Single-cell paired micro-fluidic chip, micro-fluidic chip runner and preparation method of micro-fluidic chip runner

Abstract

The invention discloses a single-cell paired micro-fluidic chip, a micro-fluidic chip runner and a preparation method thereof, the micro-fluidic chip runner comprises a microsphere runner and at least one separation device, the microsphere runner is provided with a microsphere inlet and a microsphere outlet, the separation device is arranged in the microsphere runner, and the separation device is arranged in the microsphere runner. The microspheres flow through the separation device in the process of flowing from the microsphere inlet to the microsphere outlet, and the separation device separates the microspheres, so that the microspheres which are close to each other can be separated, the microspheres are separated, the microspheres can enter liquid drops singly, and the phenomenon of empty package or multiple package of the liquid drops is avoided. And the separation device separates the microspheres through physical action, so that the stability is good, and the separation effect is good.

Description

Technical Field

[0001] The present application relates to the technical field of microfluidic chips, and in particular to a single-cell pairing microfluidic chip, a microfluidic chip flow channel, and a preparation method thereof. Background Art

[0002] The cross structure is the most common channel structure in the commercial application of microfluidic single-cell sequencing, which can encapsulate microspheres and cells one-to-one into droplets. By adjusting the entry rate of the microspheres and the droplet generation rate, a higher single-packaging rate can be achieved. In the above-mentioned encapsulation process, the hydrogel relies on its own elasticity and deformability to enter the flow channel at a volume fraction of nearly 100%, but due to input pressure fluctuations, system errors or hydrogel size differences, empty packages or multiple packages will appear when the microspheres enter the droplets. Summary of the invention

[0003] The present application aims to solve at least one of the technical problems existing in the prior art. To this end, the present application proposes a microfluidic chip flow channel that can separate microspheres, allowing microspheres to enter droplets individually, thereby improving the empty or multiple microsphere packing phenomenon in the droplets.

[0004] The microfluidic chip flow channel described in the first embodiment of the present application includes:

[0005] A microball flow channel, wherein the microball flow channel is provided with a microball inlet and a microball outlet, and a plurality of microballs flow from the microball inlet to the microball outlet;

[0006] At least one separation device is disposed in the microsphere flow channel and is used to separate the microspheres so that there are intervals between the microspheres.

[0007] The microfluidic chip flow channel according to the embodiment of the first aspect of the present application has at least the following beneficial effects: the microfluidic chip flow channel of the present application includes a microball flow channel and at least one separator, the microball flow channel is provided with a microball inlet and a microball outlet, the separator is provided in the microball flow channel, the microball will flow through the separator in the process of flowing from the microball inlet to the microball outlet, the separator separates the microballs, and can separate the microballs with close spacing, so that there are intervals between each microball, and the microballs can enter the droplet individually, avoiding the phenomenon of empty or multiple droplets. The separator separates each microball by physical action, has good stability, and good separation effect.

[0008] According to some embodiments of the present application, a shaft body for mounting the separation device is provided on the microsphere flow channel, and the separation device can be rotatably mounted on the shaft body.

[0009] According to some embodiments of the present application, two separators are provided, and the two separators are provided on both sides of the microsphere flow channel.

[0010] According to some embodiments of the present application, the separating device includes a plurality of separating teeth, and the plurality of separating teeth are arranged around the circumference of the shaft body, and the angles between the separating teeth are equal.

[0011] According to some embodiments of the present application, the separating device includes two separating teeth, and the two separating teeth are arranged opposite to each other in the radial direction of the shaft body.

[0012] According to some embodiments of the present application, the separation device is configured as a swing rod that can reciprocate in the microsphere flow channel, and one end of the swing rod is disposed on the shaft.

[0013] According to some embodiments of the present application, at least two of the separation devices are provided, and at least two of the separation devices are arranged sequentially in a direction from the microsphere inlet toward the microsphere outlet.

[0014] According to some embodiments of the present application, the microsphere flow channel is provided with a containing cavity, and the partition device is disposed in the containing cavity.

[0015] According to some embodiments of the present application, the width of the microsphere flow channel is 0-999 μm.

[0016] The single-cell pairing microfluidic chip described in the second embodiment of the present application includes the microfluidic chip channel of the first embodiment.

[0017] The single-cell pairing microfluidic chip according to the second aspect of the embodiment of the present application has at least the following beneficial effects: the single-cell pairing microfluidic chip of the present application can separate the microspheres by using the microfluidic chip channel, separate the microspheres with close spacing, and create gaps between the microspheres. The microspheres can enter the droplets individually, avoiding the phenomenon of empty or multiple droplets, thereby ensuring the success rate of single-cell sequencing.

[0018] According to some embodiments of the present application, the single-cell pairing microfluidic chip includes a dispersed phase flow channel, a continuous phase flow channel and a converging flow channel, the dispersed phase flow channel is connected to the microsphere flow channel, the continuous phase flow channel is connected to the microsphere flow channel, and the converging flow channel and the microsphere flow channel are arranged on the same axis.

[0019] According to some embodiments of the present application, the dispersed phase flow channel is configured as a cell flow channel, the continuous phase flow channel is configured as an oil flow channel, and the cell flow channel and the oil flow channel are sequentially arranged in a direction from the microsphere inlet toward the microsphere outlet.

[0020] The method for preparing the flow channel of the microfluidic chip described in the third embodiment of the present application comprises the following steps:

[0021] preparing a template for a microsphere flow channel;

[0022] pouring PDMS onto the template of the microsphere flow channel to form the microsphere flow channel;

[0023] preparing a separator;

[0024] Installing the separation device on the microsphere flow channel to obtain a microfluidic chip flow channel;

[0025] The flow channel of the microfluidic chip is processed by plasma bonding.

[0026] According to the method for preparing the microfluidic chip channel of the third aspect embodiment of the present application, at least the following beneficial effects are achieved: the method for preparing the microfluidic chip channel of the present application forms the microsphere flow channel by inverting PDMS onto the template of the microsphere flow channel, installing the prepared partition device on the microsphere flow channel, and assembling the partition device and the microsphere flow channel after preparing them separately, which can simplify the production process and reduce the difficulty of preparation. Using the partition device to separate the microspheres can make intervals between the microspheres, and the microspheres can enter the droplets individually, avoiding the phenomenon of empty or multiple droplets.

[0027] According to some embodiments of the present application, the template for preparing the microsphere flow channel further includes:

[0028] The template of the microsphere flow channel is prepared by soft lithography technology.

[0029] According to some embodiments of the present application, in the step of preparing the separation device, the step further includes:

[0030] The separation device was prepared by two-photon printing technology.

[0031] Additional aspects and advantages of the present application will be given in part in the description below, and in part will become apparent from the description below, or will be learned through the practice of the present application. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] The above and / or additional aspects and advantages of the present application will become apparent and easily understood from the description of the embodiments in conjunction with the following drawings, in which:

[0033] Figure 1 It is a schematic diagram of the structure in which the multi-pack phenomenon occurs in the prior art;

[0034] Figure 2 is a schematic structural diagram of a microfluidic chip channel according to an embodiment of the present application;

[0035] Figure 3 yes Figure 2 A schematic diagram of the top view of the microsphere flow channel of the microfluidic chip channel shown;

[0036] Figure 4 yes Figure 2 A schematic diagram of the side view structure of a microsphere flow channel of a microfluidic chip channel is shown;

[0037] Figure 5 is a schematic structural diagram of a microfluidic chip channel according to another embodiment of the present application;

[0038] Figure 6 is a structural schematic diagram of a microfluidic chip channel in another embodiment of the present application;

[0039] Figure 7 is a structural schematic diagram of a microfluidic chip channel in another embodiment of the present application;

[0040] Figure 8 is a structural schematic diagram of a microfluidic chip channel in another embodiment of the present application;

[0041] Fig. 9 is a structural schematic diagram of a microfluidic chip channel in another embodiment of the present application;

[0042] Fig.10 This is a schematic diagram of the structure of a single-cell pairing microfluidic chip according to an embodiment of the present application;

[0043] Fig.11 It is a flow chart of a method for preparing a microfluidic chip channel according to an embodiment of the present application.

[0044] Reference numerals:

[0045] Microsphere flow channel 100, microsphere 110, shaft 120, microsphere inlet 121, microsphere outlet 122, accommodating chamber 130;

[0046] Cell flow channel 200, cell 210;

[0047] Oil flow channel 300;

[0048] A collecting channel 400;

[0049] The partition device 500 , the partition teeth 510 , and the rocker rod 520 . DETAILED DESCRIPTION

[0050] The embodiments of the present application are described in detail below, and examples of the embodiments are shown in the accompanying drawings, wherein the same or similar reference numerals throughout represent the same or similar elements or elements having the same or similar functions. The embodiments described below with reference to the accompanying drawings are exemplary and are only used to explain the present application, and cannot be understood as limiting the present application.

[0051] In the description of the present application, it should be understood that descriptions involving orientation, such as up, down, front, back, left, right, inside, outside, etc., indicating orientations or positional relationships, are based on the orientations or positional relationships shown in the accompanying drawings, and are only for the convenience of describing the present application and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore should not be understood as a limitation on the present application.

[0052] In the description of this application, "several" means one or more, "more" means more than two, "greater than", "less than", "exceed", etc. are understood to exclude the number itself, and "above", "below", "within", etc. are understood to include the number itself. If there is a description of "first" or "second", it is only used for the purpose of distinguishing technical features, and cannot be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features or implicitly indicating the order of the indicated technical features.

[0053] In the description of this application, unless otherwise clearly defined, terms such as setting, installing, connecting, assembling, and matching should be understood in a broad sense, and technicians in the relevant technical field can reasonably determine the specific meanings of the above terms in this application based on the specific content of the technical solution.

[0054] Reference Figure 1 In the related art, due to input pressure fluctuations, system errors or hydrogel size differences, empty or multiple packages may appear when the microspheres enter the droplets.

[0055] Refer to the following Figures 2 to 9 The microfluidic chip channel of the embodiment of the present application is described.

[0056] The microfluidic chip channel of the embodiment of the present application refers to Figures 2 to 9 , comprising a microsphere flow channel 100 and at least one partitioning device 500, the microsphere flow channel 100 is provided with a microsphere inlet 121 and a microsphere outlet 122, and a plurality of microspheres 110 enter the microsphere flow channel 100 from the microsphere inlet 121 and flow out from the microsphere outlet 122. In the related art, the phenomenon of empty or multiple microspheres 110 is avoided by adjusting the entry rate of the microspheres 110, but due to input pressure fluctuations, system errors or hydrogel size differences, a plurality of microspheres 110 still enter the microsphere flow channel 100 together with a relatively close spacing. The separator 500 is disposed in the microsphere flow channel 100. The microspheres 110 flow through the separator 500 when flowing from the microsphere inlet 121 to the microsphere outlet 122. The separator 500 separates the microspheres 110, and can separate the microspheres 110 with close spacing, so that there are intervals between the microspheres 110. The microspheres 110 can enter the droplet individually, avoiding the phenomenon of empty or multiple droplets. The separator 500 separates the microspheres 110 by physical action, and has good stability and good separation effect.

[0057] It should be noted that, in some embodiments, the microsphere flow channel 100 is used to transport hydrogel microspheres. In other embodiments, the microsphere flow channel 100 can also be used to transport other microspheres such as protein microspheres and composite microspheres.

[0058] Reference Figure 2 In some embodiments, the microsphere flow channel 100 is provided with an axis 120 for installing the separator 500, and the separator 500 is rotatably installed on the axis 120. The separator 500 is provided on the microsphere flow channel 100. Specifically, the microsphere flow channel 100 is provided with an axis 120, and the separator 500 is rotatably installed on the axis 120. When the microsphere 110 flows through the separator 500, it can drive the separator 500 to rotate. During the rotation process, the rotatable separator 500 can hinder the movement of the next adjacent microsphere 110, so that a gap appears between two adjacent microspheres 110, so as to separate the microspheres 110, so that the microspheres 110 enter the droplet individually, and improve the single package rate. It should be noted that, since the microspheres 110 are arranged in a row in the microsphere flow channel 100 for unidirectional movement, the microspheres 110 that first leave the separation device 500 will move a longer distance than the adjacent blocked microspheres 110, thereby causing a gap between two adjacent microspheres 110. In some embodiments, the shaft 120 and the microsphere flow channel 100 are designed in an integrated manner. In other embodiments, the shaft 120 and the microsphere flow channel 100 are designed in a split manner.

[0059] Reference Figure 2 In some embodiments, two partitioning devices 500 are provided, and the two partitioning devices 500 are provided on both sides of the microsphere flow channel 100. The direction from the microsphere inlet 121 to the microsphere outlet 122 is the length direction of the microsphere flow channel 100 (refer to Figure 2 , Figure 2 The X-axis direction is the length direction), and the width direction is set perpendicular to the length direction (refer to Figure 2 , Figure 2 The Y-axis direction is the width direction), and the two separators 500 are arranged on both sides of the microsphere flow channel 100 in the width direction. During the hydrogel delivery process, the two separators 500 separate the microspheres 110 from both sides in the width direction, and the separation effect of the microspheres 110 is better. It should be noted that in some embodiments, the two separators 500 are arranged relative to each other in the width direction, and the two relatively arranged separators 500 can achieve synchronous separation of the microspheres 110. It can be understood that the separation of the microspheres 110 by the two separators 500 can be performed synchronously or asynchronously. Figure 7 In some embodiments, the two partitioning devices 500 move synchronously under the action of the microsphere 110, and the two partitioning devices 500 simultaneously hinder the movement of one microsphere 110. Figure 8In some other embodiments, the two separation devices 500 move asynchronously, wherein one separation device 500 blocks the movement of the microsphere 110 and the other separation device 500 blocks the movement of another adjacent microsphere 110 , thereby also achieving the separation of the microspheres 110 .

[0060] Reference Figure 2 and Figure 5 In some embodiments, the separator 500 includes a plurality of separator teeth 510, which are arranged around the circumference of the shaft body 120, and the angles between the separator teeth 510 are equal. The separator 500 is rotatably mounted on the shaft body 120, and includes a plurality of separator teeth 510. When the microspheres 110 flow through the separator 500, the separator teeth 510 of the separator 500 are driven to rotate. There are a plurality of separator teeth 510. When the microsphere 110 abuts against the first separator tooth 510 to drive the separator 500 to rotate, the other rotating separator teeth 510 will hinder the next microsphere 110, so that a gap appears between two adjacent microspheres 110, so as to separate the microspheres 110, so that the microspheres 110 enter the droplet individually, and improve the single package rate. The plurality of separation teeth 510 are arranged around the circumference of the shaft body 120, and the angles between the separation teeth 510 are equal. The plurality of separation teeth 510 can circulate around the shaft body 120 under the action of the plurality of microspheres 110, and continuously separate the microspheres 110. In some embodiments, the number of the separation teeth 510 can be set to three, four, five or more, all of which can achieve the separation of the microspheres 110.

[0061] Reference Figure 7 and Figure 8 In some embodiments, the separation device 500 includes two separation teeth 510, and the two separation teeth 510 are relatively arranged in the radial direction of the shaft body 120. The two separation teeth 510 are relatively arranged in the radial direction of the shaft body 120, and the two separation teeth 510 can rotate around the shaft body 120. The microsphere 110 abuts against one end of the separation tooth 510 and drives the separation tooth 510 to rotate. The other end of the rotating separation tooth 510 will hinder the next microsphere 110, so that a gap appears between two adjacent microspheres 110, so as to separate the microspheres 110, so that the microspheres 110 enter the droplet individually, and improve the single package rate. In some embodiments, the two separation teeth 510 adopt a split design. In some other embodiments, the two separation teeth 510 adopt an integrated design.

[0062] Reference Fig. 9It is understood that in some other embodiments, the separation device 500 can also be configured as a swing rod 520 that can reciprocate in the microsphere flow channel 100, one end of the swing rod 520 is set on the shaft body, the swing rod 520 blocks the microsphere 110 when the microsphere 110 passes, the microsphere 110 abuts and drives the swing rod 520 to move, after the microsphere 110 passes the position of the swing rod 520, the swing rod 520 returns to the lowest point to block the subsequent microsphere 110, so that there is a gap between two adjacent microspheres 110, and the separation of the microspheres 110 can also be achieved. It is understood that the reciprocating motion of the swing rod 520 in the microsphere flow channel 100 can be a reciprocating swing between a high point and a low point, or a reciprocating rotation in the microsphere flow channel 100.

[0063] Reference Figure 6 In some embodiments, two partitioning devices 500 are provided, and the two partitioning devices 500 are sequentially arranged in the direction from the microsphere inlet 121 to the microsphere outlet 122. The direction from the microsphere inlet 121 to the microsphere outlet 122 is the length direction of the microsphere flow channel 100 (refer to Figure 6 , Figure 6 The X-axis direction is the length direction), and two separators 500 are sequentially arranged in the length direction of the microsphere flow channel 100. When multiple microspheres 110 move from the flow inlet to the flow outlet, the microspheres 110 will pass through the two separators 500 in sequence. The two separators 500 arranged in sequence can perform secondary separation on the microspheres 110 to avoid the adjacent microspheres 110 not being separated in the first separation, thereby ensuring the separation effect and improving the single package rate. It is understandable that in some other embodiments, the number of separators 500 can also be set to multiple, and multiple separators 500 can perform multiple separations on the microspheres 110, and the separation effect is better.

[0064] Reference Figure 5 In some embodiments, the microsphere flow channel 100 is provided with a receiving chamber 130, and the partition device 500 is provided in the receiving chamber 130. The direction from the microsphere inlet 121 to the microsphere outlet 122 is the length direction of the microsphere flow channel 100 (refer to Figure 5 , Figure 5 The X-axis direction is the length direction), and the width direction is set perpendicular to the length direction (refer to Figure 5 , Figure 5 The Y-axis direction is the width direction), the accommodating cavity 130 is arranged in the width direction of the microsphere flow channel 100, and the accommodating cavity 130 is used to accommodate the separation device 500, and can provide space for the separation device 500, so that the separation device 500 can move in the microsphere flow channel 100 to separate the microspheres 110 flowing in the microsphere flow channel 100. It can be understood that, referring to Figure 2 , Figure 3 and Figure 4In some embodiments, two partitioning devices 500 are provided, and two accommodating chambers 130 are correspondingly provided. The two accommodating chambers 130 are relatively arranged in the width direction of the microsphere flow channel 100 to accommodate the two partitioning devices 500.

[0065] In some embodiments, the width of the microsphere flow channel 100 is 0-999 μm. The direction from the microsphere inlet 121 to the microsphere outlet 122 is the length direction of the microsphere flow channel 100 (refer to Figure 5 , Figure 5 The X-axis direction is the length direction), and the width direction is set perpendicular to the length direction (refer to Figure 5 , Figure 5 The Y-axis direction is the width direction), and the width of the microsphere flow channel 100 is 0-999 μm, which can transport the micron-sized microspheres 110.

[0066] Reference Fig.10 The single-cell pairing microfluidic chip of the second embodiment of the present application includes the microfluidic chip channel of the first embodiment. The single-cell pairing microfluidic chip of the present application can separate the microspheres 110 by using the microfluidic chip channel, separate the microspheres 110 with close spacing, and make intervals between the microspheres 110. The microspheres 110 can enter the droplet individually, avoiding the phenomenon of empty or multiple droplets, and ensuring the success rate of single-cell sequencing.

[0067] In some embodiments, the single-cell paired microfluidic chip includes a dispersed phase flow channel, a continuous phase flow channel and a converging flow channel 400, the dispersed phase flow channel is connected to the microsphere flow channel 100, the continuous phase flow channel is connected to the microsphere flow channel 100, and the converging flow channel 400 and the microsphere flow channel 100 are arranged on the same axis. A dispersed phase fluid is transported in the dispersed phase flow channel, and cells 210 are carried in the dispersed phase fluid. By adjusting the scale of the dispersed phase flow channel and the flow rate of the fluid to appropriate parameters, the cells 210 can enter the droplets one by one. Since the droplets are independent of each other, each droplet can provide a relatively closed micro-reaction environment, and there will be no cross-infection between the cells 210, so the droplets can be used as a very ideal microreactor. In addition to cells 210, droplets can also be used for targets such as proteins and nucleic acids. The dispersed phase fluid and the continuous phase fluid are immiscible or partially miscible. In some embodiments, a single-cell pairing microfluidic chip is used to construct a single-cell droplet with a microsphere 110, the dispersed phase flow channel is set as a cell flow channel 200, and the continuous phase flow channel is set as an oil flow channel 300. The cell flow channel 200 and the oil flow channel 300 are arranged in sequence in the direction from the microsphere inlet 121 to the microsphere outlet 122. The cell flow channel 200 of the single-cell pairing microfluidic chip carries cells 210, and the microsphere flow channel 100 carries microspheres 110 separated by a separator 500. The continuous phase flow channel is the oil flow channel 300. The cell flow channel 200 and the oil flow channel 300 are arranged in sequence in the direction from the microsphere inlet 121 to the microsphere outlet 122. When the cell 210 and the microsphere 110 converge, the two will combine together, one cell 210 matches one microsphere 110, and the matched cell 210-microsphere 110 combination continues to move through the oil flow channel 300 to form a cell 210-microsphere 110 droplet wrapped by the continuous phase and enters the converging flow channel 400.

[0068] Reference Fig.11 The method for preparing the microfluidic chip flow channel of the second aspect of the present application comprises the following steps:

[0069] Step S1: preparing a template of the microsphere flow channel 100 .

[0070] In some embodiments, a shaft 120 for cooperating with the separation device 500 is disposed on the template of the microsphere flow channel 100 , and the shaft 120 and the microsphere flow channel 100 are integrally formed to facilitate subsequent assembly of the separation device 500 .

[0071] In some other embodiments, the microsphere flow channel 100 is further provided with a receiving cavity 130 , and the receiving cavity 130 is integrally formed with the microsphere flow channel 100 to receive and assemble the separation device 500 .

[0072] In some embodiments, in step S1 of preparing the template of the microsphere flow channel 100 , the template of the microsphere flow channel 100 is prepared by soft lithography technology.

[0073] Step S2: pouring PDMS onto the template of the microsphere flow channel 100 to form the microsphere flow channel 100: The microsphere flow channel 100 is prepared using a PDMS pouring process. Specifically, the PDMS mixture is poured onto the template of the microsphere flow channel 100, and the microsphere flow channel 100 is demolded after curing.

[0074] Step S3: preparing the partition device 500: preparing the partition device 500.

[0075] In some embodiments, in step S3 of preparing the partition device 500 , the partition device 500 is prepared by two-photon printing technology.

[0076] Step S4: Install the separator 500 on the microsphere flow channel 100 to obtain a microfluidic chip flow channel: Install the prepared separator 500 on the microsphere flow channel 100. The separator 500 and the microsphere flow channel 100 are prepared separately and then assembled, which can simplify the production process and reduce the difficulty of preparation.

[0077] Step S5: Processing the flow channel of the microfluidic chip by plasma bonding.

[0078] The preparation method of the microfluidic chip channel of the present application is to form the microsphere flow channel 100 by inverting PDMS onto the template of the microsphere flow channel 100, and to install the prepared separation device 500 on the microsphere flow channel 100. The separation device 500 and the microsphere flow channel 100 are prepared separately and then assembled, which can simplify the production process and reduce the difficulty of preparation. The separation device 500 is used to separate the microspheres 110, so that there are intervals between each microsphere 110, and the microspheres 110 can enter the droplet individually, avoiding the phenomenon of empty or multiple droplets.

[0079] The embodiments of the present application are described in detail above in conjunction with the accompanying drawings, but the present application is not limited to the above embodiments, and various changes can be made within the knowledge scope of ordinary technicians in the relevant technical field without departing from the purpose of the present application.

Claims

1. A microfluidic chip flow channel, characterized in that: include: A microball flow channel, wherein the microball flow channel is provided with a microball inlet and a microball outlet, and a plurality of microballs flow from the microball inlet to the microball outlet; At least one separation device is disposed in the microsphere flow channel and is used to separate the microspheres so that there are intervals between the microspheres.

2. The microfluidic chip flow channel according to claim 1, characterized in that: The microsphere flow channel is provided with an axis body for mounting the separation device, and the separation device can be rotatably mounted on the axis body.

3. The microfluidic chip flow channel according to claim 1, characterized in that: There are two partitioning devices, and the two partitioning devices are arranged on both sides of the microsphere flow channel.

4. The microfluidic chip flow channel according to claim 2, characterized in that: The separation device comprises a plurality of separation teeth, and the plurality of separation teeth are arranged around the circumference of the shaft body, and the angles of intervals between the separation teeth are equal.

5. The microfluidic chip flow channel according to claim 2, characterized in that: The separation device comprises two separation teeth, and the two separation teeth are arranged opposite to each other in the radial direction of the shaft body.

6. The microfluidic chip flow channel according to claim 2, characterized in that: The separation device is configured as a swing rod that can reciprocate in the microsphere flow channel, and one end of the swing rod is configured on the shaft body.

7. The microfluidic chip flow channel according to claim 1, characterized in that: At least two of the separation devices are provided, and the at least two separation devices are arranged sequentially in the direction from the microsphere inlet toward the microsphere outlet.

8. The microfluidic chip flow channel according to claim 1, characterized in that: The microball flow channel is provided with a containing cavity, and the partition device is arranged in the containing cavity.

9. The microfluidic chip flow channel according to claim 1, characterized in that: The width of the microsphere flow channel is 0-999 μm.

10. Single cell pairing microfluidic chip, characterized in that: The microfluidic chip flow channel comprises any one of claims 1 to 9.

11. The single-cell pairing microfluidic chip according to claim 10, characterized in that: The single-cell pairing microfluidic chip comprises a dispersed phase flow channel, a continuous phase flow channel and a converging flow channel, the dispersed phase flow channel is connected to the microsphere flow channel, the continuous phase flow channel is connected to the microsphere flow channel, and the converging flow channel and the microsphere flow channel are arranged on the same axis.

12. The single-cell pairing microfluidic chip according to claim 11, characterized in that: The dispersed phase flow channel is configured as a cell flow channel, the continuous phase flow channel is configured as an oil flow channel, and the cell flow channel and the oil flow channel are sequentially arranged in a direction from the microsphere inlet toward the microsphere outlet.

13. A method for preparing a flow channel of a microfluidic chip, characterized in that: The following steps are involved: preparing a template for a microsphere flow channel; pouring PDMS onto the template of the microsphere flow channel to form the microsphere flow channel; preparing a separator; Installing the separation device on the microsphere flow channel to obtain a microfluidic chip flow channel; The flow channel of the microfluidic chip is processed by plasma bonding.

14. The method for preparing a flow channel of a microfluidic chip according to claim 13, characterized in that: The template for preparing the microsphere flow channel also includes: The template of the microsphere flow channel is prepared by soft lithography technology.

15. The method for preparing a flow channel of a microfluidic chip according to claim 13, characterized in that: In the preparation of the separation device, the method further comprises: The separation device was prepared by two-photon printing technology.

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