figure 1 , Figure 1 It is a system structure schematic diagram of a single droplet printing system and method, which comprises a single droplet printing chip, an image recognition device and a droplet printing device.

[0069] wherein, referring to Figure 2 , Figure 2 The single-droplet printing chip shown in the figure is combined with a visual droplet recognition technology and a droplet double-wrapping technology. The single-droplet printing chip is transparent, so that a subsequent image recognition device can be placed at a corresponding position to recognize the droplets. Therefore, the material of the single-droplet printing chip includes but is not limited to PMMA, COC, PDMS, and glass.

[0070] Specifically, the single-droplet printing chip includes a sample input module, a first sheath liquid input module, a second sheath liquid input module, an image recognition area, and a droplet output interface.

[0071] The sample input module includes a sample bin. The sample bin is used to store a single-droplet sample. The single-droplet sample can be a water-in-oil droplet, that is, a processed droplet containing bacteria or cells. In the present application, the droplet can be a pinoliter-level microdroplet.

[0072] One end of the sample bin is connected to a first pipeline, and the other end can be connected to a power device through an interface outside the chip or a sample bin interface. Then, the power device pushes the droplets into the sample bin through gas or oil phase, so that the droplets are closely arranged in the first pipeline. The power device connected to the sample bin can be a power pump.

[0073] Further, the first sheath liquid input module includes a first sheath liquid bin and a second pipeline. The second pipeline includes the target pipeline. At least two communication interfaces are arranged on the second pipeline. The first sheath liquid bin is used to store a first sheath liquid.

[0074] Further, the second pipeline includes a first communication interface. The first pipeline corresponding to the sample bin communicates with the second pipeline through the first communication interface, and a T-shaped structure is formed between the first pipeline and the second pipeline.

[0075] It should be noted that in the embodiment of the present application, the sample bin and the first sheath liquid input module are in communication. The first sheath liquid input module includes a second pipeline. The first pipeline connected to the sample bin communicates through the first communication interface arranged on the second pipeline. The communication structure is T-shaped.

[0076] The T-shaped structure is not limited to the complete right-angle communication of the two pipelines. In the embodiment of the present application, the length and shape of the pipeline are not limited. The pipeline is a channel arranged inside the chip substrate, which can meet the flow of droplets and sheath liquid.

[0077] Specifically, the first sheath liquid is pushed by the power device to flow along the second pipeline from the first sheath liquid warehouse, and under the action of the first sheath liquid, the droplets closely arranged in the first pipeline are slowly released at the T-shaped channel where the first pipeline and the second pipeline are communicated, wherein the first sheath liquid can be fluorinated oil, and the droplets can also be water-in-oil droplets wrapped by fluorinated oil, at this time, the first sheath liquid is the continuous phase, and the droplets are the dispersed phase, therefore, the droplets will be wrapped by the continuous phase first sheath liquid and flow along the second pipeline in the same direction with the first sheath liquid, and then pass through the image recognition area at a uniform speed, as shown in Figure 4 Figure 4 As can be seen from the schematic view, after the droplets pass through the second pipeline, they will be dispersed by the first sheath liquid, which has already wrapped the droplets in front and back at this time, and the shape of the droplets wrapped on the second pipeline is not approximately circular, but closely adheres to the front and back of the second pipeline, and then through the interface between the second pipeline and the third pipeline, a wrapped droplet shape can be formed by the surface tension between the force and the liquid, and then flows into the third pipeline.

[0078] Further, the image recognition area is arranged between the first communication interface and the second communication interface, and close to the position of the second communication interface.

[0079] Further, the first sheath liquid input module includes an image recognition area, wherein the image recognition area is arranged close to the position of the second sheath liquid module, specifically, the image recognition area refers to the part of the second pipeline close to the corresponding interface position of the first sheath liquid input module and the second sheath liquid input module, which is hereinafter referred to as the target pipeline, and the image recognition device is arranged around the target pipeline, which can be a miniature camera, and the image recognition device is used to identify any one droplet of the single-droplet sample passing through the target pipeline, when the droplet is pushed to the image recognition area, the identification program of the image recognition device is triggered to identify that there is a droplet passing through the target position at present, and the target position is the position point on the second pipeline calibrated by the software identification program.

[0080] Further, the first sheath liquid input module and the second sheath liquid input module are communicated, and the second sheath liquid input module is used to wrap any one droplet in the single-droplet sample and the corresponding first sheath liquid to form a target droplet.

[0081] Specifically, the second sheath liquid input module can include a second sheath liquid warehouse and a third pipeline, wherein the second sheath liquid warehouse can also be directly replaced by a sample bottle.

[0082] ​The second sheath liquid can be silicone oil or mineral oil, the density of the second sheath liquid is less than that of the first sheath liquid, and the first sheath liquid and the second sheath liquid are incompatible, so that the second sheath liquid (low density) is on the top and the first sheath liquid (high density containing single droplet samples) is on the bottom, and the stable stratified flow lays a foundation for the formation of the wrapped droplet.

[0083] At this time, the first sheath liquid mixes the droplet to become a dispersed phase, and the second sheath liquid serves as a continuous phase, that is, the high-throughput picoliter single droplet generation sorts the droplet, with the help of the supplement and spacing of the first sheath liquid, flows to the T-shaped channel formed by the second pipeline and the third pipeline, and then is subjected to transverse shearing of the third phase (the second sheath liquid) to form a double-wrapped droplet, that is, the droplet is wrapped again by the second sheath liquid and part of the first sheath liquid to form a large-volume droplet, that is, a target droplet. The stable stratified flow makes the generation frequency, size and wrapping rate of the droplet more uniform and controllable.

[0084] Further, the second sheath liquid input module comprises a third pipeline and a second sheath liquid storage;

[0085] The second sheath liquid storage is used for storing the second sheath liquid;

[0086] The second sheath liquid moves along the third pipeline through the power device.

[0087] Further, the first sheath liquid input module and the second sheath liquid input module are communicated;

[0088] The second pipeline comprises a second communication interface;

[0089] The third pipeline comprises a third communication interface;

[0090] The second pipeline in the first sheath liquid input module is communicated with the third communication interface in the third pipeline in the second sheath liquid input module through the second communication interface;

[0091] The third pipeline and the second pipeline form a T-shaped structure.

[0092] It should be noted that, in the embodiments of the present application, the second sheath liquid input module can comprise a third communication interface, which is mainly communicated with the second pipeline of the first sheath liquid input module, so as to realize secondary wrapping of the single droplet sample. Similarly, the third pipeline and the second pipeline also form a T-shaped structure in the same way as the communication mode of the first pipeline and the second pipeline. The T-shaped structure is not limited to the complete right angle communication between the two pipelines. In the embodiments of the present application, the length and shape of the pipeline are not limited. The pipeline is a channel arranged inside the chip substrate, which can meet the flow of the droplet and the sheath liquid.

[0093] Furthermore, the droplet printing device is used to receive the target droplet output from the droplet output interface and drop it into the corresponding position of the target collection container.

[0094] Furthermore, one end of the third pipeline is connected to the droplet output interface, and the other end is connected to the second sheath fluid reservoir;

[0095] Driven by the first sheath fluid, the single droplet sample and the first sheath fluid continue to move along the second pipeline and enter the third pipeline. The second sheath fluid encapsulates any one droplet in the single droplet sample and the corresponding first sheath fluid to form a target droplet.

[0096] The droplet output interface is used to drop the target droplet into the corresponding position of the target collection container through the droplet printing device.

[0097] It should be noted that, in the embodiments of this application, one end of the third pipeline is connected to the droplet output interface, and the other end can be connected to the second sheath liquid chamber or the corresponding sample inlet bottle.

[0098] Furthermore, when a large-volume target droplet is formed with double-layer encapsulation, the target droplet is pushed to the droplet output interface of the single-droplet printing chip under the action of the second sheath fluid. The droplet output interface is connected to a droplet printing device. Then, through the droplet printing device, the double-encapsulated target droplet is directly dripped into the collection container by gravity. The collection container can be a porous plate.

[0099] Specifically, through the functions of the first sheath fluid input module and the second sheath fluid input module, a double-layered droplet can be formed. This allows the droplet containing a single clone to be encased in two types of sheath fluid, thus increasing the volume of the individual. The collected droplets at the picolinate level can be enlarged to the microliter level, which means that any volume of droplet cultivated can be precisely docked with the droplet printing device. This allows for precise control of the droplets while enabling single droplet printing to be completed by gravity.

[0100] Furthermore, the image recognition device and the droplet printing device are communicatively connected; the image recognition device is used to calculate the target time corresponding to the movement of any droplet to the droplet output interface based on the second sheath fluid flow rate and the preset distance of the droplet from the droplet output interface when it detects that any droplet in the single droplet sample has passed through the image recognition area. The image recognition detection device transmits the target time to the droplet printing device so that the droplet printing device performs a single droplet printing countdown operation according to the target time.

[0101] It should be noted that, in the embodiments of this application, the image recognition device needs to perform a countdown operation in addition to performing the droplet recognition function.

[0102] Specifically, under the action of the second sheath fluid, large-volume droplets will be continuously formed. However, only some of these droplets contain the target individual, i.e., the droplets in the single droplet sample, i.e., the target droplets. Therefore, after the software identifies any droplet in the single droplet sample, it starts a countdown operation. When the countdown ends, the target droplet flows to the printing position, at which point the printing device is triggered, causing the target droplet to fall into the collection container.

[0103] The countdown operation is based on the initial position of the droplet in the single droplet sample appearing in the image recognition area and being recognized by the image recognition device, with the droplet outlet as the endpoint. The flow rates of the first and second sheath fluids can be pre-adjusted based on the power device. Therefore, after the droplet in the single droplet sample is encapsulated a second time to form the target droplet, it can move according to the preset interval and preset flow rate. That is, the droplet flows at a known and stable speed under the push of the second sheath fluid. Therefore, the time required for this distance is fixed, i.e., ΔT = distance / speed.

[0104] Furthermore, it is also possible to calculate the first time corresponding to a single droplet at the interface under the first sheath fluid flow rate, and then the second time required for the target droplet to move under the second sheath fluid flow rate after being wrapped by the third pipeline.

[0105] Therefore, after the software system corresponding to the image recognition device calculates the target time corresponding to the countdown, the target time can be transmitted to the droplet printing device via communication. Then, at the corresponding time, that is, when the countdown ends, the target droplet just flows to the printing position, and the printing device is triggered, causing the droplet to fall into the collection container. Thus, by adding droplet recognition function and corresponding implementation module, this system improves the printing accuracy and reduces the amount of sheath fluid remaining in the collection container after the droplet is printed, thereby improving the survival rate of the target bacteria or cells in the target droplet.

[0106] Furthermore, the system also includes a power unit;

[0107] The power unit is used to propel the first sheath fluid along the second pipeline;

[0108] The power unit is also used to propel the single droplet sample in the sample chamber along the first pipeline connected to the sample chamber and into the second pipeline through the first communication interface. The single droplet sample serves as the dispersed phase, and the first sheath fluid serves as the continuous phase. Driven by the first sheath fluid, the single droplet sample continues to move along the second pipeline.

[0109] It should be noted that, in the embodiments of this application, the power device can be three different power devices, which are respectively connected to the sample chamber in the first sheath fluid module and the sample input module and the second sheath fluid input module. The power device can include an oil phase or gas phase driven sample injection device, such as a power pump, thereby realizing the flow of the first sheath fluid, the second sheath fluid and the single droplet sample.

[0110] A second aspect of the present invention provides a single-droplet printing method, applied to a single-droplet printing chip in any of the single-droplet printing systems described in the first aspect above, the method comprising:

[0111] Obtain pretreated single-droplet samples;

[0112] The single droplet samples are arranged at a preset distance in the first pipeline corresponding to the sample chamber;

[0113] The single droplet sample is propelled to the image recognition area by the movement of the first sheath fluid at a preset interval and a preset flow rate. When the image recognition device detects that any droplet in the single droplet sample has passed through the image recognition area, the image recognition detection device calculates the target time corresponding to the movement of any droplet to the droplet output interface based on the second sheath fluid flow rate and the preset distance of the droplet from the droplet output interface. Based on the target time, the image recognition detection device controls the droplet printing device to perform a single droplet printing countdown operation.

[0114] Through the second connecting port of the second pipe and the third connecting port of the third pipe, the second sheath fluid drives any one of the droplets in the single droplet sample mixed with the first sheath fluid to move along the third pipe toward the droplet output port, and controls the second sheath fluid to wrap any one of the droplets and the corresponding first sheath fluid to form a target droplet;

[0115] When the countdown time corresponding to the countdown operation ends, and the target droplet flows to the droplet output interface, the droplet printing device is triggered so that the droplet printing device controls the target droplet to fall into the corresponding position of the target collection container.

[0116] A second embodiment of the present invention provides a single-droplet printing method, applied to a single-droplet printing chip in any of the single-droplet printing systems described in the first embodiment above, referring to... Figure 3 , Figure 3 A flowchart illustrating the steps of a single-droplet printing method provided in this application embodiment includes:

[0117] Step 101: Obtain the pretreated single droplet sample;

[0118] It should be noted that, in the embodiments of this application, the single droplet printing chip is installed in the single droplet printing system. When the system starts working, it will obtain pre-treated bacterial or cell-containing droplets as single droplet samples through the sample introduction device.

[0119] Step 102: Arrange the single droplet samples at a preset distance in the first pipeline corresponding to the sample chamber;

[0120] Then, the single droplet samples are arranged at a preset distance in the first tube corresponding to the sample chamber. The preset distance is required to ensure that the droplets are closely arranged in the first tube. This ensures that the second sheath fluid entering the second tube can enter at a constant speed and maintain the preset interval between the droplets, so that the image recognition device can identify the droplets containing the single droplet samples.

[0121] Step 103: The single droplet sample is propelled to the image recognition area by the first sheath fluid movement at a preset interval and a preset flow rate. When the image recognition device detects that any droplet in the single droplet sample has passed through the image recognition area, the image recognition detection device calculates the target time corresponding to the movement of any droplet to the droplet output interface based on the second sheath fluid flow rate and the preset distance of the droplet from the droplet output interface. The image recognition detection device controls the droplet printing device to perform a single droplet printing countdown operation based on the target time.

[0122] It should be noted that, in the embodiments of this application, the movement of the first sheath fluid will propel the single droplet sample to the image recognition area at a preset interval and a preset flow rate, and the image recognition device and the droplet printing device are communicatively connected;

[0123] Because large-volume droplets are continuously formed under the influence of the second sheath fluid, only a portion of these droplets contain the target droplet. The target droplet is the droplet contained within the single-droplet sample, not the various large-volume droplets containing the first sheath fluid that are continuously formed under the influence of the second sheath fluid. Therefore, a countdown operation is necessary to ensure that the droplet printing device collects the target droplets.

[0124] Specifically, once the software identifies any droplet in the single droplet sample, it begins a countdown. When the countdown ends and the target droplet has flowed to the printing position, the printing device is triggered, causing the target droplet to fall into the collection container.

[0125] The countdown operation is based on the initial position of the droplet in the single droplet sample appearing in the image recognition area and being recognized by the image recognition device, with the droplet outlet as the endpoint. The flow rates of the first and second sheath fluids can be pre-adjusted based on the power device. Therefore, after the droplet in the single droplet sample is encapsulated a second time to form the target droplet, it can move according to the preset interval and preset flow rate. That is, the droplet flows at a known and stable speed under the push of the second sheath fluid. Therefore, the time required for this distance is fixed, i.e., ΔT = distance / speed.

[0126] It should be noted that, in the embodiments of this application,

[0127] Furthermore, it is also possible to calculate the first time corresponding to a single droplet at the interface under the first sheath fluid flow rate, and then the second time required for the target droplet to move under the second sheath fluid flow rate after being wrapped by the third pipeline.

[0128] Therefore, after the software system corresponding to the image recognition device calculates the target time corresponding to the countdown, the target time can be transmitted to the droplet printing device via communication. Then, at the corresponding time, that is, when the countdown ends, the target droplet just flows to the printing position, and the printing device is triggered, causing the droplet to fall into the collection container. Thus, by adding droplet recognition function and corresponding implementation module, this system improves the printing accuracy and reduces the amount of sheath fluid remaining in the collection container after the droplet is printed, thereby improving the survival rate of the target bacteria or cells in the target droplet.

[0129] Step 104: Through the second connecting port of the second pipe and the third connecting port of the third pipe, the second sheath fluid drives any one of the droplets in the single droplet sample mixed with the first sheath fluid to move along the third pipe toward the droplet output port, and controls the second sheath fluid to wrap any one of the droplets and the corresponding first sheath fluid to form a target droplet;

[0130] It should be noted that, in the embodiments of this application, the second sheath fluid can be silicone oil or mineral oil. The density of the second sheath fluid is less than that of the first sheath fluid, and the first and second sheath fluids are incompatible. Therefore, the second sheath fluid (low density) is on top, and the first sheath fluid (high density containing single droplet samples) is below. This stable stratified flow lays the foundation for the formation of encapsulated droplets.

[0131] At this point, the first sheath fluid mixed droplets become the dispersed phase, and the second sheath fluid becomes the continuous phase. That is, the high-throughput picoliter single droplet generated and sorted droplets flow to the T-shaped channel formed by the second and third pipelines with the help of the replenishment and spacing of the first sheath fluid. After being laterally sheared by the third phase (second sheath fluid), the droplets are double-encapsulated. That is, the droplets are encapsulated again by the second sheath fluid and part of the first sheath fluid, forming a large-volume droplet, i.e. the target droplet. The stable stratified flow makes the droplet generation frequency, size and encapsulation rate more uniform and controllable.

[0132] Step 105: When the countdown time corresponding to the countdown operation ends and the target droplet flows to the droplet output interface, the droplet printing device is triggered so that the droplet printing device controls the target droplet to drop into the corresponding position of the target collection container.

[0133] It should be noted that, in the embodiments of this application, when a large-volume target droplet with double-layer encapsulation is formed, the target droplet will be pushed to the droplet output interface of the single droplet printing chip under the action of the second sheath fluid. The droplet output interface is connected to a droplet printing device. Then, through the droplet printing device, the double-encapsulated target droplet will be directly dripped into the collection container by gravity. The collection container can be a porous plate.

[0134] Although preferred embodiments of the invention 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 the invention.

[0135] The embodiments of this application have been described above with reference to the accompanying drawings. However, this application is not limited to the specific embodiments described above. The specific embodiments described above are merely illustrative and not restrictive. Those skilled in the art can make many other forms under the guidance of this application without departing from the spirit and scope of the claims, and all of these forms are within the protection scope of this application.

[0136] Obviously, those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. Therefore, if these modifications and variations fall within the scope of the claims and their equivalents, this invention also intends to include these modifications and variations.

Claims

1. A single droplet printing system, characterized by, The system comprises a single droplet printing chip, an image recognition device and a droplet printing device; The single droplet printing chip comprises a sample bin, a first sheath liquid input module, a second sheath liquid input module, an image recognition area and a droplet output interface; The sample bin is used for storing a single droplet sample; The sample bin and the first sheath liquid input module are in communication; The first sheath liquid input module and the second sheath liquid input module are in communication; The first sheath liquid input module comprises an image recognition area; The image recognition area is a target pipeline close to the interface position corresponding to the communication between the first sheath liquid input module and the second sheath liquid input module, and image recognition devices are arranged around the target pipeline, which are used for identifying each droplet corresponding to the single droplet sample passing through the target pipeline; The second sheath liquid input module is used for wrapping any one droplet in the single droplet sample and corresponding first sheath liquid to form a target droplet; The droplet printing device is used for receiving the target droplet output by the droplet output interface and dropping into a corresponding position of a target collection container.

2. The system of claim 1, wherein, The image recognition device and the droplet printing device are in communication connection; The image recognition device is used for, in the case of detecting that any one droplet in the single droplet sample passes through the image recognition area, calculating a target time for any one droplet to move to the droplet output interface according to a second sheath liquid flow rate and a preset distance of the droplet from the droplet output interface, and transmitting the target time to the droplet printing device to enable the droplet printing device to perform a single droplet printing countdown operation according to the target time.

3. The system of claim 1, wherein, The first sheath liquid input module comprises a first sheath liquid bin and a second pipeline; The second pipeline comprises the target pipeline; At least two communication interfaces are arranged on the second pipeline; The first sheath liquid bin is used for storing first sheath liquid.

4. The system of claim 3, wherein, The second pipeline comprises a first communication interface; A first pipeline corresponding to the sample bin is in communication with the second pipeline through the first communication interface, and a T-shaped structure is formed between the first pipeline and the second pipeline.

5. The system of claim 4, wherein, The system further comprises a power device; The power device is used for pushing the first sheath liquid to move along the second pipeline; The power device is also used for pushing the single droplet sample in the sample bin to flow into the second pipeline through the first communication interface along a first pipeline connected to the sample bin, the single droplet sample serving as a dispersed phase and the first sheath liquid serving as a continuous phase, and the single droplet sample continues to move along the second pipeline under the driving of the first sheath liquid.

6. The system of claim 5, wherein, The second sheath liquid input module comprises a third pipeline and a second sheath liquid bin; The second sheath liquid bin is used for storing second sheath liquid; The second sheath liquid moves along the third pipeline through the power device.

7. The system of claim 6, wherein, The first sheath liquid input module and the second sheath liquid input module are in communication; The second pipeline comprises a second communication interface; The third pipeline comprises a third communication interface; The second pipeline in the first sheath liquid input module is communicated through the second communication interface and the third communication interface in the third pipeline in the second sheath liquid input module; The third pipeline and the second pipeline form a T-shaped structure.

8. The system of claim 7, wherein, The image recognition area is arranged between the first communication interface and the second communication interface and is close to the second communication interface.

9. The system of claim 7, wherein, One end of the third pipeline is connected with the droplet output interface, and the other end is connected with the second sheath liquid tank. Under the driving of the first sheath liquid, the single-droplet sample and the first sheath liquid continue to move along the second pipeline, enter the third pipeline, and the second sheath liquid wraps any one of the single-droplet sample and the corresponding first sheath liquid to form a target droplet. The droplet output interface is used for dripping the target droplet into a corresponding position of a target collection container through a droplet printing device.

10. A single droplet printing method characterized by, The single-droplet printing chip is applied to the single-droplet printing system in any one of claims 1-8, and the method comprises: acquiring a preprocessed single-droplet sample; arranging the single-droplet sample in a first pipeline corresponding to a sample tank according to a preset distance; driving the single-droplet sample to move to an image recognition area according to a preset interval and a preset flow rate through first sheath liquid movement, so that when the image recognition device detects that any one of the single-droplet sample passes through the image recognition area, the image recognition detection device calculates a target time for any one of the droplets to move to a droplet output interface according to a second sheath liquid flow rate and a preset distance of the droplet from the droplet output interface, and the image recognition detection device controls the droplet printing device to perform a single-droplet printing countdown operation based on the target time; driving any one of the droplets in the single-droplet sample mixed with the first sheath liquid to move along the third pipeline to the droplet output interface through the second communication interface of the second pipeline and the third communication interface of the third pipeline, and controlling the second sheath liquid to wrap any one of the droplets and the corresponding first sheath liquid to form a target droplet; when the target droplet flows to the droplet output interface when a countdown time corresponding to the countdown operation ends, triggering the droplet printing device, so that the droplet printing device controls the target droplet to drip into a corresponding position of a target collection container.

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