Digital MIRA chip, discretization device and discretization method

Abstract

The invention belongs to the technical field of micro-fluidic chips, and particularly provides a digital MIRA chip, a discretization device and a discretization method.According to the digital MIRA chip, highly-asymmetric contact angle composite interfaces are constructed on the upper surface and the lower surface of a micro-bin module of a structural layer and a micro-well array module, the upper surface is designed to be a high contact angle, and the lower surface is designed to be a high contact angle; the spreadability of the reagent is weakened, the flowing of the reagent in the horizontal direction is delayed, the lower surface and the micro-well array module are designed to have low contact angles, and the retention of the reagent at an inlet of the micro-well array module is reduced, so that the reagent is promoted to smoothly enter a micro-well below, and the problems that the reagent is blocked to flow in the micro-bin module and is difficult to enter the micro-well array module are solved; the reagent can be quickly and uniformly distributed in each micro-well of the micro-well array module, so that the absolute quantitative analysis accuracy of the reagent is ensured.

Description

Technical Field

[0001] The present invention belongs to the technical field of microfluidic chips, and in particular relates to a digital MIRA chip, a discrete device and a discrete method. Background Art

[0002] Multienzyme Isothermal Rapid Amplification (MIRA) technology is a isothermal rapid nucleic acid amplification technology that relies on the synergistic action of multiple functional proteins (helicase, recombinase, single-stranded binding protein, DNA polymerase, etc.) to achieve rapid nucleic acid amplification at room temperature.

[0003] Traditional MIRA reactions are usually carried out in macroscopic reaction systems, and it is difficult to achieve digital nucleic acid amplification on microfluidic platforms, which limits its application in high-throughput and precision detection. The development of microfluidic technology has provided a new approach for nucleic acid amplification. By reducing the reaction system to the nanoliter level, high-sensitivity detection at the single-molecule level can be achieved, and the reaction is faster and more economical.

[0004] In the digital MIRA chip in the related art, due to the high viscosity and interface characteristics of the reagents, it is difficult to evenly distribute the reagents into each microwell, and droplets cannot be stably generated, which affects the accuracy and consistency of the quantitative results. Therefore, how to improve the fluidity and discreteness of the reagents and achieve accurate quantitative analysis is currently an urgent problem to be solved.

[0005] It should be noted that the above information disclosed in this background technology section is only used to understand the background technology of the present application concept, and therefore, the above description is not considered to constitute information of the prior art. Summary of the invention

[0006] The embodiments of the present disclosure at least provide a digital MIRA chip, a discrete device and a discrete method.

[0007] In a first aspect, an embodiment of the present disclosure provides a digital MIRA chip, comprising: A top glass slide, a structural layer, and a bottom glass slide are sequentially arranged from top to bottom; The structural layer includes a micro-warehouse module and a micro-well array module; The micro-warehouse module is suitable for filling reagents into the micro-well array module; The upper surface and lower surface of the micro-warehouse module are highly asymmetric contact angle composite interfaces, wherein the upper surface of the micro-warehouse module is a high contact angle, the lower surface of the micro-warehouse module is a low contact angle, and the surface of the micro-well array module is a low contact angle.

[0008] In an optional embodiment, the micro-warehouse module includes: Micro-warehouse channel, support column, reagent inlet and reagent outlet; The reagent inlet and the reagent outlet are respectively arranged on both sides of the micro-warehouse channel; The support column is arranged in the micro-warehouse channel and is suitable for supporting the micro-warehouse channel.

[0009] In an optional embodiment, the lower surface of the micro-warehouse channel and the micro-well array module are made of hydrophilic material; The upper surface of the micro-warehouse channel is made of hydrophobic material.

[0010] In an optional embodiment, the surface of the micro-well array module, the lower surface and the upper surface of the micro-warehouse channel are constructed with a surface treatment method to have a highly asymmetric contact angle composite interface, wherein the surface of the micro-well array module and the lower surface of the micro-warehouse channel are hydrophilic, and the upper surface of the micro-warehouse channel is hydrophobic.

[0011] In an optional embodiment, the surface treatment method is one of chemical vapor deposition, PDMS doped with surfactant, self-assembled monolayer technology or coating, or a combination of two or more thereof.

[0012] In an optional embodiment, the top slide is provided with a sample inlet and a sample outlet; The reagent inlet is connected to the sample inlet; The reagent outlet is communicated with the sample outlet.

[0013] In an optional embodiment, the structural layer is connected to the top glass slide and the bottom glass slide respectively by bonding.

[0014] In an optional embodiment, the bonding method is any one of heat-resistant transparent tape, thermal bonding, heat-resistant adhesive bonding, hot pressure sealing, and air plasma treatment bonding, or a combination of two or more of them.

[0015] In a second aspect, an embodiment of the present disclosure further provides a discrete device, including: Positive pressure generating module, negative pressure generating module, air pressure sensor and digital MIRA chip as described above; The positive pressure generating module is connected to the injection port of the digital MIRA chip through a first hose; The negative pressure generating module is connected to the sample outlet of the digital MIRA chip through a second hose; The first hose is provided with a first pinch valve; The second hose is provided with a second pinch valve and an air pressure sensor.

[0016] In a third aspect, the present disclosure also provides a discretization method for the discretization device as described above, the discretization method comprising: The reagent and the sealing oil are sucked into the first hose in sequence; Connecting the positive pressure generating module, the negative pressure generating module, the first hose and the second hose to the digital MIRA chip; Open the first pinch valve and the second pinch valve; Start the negative pressure generating module to reduce the air pressure in the digital MIRA chip to the preset negative pressure; Start the positive pressure generating device, and inject the reagent and sealing oil into the digital MIRA chip in the order of reagent first and sealing oil later; After the sealing oil enters the second hose, the first pinch valve and the second pinch valve are closed to complete the dispersion.

[0017] The beneficial effect of the present invention is that the digital MIRA chip, discrete device and discrete method construct a highly asymmetric contact angle composite interface on the upper and lower surfaces of the micro-warehouse module and the micro-well array module of the structural layer. The upper surface is designed to have a high contact angle to reduce the spreadability of the reagent and delay the flow of the reagent in the horizontal direction. The lower surface and the micro-well array module are designed to have a low contact angle to reduce the retention of the reagent at the entrance of the micro-well array module, thereby promoting the reagent to smoothly enter the micro-well below, solving the problem of obstructed flow of the reagent in the micro-warehouse module and difficulty in entering the micro-well array module, ensuring that the reagent can be quickly and evenly distributed in each micro-well of the micro-well array module, thereby ensuring the accuracy of the absolute quantitative analysis of the reagent.

[0018] Other features and advantages of the present invention will be described in the following description, and partly become apparent from the description, or understood by practicing the present invention. The purpose and other advantages of the present invention are realized and obtained by the structures particularly pointed out in the description, claims and drawings.

[0019] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, this article specifically cites preferred embodiments and provides detailed descriptions as follows in conjunction with the attached drawings. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the specific implementation methods of the present invention or the technical solutions in the prior art, the drawings required for use in the specific implementation methods or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are some implementation methods of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying creative work.

[0021] Figure 1 An exploded diagram of a digital MIRA chip provided by an embodiment of the present disclosure; Figure 2 A schematic diagram of the structure of the structural layer provided in the embodiment of the present disclosure; Figure 3 Actual discrete schematic diagram of a digital MIRA chip for a highly asymmetric contact angle composite interface provided by an embodiment of the present disclosure; Figure 4 An actual discrete schematic diagram of a digital MIRA chip with hydrophobic contact angles provided in an embodiment of the present disclosure; Figure 5 An actual discrete schematic diagram of a digital MIRA chip with hydrophilic contact angles provided in an embodiment of the present disclosure; Figure 6 A schematic diagram of the structure of a discrete device provided in an embodiment of the present disclosure; Figure 7 A flowchart of a discrete method provided in an embodiment of the present disclosure.

[0022] In the figure: 100, digital MIRA chip; 110, top glass slide; 111, sample inlet; 112, sample outlet; 120, structural layer; 121, micro-warehouse module; 1211, micro-warehouse channel; 1212, support column; 1213, reagent inlet; 1214, reagent outlet; 122, micro-well array module; 130, bottom glass slide; 200, positive pressure generating module; 210, first hose; 220, first pinch valve; 300, negative pressure generating module; 310, second hose; 320, second pinch valve; 330, air pressure sensor. DETAILED DESCRIPTION

[0023] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution of the present invention will be clearly and completely described below in conjunction with the accompanying drawings. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0024] It should be noted that similar reference numerals and letters represent similar items in the following drawings, so once an item is defined in one drawing, it does not need to be further defined and explained in the subsequent drawings. In addition, in the drawings, the thickness of the components may be exaggerated or reduced in order to effectively describe the technical content.

[0025] Some embodiments of the present invention are described in detail below in conjunction with the accompanying drawings. In the absence of conflict, the following embodiments and features in the embodiments can be combined with each other.

[0026] See also Figure 1 , Figure 1An exploded view of the digital MIRA chip 100 is shown, which includes a top glass slide 110, a structural layer 120 and a bottom glass slide 130 arranged in sequence from top to bottom; the structural layer 120 includes a micro-warehouse module 121 and a micro-well array module 122; the micro-warehouse module 121 is suitable for filling reagents into the micro-well array module 122; the upper surface and the lower surface of the micro-warehouse module 121 are highly asymmetric contact angle composite interfaces, wherein the upper surface of the micro-warehouse module 121 has a high contact angle, and the lower surface of the micro-warehouse module 121 has a low contact angle. A highly asymmetric contact angle composite interface is constructed on the upper and lower surfaces of the micro-warehouse module 121 of the structural layer and the surface of the micro-well array module 122. The upper surface is designed to have a high contact angle to reduce the spreading of the reagent and delay the horizontal flow of the reagent. The lower surface is designed to have a low contact angle to reduce the retention of the reagent at the entrance of the micro-well array module 122, thereby facilitating the reagent to smoothly enter the micro-well below, solving the problem of the reagent being obstructed in the flow of the micro-warehouse module 121 and having difficulty entering the micro-well array module 122, ensuring that the reagent can be quickly and evenly distributed in each micro-well of the micro-well array module 122, thereby ensuring the accuracy of the absolute quantitative analysis of the reagent.

[0027] Figure 3 In the figure, a represents the upper surface of the micro-warehouse module 121 , b represents the lower surface of the micro-warehouse module 121 , and c represents the surface of the micro-well array module 122 .

[0028] The micro-well array module 122 includes a plurality of micro-wells.

[0029] In some embodiments, the top glass slide 110 and the bottom glass slide 130 are made of incompressible, airtight and thermally conductive transparent materials, such as glass, silicon wafer, and polydimethylsiloxane. The material of the structural layer 120 is made of a compressible, internally loose and porous transparent material, such as polydimethylsiloxane.

[0030] The top glass slide 110 and the bottom glass slide 130 are both made of materials with good thermal conductivity, and the top glass slide 110 and the bottom glass slide 130 can be used to control the temperature rise and fall of the reagent sample in the micro-well array module 122 in the structural layer 120 .

[0031] In at least one embodiment, the structural layer 120 is connected to the top glass slide 110 and the bottom glass slide 130 respectively by bonding. The bonding method is any one of heat-resistant transparent tape, thermal bonding, heat-resistant adhesive bonding, hot press sealing, air plasma treatment bonding, or a combination of two or more thereof.

[0032] See also Figure 1 and Figure 2In at least one embodiment, the micro-warehouse module 121 includes: a micro-warehouse channel 1211, a support column 1212, a reagent inlet 1213 and a reagent outlet 1214; the reagent inlet 1213 and the reagent outlet 1214 are respectively arranged on both sides of the micro-warehouse channel 1211; the support column 1212 is arranged in the micro-warehouse channel 1211 and is suitable for supporting the micro-warehouse channel 1211. The structural strength of the micro-warehouse channel 1211 is improved by the support column 1212 to prevent the micro-warehouse channel 1211 from being damaged in a negative pressure environment.

[0033] In a preferred embodiment, the lower surface of the micro-warehouse channel 1211 and the micro-well array module 122 are made of hydrophilic materials; the upper surface of the micro-warehouse channel 1211 is made of hydrophobic materials. Figure 3 shown.

[0034] When the contact angles of the upper and lower surfaces of the micro-tank channel 1211 are both hydrophobic, it is difficult for the reagent to fill the small-sized micro-wells due to the presence of surface tension. Figure 4 As shown; when the contact angles of the upper and lower surfaces of the micro-warehouse channel 1211 are both hydrophilic, the reagent swings along the channel surfaces on both sides of the micro-well, thereby trapping the air in the micro-well and being unable to fill the micro-well, such as Figure 5 shown.

[0035] Depend on Figure 3-Figure 5 It can be seen that the highly asymmetric contact angle composite interface can effectively accelerate the filling of the micro-wells and delay the flow of reagents along the upper surface of the micro-tank, thereby improving the filling performance of the reagents and making the discreteness more accurate and efficient.

[0036] In an optional embodiment, the surface of the micro-well array module (122), the lower surface and the upper surface of the micro-warehouse channel 1211 are constructed by surface treatment methods to form a highly asymmetric contact angle composite interface, wherein the surface of the micro-well array module (122) and the lower surface of the micro-warehouse channel 1211 are hydrophilic, and the upper surface of the micro-warehouse channel 1211 is hydrophobic.

[0037] Specifically, the surface treatment method is one of chemical vapor deposition, PDMS doped with surfactant, self-assembled monolayer technology or coating, or a combination of two or more of them.

[0038] In an optional embodiment, the top glass slide 110 is provided with a sample inlet 111 and a sample outlet 112 ; the reagent inlet 1213 is communicated with the sample inlet 111 ; and the reagent outlet 1214 is communicated with the sample outlet 112 .

[0039] See also Figure 6 The embodiment of the present disclosure also provides a discrete device, including: a positive pressure generating module 200, a negative pressure generating module 300, an air pressure sensor 330 and the digital MIRA chip 100 as described above; the positive pressure generating module 200 is connected to the sample inlet 111 of the digital MIRA chip 100 through a first hose 210; the negative pressure generating module 300 is connected to the sample outlet 112 of the digital MIRA chip 100 through a second hose 310; a first pinch valve 220 is provided on the first hose 210; a second pinch valve 320 and an air pressure sensor 330 are provided on the second hose 310.

[0040] See also Figure 7 The embodiment of the present disclosure further provides a discretization method for the discretization device as described above, the discretization method comprising: S110 : The reagent and the sealing oil are sucked into the first hose 210 in sequence.

[0041] S120 : Connecting the positive pressure generating module 200 , the negative pressure generating module 300 , the first hose 210 , and the second hose 310 to the digital MIRA chip 100 .

[0042] S130: Open the first pinch valve 220 and the second pinch valve 320.

[0043] S140: Start the negative pressure generating module 300 to reduce the air pressure in the digital MIRA chip 100 to a preset negative pressure.

[0044] Among them, the preset negative pressure is -30kPa.

[0045] S150: Start the positive pressure generating device, and inject the reagent and the sealing oil into the digital MIRA chip 100 in sequence in the order of the reagent in front and the sealing oil in the back.

[0046] S160: After the sealing oil enters the second hose 310, the first pinch valve 220 and the second pinch valve 320 are closed to complete the dispersion.

[0047] In summary, the digital MIRA chip 100, the discrete device and the discrete method construct a highly asymmetric contact angle composite interface on the upper and lower surfaces of the micro-warehouse module 121 of the structural layer and the surface of the micro-well array module 122. The upper surface is designed to have a high contact angle to reduce the spread of the reagent and delay the horizontal flow of the reagent. The lower surface is designed to have a low contact angle to reduce the retention of the reagent at the entrance of the micro-well array module 122, thereby facilitating the smooth entry of the reagent into the micro-well below, solving the problem of the reagent being obstructed in the flow of the micro-warehouse module 121 and having difficulty entering the micro-well array module 122, ensuring that the reagent can be quickly and evenly distributed in each micro-well of the micro-well array module 122, thereby ensuring the accuracy of the absolute quantitative analysis of the reagent.

[0048] Spatially relative terms, such as "inside", "outside", "below", "below", "down", "above", "on", etc., may be used herein to facilitate description of the relationship of one element or feature to another element or feature as illustrated in the figure. In addition to the orientation depicted in the figure, spatially relative terms may be intended to cover different orientations of the device in use or operation. For example, if the device in the figure is turned over, the elements described as "below" or "below" other elements or features will be oriented to be "above" other elements or features. Therefore, the example term "below" can cover the orientation above and below. The device can be oriented in other ways (rotated 90 degrees or in other orientations), and the spatially relative descriptors used herein are interpreted accordingly.

[0049] Based on the above ideal embodiments of the present invention, the relevant staff can make various changes and modifications without departing from the technical concept of the present invention through the above description. The technical scope of the present invention is not limited to the contents of the specification, and its technical scope must be determined according to the scope of the claims.

Claims

1. A digital MIRA chip, characterized in that: include: A top glass slide (110), a structural layer (120), and a bottom glass slide (130) are sequentially arranged from top to bottom; The structural layer (120) comprises a micro-warehouse module (121) and a micro-well array module (122); The micro-tank module (121) is suitable for filling the reagent into the micro-well array module (122); The upper surface and the lower surface of the micro-warehouse module (121) are highly asymmetric contact angle composite interfaces, wherein the upper surface of the micro-warehouse module (121) has a high contact angle, the lower surface of the micro-warehouse module (121) has a low contact angle, and the surface of the micro-well array module (122) has a low contact angle.

2. The digital MIRA chip according to claim 1, characterized in that: The micro-warehouse module (121) comprises: A micro-warehouse channel (1211), a support column (1212), a reagent inlet (1213), and a reagent outlet (1214); The reagent inlet (1213) and the reagent outlet (1214) are respectively arranged on both sides of the micro-warehouse channel (1211); The support column (1212) is arranged in the micro-warehouse channel (1211), and is suitable for supporting the micro-warehouse channel (1211).

3. The digital MIRA chip as claimed in claim 2, characterized in that: The lower surface of the micro-warehouse channel (1211) and the micro-well array module (122) are made of hydrophilic material; The upper surface of the micro-warehouse channel (1211) is made of a hydrophobic material.

4. The digital MIRA chip as claimed in claim 2, characterized in that: The surface of the micro-well array module (122), the lower surface and the upper surface of the micro-warehouse channel (1211) are constructed with a highly asymmetric contact angle composite interface using a surface treatment method, wherein the surface of the micro-well array module (122) and the lower surface of the micro-warehouse channel (1211) are hydrophilic, and the upper surface of the micro-warehouse channel (1211) is hydrophobic.

5. The digital MIRA chip as claimed in claim 4, characterized in that: The surface treatment method is one of chemical vapor deposition, PDMS doped with surfactant, self-assembled monolayer technology or coating, or a combination of two or more of them.

6. The digital MIRA chip as claimed in claim 2, characterized in that: The top glass slide (110) is provided with a sample inlet (111) and a sample outlet (112); The reagent inlet (1213) is in communication with the sample inlet (111); The reagent outlet (1214) is in communication with the sample outlet (112).

7. The digital MIRA chip (100) according to claim 1, characterized in that: The structural layer (120) is respectively connected to the top glass slide (110) and the bottom glass slide (130) by bonding.

8. The digital MIRA chip (100) according to claim 7, characterized in that: The bonding method is any one of heat-resistant transparent tape, thermal bonding, heat-resistant adhesive bonding, hot pressing sealing, and air plasma treatment bonding, or a combination of two or more thereof.

9. A discrete device, characterized in that: include: A positive pressure generating module (200), a negative pressure generating module (300), an air pressure sensor (330), and a digital MIRA chip (100) as claimed in any one of claims 1 to 8; The positive pressure generating module (200) is connected to the injection port (111) of the digital MIRA chip (100) via a first hose (210); The negative pressure generating module (300) is in communication with the sample outlet (112) of the digital MIRA chip (100) via a second hose (310); The first hose (210) is provided with a first pinch valve (220); The second hose (310) is provided with a second pinch valve (320) and an air pressure sensor (330).

10. A method for discrete devices according to claim 9, characterized in that: The discrete method includes: The reagent and the sealing oil are sucked into the first hose (210) in sequence; Connecting the positive pressure generating module (200), the negative pressure generating module (300), the first hose (210), and the second hose (310) to the digital MIRA chip (100); Opening the first pinch valve (220) and the second pinch valve (320); Starting the negative pressure generating module (300) to reduce the air pressure in the digital MIRA chip (100) to a preset negative pressure; The positive pressure generating device is started, and the reagent and the sealing oil are sequentially poured into the digital MIRA chip (100) in the order of the reagent first and the sealing oil later; After the sealing oil enters the second hose (310), the first pinch valve (220) and the second pinch valve (320) are closed to complete the separation.

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