A droplet reading chip and methods of use thereof

Abstract

The present application relates to the field of droplet microfluidics, and more particularly to a droplet reading chip and a method of using the same. The disclosed droplet reading chip comprises a microcavity for tiling droplets and a pipette for pipetting droplets into the microcavity. The chip can tile a large number of droplets at the same time, meeting the requirement of high-throughput detection of droplets. In addition, the chip can keep the droplets in a relatively closed space during detection, reducing the risk of contamination.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of droplet microfluidics, and more particularly to a droplet reading chip and a method of using the same. BACKGROUND

[0002] Microfluidics refers to a technology for manipulating fluid in a micron scale space, which can miniaturize the basic functions of a chemical or biological laboratory onto a chip of a few square centimeters, and is therefore also known as a "lab-on-a-chip". Droplet microfluidics is an important branch of microfluidic platforms developed in recent years, which is a non-continuous flow microfluidic technology for experimental operation using dispersed microdroplets generated by two incompatible liquids, and has advantages such as reducing reagent consumption, reducing pollution risk, improving reaction accuracy, and shortening reaction time. Among them, droplets as "micro-reactors" have the characteristics of regionalization, miniaturization, controllability and high throughput.

[0003] Droplet microfluidic technology includes droplet generation, droplet manipulation, and droplet detection. Among them, the reading of high-throughput droplet signals is an important link of droplet detection technology. The existing technology mainly focuses on the research of droplet generation, droplet operation, detector performance, chip manufacturing and application of droplet microfluidic technology, and less research on how droplets enter the detector and how they are detected in a certain arrangement. In addition, droplets are often in an open state in contact with the outside atmosphere during signal reading, which is easy to cause pollution. SUMMARY

[0004] The present application provides a droplet reading chip and a method of using the same, which has low cost, is easy to use, and has low pollution risk during use, and meets the detection needs of high-throughput droplets.

[0005] The purpose of the present application is to solve the problem by the following technical scheme:

[0006] A droplet reading chip, characterized in that it comprises a microcavity and a sample suction needle; the microcavity is used for paving droplets; the sample suction needle is used for sucking droplets into the microcavity. The sample suction needle is in communication with the microcavity and can exchange fluid. The microcavity can be a closed cavity, and droplets can enter the microcavity through the sample suction needle by pressurizing the solution containing droplets. A hole can also be provided on the microcavity, and droplets can enter the microcavity through the sample suction needle by suction at the hole.

[0007] Preferably, the chip comprises a channel in communication with the outside, which is not in communication with the microcavity.

[0008] Further, when the channel and the sample suction needle are connected to the same closed space, the channel can make the solution containing droplets in the closed space enter the microcavity through the sample suction needle by flowing with the outside gas.

[0009] Further, the channel can discharge gas in the closed space and the microcavity. The gas in the microcavity and the closed space can be discharged through the channel to form a negative pressure with the same pressure.

[0010] Further, the channel can input gas to the closed space, the pressure of the closed space increases, the negative pressure in the microcavity remains unchanged, and a pressure difference is formed between the microcavity and the closed space. Under the action of the pressure difference, the solution containing droplets in the closed space enters the microcavity through the sample needle.

[0011] Further, in the process of discharging gas, one end of the sample needle is immersed in the solution containing droplets in the closed space, and the other end of the channel is not immersed in the liquid containing droplets.

[0012] Further, the other end of the sample needle communicates with the microcavity, and the other end of the channel communicates with the external atmosphere.

[0013] Further, the closed space is formed by a sealed sample tube, the sample tube contains a solution containing droplets, and there is a gas zone above the solution. The sample tube can be a PCR tube, a 96-well plate, a 384-well plate, a centrifuge tube, a liquid phase vial, or other containers containing droplets. The gas zone is a blank space for gas

[0014] Preferably, a filter screen is arranged in the channel, the filter screen has a mesh structure for preventing aerosol pollution; the filter screen can further be a microporous filter membrane structure with smaller pore size, which allows gas to pass through but does not allow microorganisms such as bacteria to pass through, for example, a 0.22 μm microporous filter membrane, so that the risk of pollution can be further reduced.

[0015] Preferably, the chip includes a serpentine cavity, the microcavity is connected to the serpentine cavity, and the serpentine cavity is generally arranged between the microcavity and the sample needle to trap possible bubbles and prevent the bubbles from entering the microcavity.

[0016] Preferably, the chip includes a liquid storage cavity, the microcavity is connected to the liquid storage cavity, and the liquid storage cavity is used to store the continuous phase such as the oil phase or the aqueous phase in the solution containing droplets.

[0017] Preferably, the chip includes a transparent upper cover plate and a microfluidic plate; the transparent upper cover plate and the microfluidic plate are sealed to form the microcavity. The transparent upper cover plate and the microfluidic plate are provided with notches to facilitate the embedding and sealing of the two.

[0018] Preferably, the microfluidic plate includes a serpentine channel, and the serpentine channel is sealed with the transparent upper cover plate to form the serpentine cavity.

[0019] Preferably, the microfluidic plate comprises a reservoir, which forms a reservoir cavity in sealing with the transparent upper cover plate.

[0020] Further, the sample suction needle comprises a base and a needle, and the base is used to fit with the microfluidic plate.

[0021] Further, the transparent upper cover plate, the microfluidic plate and the base are all provided with a through hole, which is used to form the channel after combination.

[0022] Preferably, the microcavity limits the height of the droplet, and the height is 0.5-1.2 times of the diameter of the droplet, so that the droplets are laid flat in a layer, which is beneficial to the collection of subsequent droplet signals and reduces the interference between each other. Most preferably, the height of the microcavity is the diameter of the droplet.

[0023] Further, the number of the microcavity and the sample suction needle is not less than 1. The microcavity and the sample suction needle can be an independent system corresponding to each other, and the number of the microcavity and the sample suction needle can be adjusted according to the use requirement, so that multiple droplet samples can be read at the same time.

[0024] Preferably, the sample suction needle comprises a conical hollow needle, which is easier to enter the sample solution.

[0025] Preferably, the chip comprises an auxiliary sealing member, which is used to seal the closed space.

[0026] Preferably, the auxiliary sealing member is provided with a sealing groove and a tearing groove. The sealing groove is used to seal the sample tube and ensure the sealing of the closed space; the tearing groove can be made thinner or made of softer material, so as to facilitate the penetration of the sample suction needle.

[0027] Further, the auxiliary sealing member is a silica gel cap. In addition, the auxiliary sealing member can also be made of other elastic materials such as rubber, resin and plastic. In order to ensure sealing, an aluminum foil cover or the like can be further arranged on the periphery.

[0028] Referring to Figure 1 A use method of the droplet reading chip, which comprises the following steps:

[0029] Step (1): a solution containing a droplet is located in a sealed sample tube to form a closed space, and a gas zone is formed at the upper end of the closed space, which is recorded as B zone. The B zone is communicated with a channel, one end of the channel is communicated with the B zone, and the other end is communicated with the outside. The gas exchange between the closed space and the outside can be carried out through the channel. The sample suction needle enters the solution containing the droplet.

[0030] Step (2): the gas in the B zone is discharged through the channel. If there is gas in the microcavity, it will be discharged through the sample suction needle, the B zone and the channel. At this time, the gas pressure in the microcavity is recorded as P.aim The gas in the B area can be discharged by pumping the gas out of the end of the channel which is in communication with the outside.

[0031] Step (3): gas is introduced into the B area through the channel, at this time, the gas pressure in the B area is P0;

[0032] Step (4): P0 is higher than P aim A negative pressure is formed in the microcavity, and the solution containing droplets is introduced into the microcavity through the sample needle until the gas pressure in the B area and the microcavity is equal.

[0033] Step (5): the droplets are spread in the microcavity, and the chip is placed in an optical detection system to read the signal of the droplets in the microcavity.

[0034] Further, by controlling P0 and P aim A specific negative pressure is formed in the microcavity, so as to control the amount of the solution containing droplets introduced into the microcavity.

[0035] Further, in step (3), the channel can be in communication with the outside air, so that air is introduced into the B area, at this time, P0 is the atmospheric pressure; or the channel can be used to fill gas into the B area to reach a specific pressure.

[0036] Further, the gas in step (3) can be air, or inert gases such as nitrogen, helium and argon, which can prevent the solution containing droplets from contacting air and further reduce the risk of pollution, but will increase the cost.

[0037] Further, the channel can be arranged on the droplet reading chip, such as passing through the droplet reading chip to communicate one end with the B area; or arranged on a sealed sample tube, such as an auxiliary sealing device for sealing the sample tube; or directly arranged on the sample tube.

[0038] Preferably, a device is arranged at one end of the channel which is in communication with the outside, so as to make the end of the channel in an open or closed state. The end is in a closed state when the channel does not exchange gas, which can further reduce the communication between the sealed space and the outside and reduce the risk of pollution. The device can be a piston or a switching valve structure.

[0039] The advantages of the present application are as follows:

[0040] The present application provides a droplet reading chip and a use method, which provides a feasible scheme for high-throughput detection of droplets. The chip of the present application provides a relatively sealed system, which reduces the risk of pollution, and the chip structure of the present application is simple and convenient to use, which can reduce the cost of droplet detection. BRIEF DESCRIPTION OF DRAWINGS

[0041] The features, advantages and technical effects of exemplary embodiments of the present invention will now be described with reference to the accompanying drawings.

[0042] Figure 1 A schematic diagram of a droplet reading chip and its usage method provided in an embodiment of the present invention.

[0043] Figure 2 A droplet reading chip provided in an embodiment of the present invention

[0044] Figure 3 An exploded view of a droplet reading chip provided in an embodiment of the present invention.

[0045] Figure 4 For the appendix Figure 3 Top view of the droplet reading chip

[0046] Figure 5 For the appendix Figure 3 Front view of the droplet reading chip

[0047] Figure 6 A schematic diagram of an auxiliary sealing silicone cap and its application according to an embodiment of the present invention.

[0048] In the diagram: 8-Auxiliary sealing element, 9-Sealed space, 10-Sample tube, 71-Transparent top cover, 72-Microfluidic plate, 73-Aspiration needle, 74-Oil reservoir, 75-Microcavity, 76-Serpentine cavity, 77-Channel, 78-Filter sieve, 711-Pore, 712-Notch, 721-Oil reservoir, 722-Liquid surface spreading groove, 723-Serpentine channel, 724-Inlet, 731-Outlet, 732-Needle, 733-Needle hub, 81-PCR tube groove, 82-Tear groove, 91-Sample solution, 911-Droplet, 912-Continuous phase. Detailed Implementation

[0049] The following detailed description should be read with reference to the accompanying drawings, in which similar elements in different figures are denoted by the same reference numerals. The drawings (which are not necessarily to scale) illustrate illustrative embodiments and are not intended to limit the scope of the invention.

[0050] Figure 1 A droplet reading chip according to the present invention is shown. Figure 1 a. The chip includes a microcavity 75, a sampling needle 73, a liquid reservoir 74, a channel 77, and an auxiliary sealing element 8. The sampling needle 73 and the channel 77 are connected to the same sealed space 9. The sealed space 9 is formed by the auxiliary sealing element 8 and the sample tube 10. The sealed space 9 contains a sample solution 91, and above the sample solution 91 is a gas zone B. Figure 1b) The sample needle 73 penetrates the auxiliary seal 8 to the bottom of the sample solution 91, thus communicating with the closed space 9. The channel 77 penetrates the auxiliary seal 8 to communicate with the B region of the closed space 9. The sample solution 91 includes a droplet 911 and a continuous phase 912.

[0051] In addition, Figure 1 A method of using the chip is shown, which includes the following processes:

[0052] (1) As Figure 1 a, the volume of the droplet 911 is V drop , and the volume of the continuous phase 912 is V oil . The other end of the channel 77 is in communication with the atmosphere.

[0053] (2) As Figure 1 b, a negative pressure is created by drawing air at the end of the channel 77 in communication with the atmosphere, and the gas in the B region is discharged to generate an air pressure Paim of a specified vacuum degree. At this time, if the air pressure in the A region formed by the microcavity 75, the liquid storage cavity 74, and the sample needle 73 is higher than that in the B region, the gas in the A region will be discharged through the B region and then through the channel 77 until the air pressure in the A region is equal to P aim . The air drawing is stopped.

[0054] (3) As Figure 1 c, the channel 77 is again in communication with the atmosphere, and air enters the B region, so that the air pressure in the B region is equal to the external pressure P0.

[0055] (4) As Figure 1 d, the air pressure P0 in the B region is greater than the air pressure P aim in the A region, and under the action of the pressure difference, the sample solution 91 enters the microcavity 75 through the sample needle 73, and the process of entering the sample solution 91 ends when the air pressure in the A region is equal to the air pressure in the B region.

[0056] (5) As Figure 1 e, the most ideal case is that the continuous phase 912 is completely pressed into the liquid storage cavity 74, and the droplet 911 is completely laid in the microcavity 75 to form a layer of laid droplets. The upper surface of the microcavity 75 is transparent to allow light to pass through. The optical detection system is started to perform signal reading and other operations on the laid droplets.

[0057] In this process, the volume of the solution pressed into the A region is V = (Vcc + Vcd + Vcn) * (P0 - P aim ) / P0. Wherein, Vcc is the volume of the liquid storage cavity 74, Vcd is the volume of the microcavity 75, and Vcn is the volume of the sample needle 73. Therefore, by controlling P aim , the amount of solution sucked into the chip can be effectively controlled.

[0058] Figure 2 A droplet reading chip according to the present application is shown, and Figure 1The difference of the chip in a is that a filter screen 78 is added at the channel 77. The filter screen 78 is in a mesh structure for preventing aerosol pollution; it can further be a microporous filter membrane structure with smaller pore size, which allows gas to pass through but does not allow microorganisms such as bacteria to pass through, for example, a 0.22 μm microporous filter membrane, so that the pollution risk can be more effectively reduced.

[0059] Figures 3 to 5 A droplet reading chip according to the present application is shown. As Figure 3 , the chip is composed of three components: a transparent upper cover plate 71, a microfluidic plate 72, and a sample suction needle 73. The transparent upper cover plate 71 is composed of a transparent material, allowing signals such as light to pass through. The microfluidic plate 72 is provided with a liquid storage pool 721, a liquid surface flattening groove 722, and a serpentine channel 723. The transparent upper cover plate 71 and the microfluidic plate 72 are both provided with notches 712 to facilitate their fitting together, and after fitting together, they form a liquid storage cavity 74, a microcavity 75, and a serpentine cavity 76 (as Figure 4 ) in a sealed manner. In use, the liquid storage cavity 74 is used to store the continuous phase; the microcavity 75 is used to flatten the droplets, which are limited in height so that the droplets are flattened into a layer, and the height is 0.5-1.2 times the diameter of the droplets, and the optimal height is the same as the diameter of the droplets; the serpentine cavity 76 is curved and located at the front end of the microcavity 75, which can buffer the bubbles that may enter and trap them in the channel, avoiding the entry of bubbles into the microcavity 75. The sample suction needle 73 includes a base 733 and a needle 732. The sample suction needle 73 is installed below the microfluidic plate 72. The sample outlet 731 of the needle 732 is connected to the sample inlet 724 of the microfluidic plate 72, which is used for the sample solution to enter the serpentine cavity 76 from the sample suction needle; further, the droplets in the sample solution enter the microcavity 75, and the continuous phase enters the liquid storage cavity 74. The needle 732 can be a conical hollow needle, which facilitates the piercing of the auxiliary sealing member to enter the bottom of the sealed space. Through holes 711 are provided on the transparent upper cover plate 71, the microfluidic plate 72, and the sample suction needle 73, and after the three are combined, a channel 77 is formed.

[0060] It should be noted that Figures 3 to 5 the transparent upper cover plate 71, the microfluidic plate 72, and the sample suction needle 73 of the chip are combined to form eight independent cavities (composed of needles, serpentine cavities, microcavities, and liquid storage cavities), which can be used for the detection of eight sample solutions at the same time, and in actual use, the number of cavities can be adjusted as needed.

[0061] Figure 6 An auxiliary sealing member, a silica gel cap, for an implementation method of the fundamental application is shown. The silica gel cap is designed for a PCR tube. As Figure 6 a, the silica gel cap can be clamped at the opening of the PCR tube to seal the PCR tube. As Figure 6b, the silica gel cap 8 comprises a PCR tube slot 25 and a tearing slot 26, the PCR tube slot 26 is used to clamp the PCR tube; the silica gel mold of the tearing slot 26 is relatively thin, such as 100 μm, so that the sample needle can easily enter the PCR tube.

[0062] It should be noted that the method of use of the present application is not limited to Figure 1 In addition to the negative pressure control of droplet extraction shown, the droplet sample can also be directly pressed into the microcavity 75 by pressurization, or a suction device is connected to the microcavity 75, and the droplet sample is sucked into the microcavity 75 by the sample needle 73. The channel 77 for gas inlet and outlet described above can be provided on the device of the present application, or on the device containing the droplet sample or other auxiliary use device. The sample tube 10 described above can be a PCR tube, a 96-well plate, a 384-well plate, a centrifuge tube, a liquid phase vial and other containers for storing solutions.

[0063] It should be understood that the above embodiments can be appropriately changed or modified by those skilled in the art without departing from the spirit and scope of the present application. For example, the relative positions of the liquid storage cavity 74, the microcavity 75 and the serpentine cavity 76 can be appropriately adjusted, for example, the liquid storage cavity 74 can be located at the top end of the microcavity 75. The droplet can be O / W type, and the liquid storage cavity 74 is used to store the oil phase; the droplet can also be W / O type, and the liquid storage cavity 74 is used to store the water phase.

[0064] The above is only used to illustrate the technical idea and characteristics of the present application, and cannot limit the protection scope of the present application. Any modification made on the basis of the technical solution according to the technical idea of the present application is within the protection scope of the present application. The technology not involved in the present application can be realized by the existing technology.

Claims

1. A droplet reader chip, characterized by, It is composed of transparent upper cover plate, microfluidic plate, sample needle and auxiliary sealing piece; the microfluidic plate is provided with liquid storage pool, liquid surface flat slot and serpentine channel; the transparent upper cover plate and the microfluidic plate are embedded and sealed to form liquid storage cavity, microcavity and serpentine cavity; the microcavity is used for flat liquid drop; the microcavity is connected with the serpentine cavity; the microcavity is connected with the liquid storage cavity; the sample needle is used for sucking liquid drop into the microcavity; the sample needle is communicated with the microcavity and can exchange fluid; the transparent upper cover plate, the microfluidic plate and the base are provided with hole for forming the channel after combination; the channel is not communicated with the microcavity; the auxiliary sealing piece is provided with sealing groove and tearing groove; The use method of the liquid drop reading chip is composed of the following steps: Step (1): the solution containing liquid drop is located in a closed sample tube to form a closed space, the upper end of the closed space is a gas zone, recorded as B zone, the B zone is communicated with a channel, one end of the channel is communicated with the B zone, the other end is communicated with the outside, the gas exchange between the closed space and the outside can be carried out through the channel; the sample needle enters the solution containing liquid drop; Step (2): the gas in the B zone is discharged through the channel, if there is gas in the microcavity, it will be discharged through the sample needle, the B zone and the channel, at this time, the gas pressure in the microcavity is recorded as Paim; Step (3): the gas enters the B zone through the channel, at this time, the gas pressure in the B zone is P0; Step (4): the P0 is higher than the Paim, the negative pressure is formed in the microcavity, the solution containing liquid drop enters the microcavity through the sample needle, until the gas pressure in the B zone and the microcavity is equal; Step (5): the chip is placed in the optical detection system, and the liquid drop signal in the microcavity is read.

2. The droplet reader chip of claim 1, wherein, When the channel and the sample needle are connected with the same closed space, the channel can make the solution containing liquid drop in the closed space enter the microcavity through the sample needle by flowing with the outside gas; the channel can discharge the gas in the closed space and the microcavity; the gas is input into the closed space through the channel, the pressure difference is formed between the microcavity and the closed space, under the action of the pressure difference, the solution containing liquid drop in the closed space enters the microcavity through the sample needle; the microcavity allows the liquid drop to be flat; the channel is provided with filter screen; the sample needle includes base and needle, the base is used for embedding with the microfluidic plate; one end of the sample needle is immersed in the solution containing liquid drop, one end of the channel is not immersed in the solution containing liquid drop; the other end of the sample needle is communicated with the microcavity, the other end of the channel is communicated with the outside; the closed space is formed by the sealed sample tube, the sample tube includes the solution containing liquid drop, and there is a gas zone above the solution; the number of the microcavity and the sample needle is not less than 1; the sample needle includes conical hollow needle; the auxiliary sealing piece is silica gel cap.

3. The droplet reader chip of claim 1, wherein, The amount of the solution containing the droplets into the microcavity is controlled by controlling the P0 and Paim to form a specific negative pressure in the microcavity, wherein the P0 is the atmospheric pressure; and the gas in step (3) includes air, nitrogen, helium and argon.

4. A method of using a droplet reading chip, characterized by, The method for using the droplet reading chip according to any one of claims 1-3.

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