Detection chip and preparation method and sample injection method thereof

Abstract

The application discloses a detection chip, wherein the detection chip is divided into at least one functional area (1), the functional area (1) comprises a reaction area (101) and a non-reaction area (102) surrounding the reaction area (101), and the detection chip comprises oppositely arranged first and second substrates (9 and 10). The side of the first substrate (9) facing the second substrate (10) is provided with a plurality of reaction grooves (4) arranged in an array along a first direction (X) and a second direction (Y) in the reaction area (101). A first flow-through groove (5) connected with two adjacent reaction grooves (4) in the first direction (X) is arranged between the two reaction grooves (4), and the first flow-through groove (5) extends along the first direction (X). A second flow-through groove (6) connected with two adjacent first flow-through grooves (5) in the second direction (Y) is arranged between the two first flow-through grooves (5), and the second flow-through groove (6) extends along the second direction (Y). The first direction (X) intersects with the second direction (Y). A detection method and a sample injection method of the detection chip are also provided.

Description

Technical Field

[0001] The embodiments disclosed herein relate to a detection chip and its fabrication and injection methods. Background Technology

[0002] Digital polymerase chain reaction (DPCR) is a rapidly developing third-generation nucleic acid molecule quantitative analysis technology. Its principle involves evenly distributing a sample into tens of thousands of different reaction units, each containing at least one copy of the target DNA template. PCR amplification is then performed separately in each reaction unit, and the fluorescence signals from each unit are statistically analyzed after amplification. This technology is independent of standard curves, less affected by amplification efficiency, and boasts excellent accuracy and reproducibility, enabling absolute quantitative analysis. It demonstrates significant technological advantages in nucleic acid detection and identification research. Compared to traditional real-time quantitative PCR, it is particularly suitable for copy number variation, rare mutation detection and genotyping, NGS verification, and single-cell expression analysis.

[0003] Currently, digital PCR is mainly implemented in two forms: array-based and droplet-based. Among them, array-based digital PCR detection chips generate more uniform microreaction volumes, have higher stability, and less influence between systems compared to droplet-based chips, which is more conducive to obtaining highly accurate analytical results. However, the fabrication of microarrays is relatively complex, and the process of sample solution introduction into the detection chip, that is, the process of sample solution entering each microreaction chamber, is often inefficient. The sample solution cannot smoothly fill the entire chamber, resulting in uneven distribution of the sample solution in each microreaction chamber, which directly affects amplification efficiency and result interpretation, thus limiting the application of array-based digital PCR detection chips. Summary of the Invention

[0004] In a first aspect, embodiments of this disclosure provide a detection chip, wherein at least one functional region is defined, the functional region including: a reaction region and a non-reaction region surrounding the reaction region, the detection chip including: a first substrate and a second substrate disposed opposite to each other, the first substrate having a plurality of reaction slots arranged in an array along a first direction and a second direction on a side facing the second substrate and located within the reaction region;

[0005] A first flow channel is provided between two adjacent reaction channels in the first direction, and the first flow channel extends along the first direction.

[0006] A second flow groove connected to the two first flow grooves is provided between two adjacent first flow grooves in the second direction, and the second flow groove extends along the second direction.

[0007] The first direction intersects with the second direction.

[0008] In some embodiments, the width of the first flow channel is greater than the width of the second flow channel.

[0009] In some embodiments, the depth of the reaction tank is greater than or equal to the depth of the second flow tank.

[0010] In some embodiments, the first flow channel includes: a first portion, a second portion, and a third portion connected sequentially along a first direction, wherein the first portion and the third portion are respectively connected to two adjacent reaction channels, and the second portion is connected to the second flow channel;

[0011] The depth of the first part and the depth of the third part are both greater than or equal to the depth of the second flow channel;

[0012] The depth of the second part is equal to the depth of the second flow channel.

[0013] In some embodiments, a first liquid inlet and a first liquid outlet penetrating the second substrate are provided on the second substrate and located in the non-reactive region;

[0014] A plurality of reaction tanks and a plurality of first flow tanks arranged alternately in a first direction constitute a first flow channel, and the two ends of the first flow channel are respectively connected to the first liquid inlet and the first liquid outlet.

[0015] In some embodiments, the first inlet and the first outlet are located on opposite sides of the reaction region in the first direction.

[0016] In some embodiments, the line connecting the center of the first inlet and the center of the first outlet extends along a first direction and passes through the center of the reaction region.

[0017] In some embodiments, a first liquid inlet connection groove and a first liquid outlet connection groove corresponding to the first flow channel are further provided on the side of the first substrate facing the second substrate.

[0018] One end of the first liquid inlet connection groove is connected to one end of the corresponding first flow channel, and the other end of the first liquid inlet connection groove extends to the non-reaction area and is connected to the first liquid inlet.

[0019] One end of the first liquid outlet connection groove is connected to the other end of the corresponding first flow channel, and the other end of the first liquid inlet connection groove extends to the non-reaction area and is connected to the first liquid outlet.

[0020] In some embodiments, a second liquid inlet and a second liquid outlet are formed on the second substrate and located in the non-reactive region, penetrating the second substrate;

[0021] The second flow channels arranged in the second direction constitute a second flow channel, and the two ends of the second flow channel are respectively connected to the second liquid inlet and the second liquid outlet.

[0022] In some embodiments, the second inlet and the second outlet are located on opposite sides of the reaction region in the second direction.

[0023] In some embodiments, the line connecting the center of the second inlet and the center of the second outlet extends along a second direction and passes through the center of the reaction region.

[0024] In some embodiments, a second liquid inlet connection groove and a second liquid outlet connection groove corresponding to the second flow channel are further provided on the side of the first substrate facing the second substrate.

[0025] One end of the second liquid inlet connection groove is connected to one end of the corresponding second flow channel, and the other end of the second liquid inlet connection groove extends to the non-reaction area and is connected to the second liquid inlet.

[0026] One end of the second liquid outlet connection groove is connected to the other end of the corresponding second flow channel, and the other end of the second liquid inlet connection groove extends to the non-reaction area and is connected to the second liquid outlet.

[0027] In some embodiments, the width of the first flow channel ranges from 20µm to 30µm;

[0028] The width of the second flow channel ranges from 10µm to 20µm.

[0029] In some embodiments, the first substrate includes: a substrate and a hole defining layer located on the side of the substrate facing the second substrate;

[0030] The hole limiting layer has a first hole structure in the area where the first flow groove is to be formed, and the first flow groove includes the first hole structure.

[0031] The hole limiting layer has a second hole structure in the area where the second flow groove is to be formed, and the second flow groove includes the second hole structure.

[0032] A third hole structure is provided on the hole-defining layer in the area where the reaction tank is to be formed, and the reaction tank includes the third hole structure.

[0033] In some embodiments, when a first liquid inlet connection groove and a first liquid outlet connection groove are provided on the side of the first substrate facing the second substrate, a fourth hole structure is provided on the hole limiting layer in the area where the first liquid inlet connection groove is to be formed, and a fifth hole structure is provided on the hole limiting layer in the area where the first liquid outlet connection groove is to be formed. The first liquid inlet connection groove includes the fourth hole structure, and the first liquid outlet connection groove includes the fifth hole structure.

[0034] In some embodiments, when a second liquid inlet connection groove and a second liquid outlet connection groove are provided on the side of the first substrate facing the second substrate, a sixth hole structure is provided on the hole limiting layer in the area where the second liquid inlet connection groove is to be formed, and a seventh hole structure is provided on the hole limiting layer in the area where the second liquid outlet connection groove is to be formed. The second liquid inlet connection groove includes the sixth hole structure, and the second liquid outlet connection groove includes the seventh hole structure.

[0035] In some embodiments, a heating electrode is disposed between the substrate and the hole defining layer, the heating electrode being configured to heat the region where the reaction tank is located.

[0036] In some embodiments, a control electrode is disposed between the heating electrode and the substrate, a first insulating layer is disposed between the control electrode and the heating electrode, the control electrode is connected to the heating electrode through a via on the first insulating layer, and the control electrode is configured to apply an electrical signal to the heating electrode.

[0037] In some embodiments, a second insulating layer is provided between the heating electrode and the hole defining layer, and a light-shielding layer is provided between the second insulating layer and the hole defining layer. A hollow structure is provided on the light-shielding layer in the area where the reaction tank is to be formed.

[0038] The reaction tank also includes the hollow structure.

[0039] In some embodiments, the substrate is provided with a first receiving groove on the side facing the hole defining layer and in the region where the reaction tank is to be formed;

[0040] The reaction tank also includes the first receiving tank.

[0041] In some embodiments, the first flow channel includes: a first portion, a second portion, and a third portion connected sequentially along a first direction, wherein the first portion and the third portion are respectively connected to two adjacent reaction channels, and the second portion is connected to the second flow channel;

[0042] A second receiving groove is provided on the substrate in the region where the first part is to be formed, and a third receiving groove is provided on the substrate in the region where the third part is to be formed. Both the second receiving groove and the third receiving groove are connected to the corresponding first receiving groove.

[0043] The first flow channel also includes the second receiving channel and the third receiving channel.

[0044] In some embodiments, a light-shielding layer is provided between the substrate and the hole-defining layer, and a hollow structure is provided on the light-shielding layer in the area where the reaction tank is to be formed.

[0045] The reaction tank also includes the hollow structure.

[0046] In some embodiments, the second substrate includes a cover plate and a heating electrode located on the side of the cover plate facing the first substrate, the heating electrode being configured to heat the region where the reaction tank is located.

[0047] In some embodiments, a first protective layer is provided on the side of the heating electrode facing away from the cover plate.

[0048] In some embodiments, a control electrode is disposed between the heating electrode and the cover plate, and a first insulating layer is disposed between the control electrode and the heating electrode. The control electrode is connected to the heating electrode through a via on the first insulating layer, and the control electrode is configured to apply an electrical signal to the heating electrode.

[0049] In some embodiments, the material of the hole defining layer includes photoresist.

[0050] In some embodiments, a hydrophilic layer is provided on the bottom of the reaction tank, the sidewall of the reaction tank, the bottom of the first flow channel, and / or the sidewall of the first flow channel.

[0051] In some embodiments, a hydrophobic layer is provided at the bottom of the second flow channel and / or on the sidewall of the second flow channel.

[0052] In some embodiments, the number of functional areas is multiple.

[0053] Secondly, embodiments of this disclosure also provide a method for fabricating a detection chip as described in the first aspect, wherein the detection chip is divided into at least one functional region, the functional region comprising: a reactive region and a non-reactive region surrounding the reactive region, and the fabrication method comprising:

[0054] A first substrate and a second substrate are prepared respectively. A plurality of reaction cells are arranged in an array along a first direction and a second direction on one side of the first substrate. A first flow channel connected to the two reaction cells is provided between two adjacent reaction cells in the first direction. The first flow channel extends along the first direction. A second flow channel connected to the two first flow channels is provided between two adjacent first flow channels in the second direction. The second flow channel extends along the second direction. The first direction and the second direction intersect.

[0055] The first substrate, which has the reaction tank, the first flow tank and the second flow tank, is positioned opposite the second substrate, and the first substrate and the second substrate are then encapsulated.

[0056] Thirdly, embodiments of this disclosure also provide a sample introduction method for the detection chip as described in the first aspect, comprising:

[0057] The sample solution is injected into the reaction tank through the first flow channel;

[0058] An oil phase is injected into the second flow channel to isolate the reaction tanks from the oil phase. Attached Figure Description

[0059] Figure 1 This is a schematic diagram of the structure of a detection chip provided in an embodiment of the present disclosure;

[0060] Figure 2 A top view schematic diagram of the detection chip provided in an embodiment of this disclosure;

[0061] Figure 3 This is a top view of the reaction area in an embodiment of this disclosure;

[0062] Figure 4 for Figure 1 A schematic diagram of a cross section along line A-A';

[0063] Figure 5 for Figure 1 A schematic diagram of a cross section along the B-B' direction;

[0064] Figure 6 for Figure 1 Another cross-sectional diagram along the A-A' direction;

[0065] Figure 7 for Figure 1 Another cross-sectional diagram along the B-B' direction;

[0066] Figure 8 for Figure 1 Another cross-sectional diagram along the A-A' direction;

[0067] Figure 9 for Figure 1 Another cross-sectional diagram along the B-B' direction;

[0068] Figure 10 This is a schematic diagram of a portion of the detection chip in an embodiment of this disclosure;

[0069] Figure 11 for Figure 10 A top view of a partial area of ​​the substrate;

[0070] Figure 12 This is a schematic diagram of another structure of the detection chip provided in an embodiment of this disclosure;

[0071] Figure 13 A flowchart illustrating a method for fabricating a detection chip according to an embodiment of this disclosure;

[0072] Figure 14 This is a flowchart of a sample introduction method for a detection chip provided in an embodiment of this disclosure. Detailed Implementation

[0073] To enable those skilled in the art to better understand the technical solutions of this disclosure, a detection chip, its preparation method, and its sample introduction method provided in this disclosure will be described in detail below with reference to the accompanying drawings.

[0074] To make the objectives, technical solutions, and advantages of the embodiments of this disclosure clearer, the technical solutions of the embodiments of this disclosure will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of this disclosure. All other embodiments obtained by those skilled in the art based on the described embodiments of this disclosure without creative effort are within the scope of protection of this disclosure.

[0075] Unless otherwise defined, the technical or scientific terms used herein should have the ordinary meaning understood by one of ordinary skill in the art to which this disclosure pertains. The terms “first,” “second,” and similar terms used in this disclosure do not indicate any order, quantity, or importance, but are merely used to distinguish different components. Similarly, terms such as “comprising” or “including” mean that the element or object preceding the word encompasses the elements or objects listed following the word and their equivalents, without excluding other elements or objects. Terms such as “connected” or “linked” are not limited to physical or mechanical connections, but can include electrical connections, whether direct or indirect. Terms such as “upper,” “lower,” “left,” and “right” are used only to indicate relative positional relationships, which may change accordingly when the absolute position of the described objects changes.

[0076] Figure 1 This is a schematic diagram of a detection chip provided in an embodiment of this disclosure. Figure 2 This is a top view schematic diagram of the detection chip provided in an embodiment of this disclosure. Figure 3 This is a top view of the reaction area in an embodiment of this disclosure. Figure 4 for Figure 1 A schematic diagram of a cross-section along line A-A'. Figure 5 for Figure 1 A schematic diagram of a cross-section along the B-B' direction. (Example) Figures 1 to 5 As shown, the detection chip is divided into at least one functional region 1, which includes a reaction region 101 and a non-reaction region 102 surrounding the reaction region 101. The detection chip includes a first substrate 9 and a second substrate 10 disposed opposite to each other. A plurality of reaction grooves 4 are arranged in an array along a first direction X and a second direction Y on the side of the first substrate 9 facing the second substrate 10 and located within the reaction region 101. A first flow groove 5 is disposed between two adjacent reaction grooves 4 in the first direction X and extends along the first direction X. A second flow groove 6 is disposed between two adjacent first flow grooves 5 in the second direction Y and extends along the second direction Y. The first direction X and the second direction Y intersect.

[0077] During sample introduction, the sample solution is first injected into the reaction tank 4 through the first flow channel 5, ensuring that the sample solution fully enters each reaction tank 4 (i.e., the aqueous phase is introduced). Then, the liquid-sealing oil phase (i.e., the oil phase is introduced) is injected into the second flow channel 6. The liquid-sealing oil phase can divide the sample solution in the first flow channel 5 into two parts, thereby achieving oil phase isolation between each reaction tank 4. The liquid-sealing oil phase can be mineral oil, liquid paraffin, isopropyl palmitate followed by butyl lauryl ester, perfluoroalkane oil, etc.

[0078] It should be noted that, in the embodiments of this disclosure, the shape of the orthographic projection of the reaction groove 4 onto the second substrate 10 can be circular, square, or other regular or irregular shapes, and this disclosure does not limit this. In some embodiments, the aperture of the micro-reaction groove 4 includes 40um to 60um, for example 50um.

[0079] The detection chip provided in this embodiment has a simple structure and is easy to prepare. At the same time, the sample introduction process of the detection chip is stable and has high sample introduction efficiency, which can make the sample solution fully enter each reaction cell 4, thereby effectively improving the uniformity of the sample solution in each reaction cell 4.

[0080] In some embodiments, the first direction X is perpendicular to the second direction Y; during the injection of the liquid seal oil phase, the oil phase flow direction is perpendicular to the extension direction of the first flow channel 5, so that the oil phase has a better cutting effect on the sample solution in the first flow channel 5.

[0081] In some embodiments, a first inlet 6a and a first outlet 6b are provided on the second substrate 10 within the non-reaction region 102, penetrating the second substrate 10. A plurality of reaction tanks 4 and a plurality of first flow tanks 5 are alternately arranged in the first direction X, constituting a first flow channel 2. The two ends of the first flow channel 2 are respectively connected to the first inlet 6a and the first outlet 6b. During sample injection, a sample solution can be injected into the first flow channel 2 through the first inlet 6a. To ensure the best possible injection effect, a negative pressure can be applied to the first outlet 6b (e.g., vacuuming the first outlet 6b) while injecting the sample solution into the first inlet 6a. The injection of the sample solution is stopped when all reaction tanks 4 are filled with sample solution. Generally, the first flow channel 2 is also filled with sample solution at this time.

[0082] In some embodiments, the first inlet 6a and the first outlet 6b are located on opposite sides of the reaction region 101 in the first direction X. This design allows the sample solution entering the first flow channel 2 to preferentially move along the first direction X, ensuring that the sample solution is rapidly injected into the reaction tank 4 located within the first flow channel 2. Furthermore, the line connecting the center of the first inlet 6a and the center of the first outlet 6b extends along the first direction X and passes through the center of the reaction region 101. This arrangement facilitates the uniform input of the sample solution into each of the first flow channels 2 and the uniform discharge of gas within the first flow channels 2, ensuring that the sample solution is fully injected into each of the reaction tanks 4.

[0083] In some embodiments, a first liquid inlet connection groove 8a and a first liquid outlet connection groove 8b corresponding to the first flow channel 2 are further provided on the side of the first substrate 9 facing the second substrate 10; one end of the first liquid inlet connection groove 8a is connected to one end of the corresponding first flow channel 2, and the other end of the first liquid inlet connection groove 8a extends to the non-reaction area 102 and is connected to the first liquid inlet 6a; one end of the first liquid outlet connection groove 8b is connected to the other end of the corresponding first flow channel 2, and the other end of the first liquid inlet connection groove 8a extends to the non-reaction area 102 and is connected to the first liquid outlet 6b.

[0084] In some embodiments, a second liquid inlet 7a and a second liquid outlet 7b are formed on the second substrate 10 and within the non-reactive region 102, penetrating the second substrate 10; a plurality of second flow channels 6 arranged in the second direction Y constitute a second flow channel 3, with both ends of the second flow channel 3 connected to the second liquid inlet 7a and the second liquid outlet 7b, respectively. After the injection of the sample solution is completed, the injected sample solution can be input into the second flow channel 3 through the second liquid inlet 7a; to ensure the injection effect of the sample oil phase as much as possible, a negative pressure can be applied to the second liquid outlet 7b (e.g., vacuum treatment of the second liquid outlet 7b) while injecting the oil phase into the second liquid inlet 7a; when no bubbles are discharged from the second liquid outlet 7b, the oil phase injection is stopped.

[0085] In some embodiments, the second inlet 7a and the second outlet 7b are located on opposite sides of the reaction zone 101 in the second direction Y. This design allows the oil phase entering the second flow channel 3 to have a faster flow velocity in the second direction Y, thereby improving the cutting effect on the sample solution in the first flow tank 5. Furthermore, the line connecting the center of the second inlet 7a and the center of the second outlet 7b extends along the second direction Y and passes through the center of the reaction zone 101. This arrangement facilitates the uniform input of the oil phase into each of the second flow channels 3 and the uniform discharge of gas within each of the second flow channels 3, ensuring that the oil phase is fully injected into each of the second connecting tanks 6.

[0086] In some embodiments, a second liquid inlet connection groove 9a and a second liquid outlet connection groove 9b corresponding to the second flow channel 3 are further provided on the side of the first substrate 9 facing the second substrate 10; one end of the second liquid inlet connection groove 9a is connected to one end of the corresponding second flow channel 3, and the other end of the second liquid inlet connection groove 9a extends to the non-reaction area 102 and is connected to the second liquid inlet 7a; one end of the second liquid outlet connection groove 9b is connected to the other end of the corresponding second flow channel 3, and the other end of the second liquid inlet connection groove 9a extends to the non-reaction area 102 and is connected to the second liquid outlet 7b.

[0087] In this embodiment of the present disclosure, a first flow channel 2 for transporting sample solution and a second flow channel 3 for transporting oil phase are respectively provided. During the sample injection process, it is only necessary to first inject the sample solution into the first flow channel 2 and then inject the oil phase into the first flow channel 2. The whole sample injection process is relatively simple and convenient to operate.

[0088] In some embodiments, the width of the first flow channel 5 is greater than the width of the second flow channel 6. That is, the width of the first flow channel 5 is relatively wide, while the width of the second flow channel 6 is relatively narrow. The wider first flow channel 5 can effectively increase the speed at which the sample solution is injected into the reaction vessel 4, which is beneficial for reducing the injection time. The injection of the oil phase is for cutting the sample solution in the first flow channel 5, and the narrower first flow channel 5 can achieve a faster flow rate of the oil phase, resulting in a better cutting effect on the sample solution in the first flow channel 5. In some embodiments, the width of the first flow channel 5 ranges from 20 μm to 30 μm; the width of the second flow channel 6 ranges from 10 μm to 20 μm.

[0089] In some embodiments, the depth of the reaction tank 4 is greater than or equal to the depth of the second flow channel 6. In embodiments of this disclosure, the depth of the reaction tank 4 may be greater than the depth of the second flow channel 6, in which case the reaction tank 4 can accommodate a larger volume of sample solution.

[0090] In some embodiments, the first flow channel 5 includes a first portion 501, a second portion 502, and a third portion 503 connected sequentially along a first direction X. The first portion 501 and the third portion 503 are respectively connected to two adjacent reaction channels, and the second portion 502 is connected to the second flow channel 6. The depths of the first portion 501 and the third portion 503 are both greater than or equal to the depth of the second flow channel 6; the depth of the second portion 502 is equal to the depth of the second flow channel 6. Setting the depth of the second portion 502 connected to the second flow channel 6 in the first flow channel 5 to be equal to the depth of the second flow channel 6 ensures that the depths are consistent throughout the second flow channel 3, which facilitates rapid flow of the oil phase within the second flow channel 3 and improves the cutting effect of the oil phase on the sample solution. Simultaneously, the depths of the first portion 501 and the third portion 503 connected to the reaction channel 4 in the first flow channel 5 can be greater than the depth of the second flow channel 6, allowing more sample solution to exist around the reaction channel 4, which is beneficial for injecting the sample solution into the reaction channel 4.

[0091] See Figure 4 and Figure 5 As shown, in some embodiments, the first substrate 9 includes a substrate 11 and a hole defining layer 12 located on the side of the substrate 11 facing the second substrate 10. The substrate 11 may be a glass substrate; the hole defining layer 12 may form a series of hole structures, which can be used to form a reaction tank 4, a first flow channel 5, a second flow channel 6, a first liquid inlet connection channel 8a, a first liquid outlet connection channel 8b, a second liquid inlet connection channel 8a, and a second liquid outlet connection channel 8b.

[0092] It should be noted that, in the embodiments of this disclosure, the various hole structures on the hole limiting layer 12 can be selectively configured as through-hole structures that penetrate the hole limiting layer 12 or blind hole structures that do not penetrate the hole limiting layer 12 (blind hole structures can be regarded as grooves formed on the hole limiting layer 12).

[0093] A first hole structure 15 is provided on the hole limiting layer 12 in the area where the first flow groove 5 is to be formed, and the first flow groove 5 includes the first hole structure 15; a second hole structure 16 is provided on the hole limiting layer 12 in the area where the second flow groove 6 is to be formed, and the second flow groove 6 includes the second hole structure 16; a third hole structure 14 is provided on the hole limiting layer 12 in the area where the reaction groove 4 is to be formed, and the reaction groove 4 includes the third hole structure 14.

[0094] In some embodiments, when the first flow channel 5 includes only the first hole structure 15, the first hole structure 15 can be a through hole structure or a blind hole structure; when the second flow channel 6 includes only the second hole structure 16, the second hole structure 15 can be a through hole structure or a blind hole structure; when the reaction channel 4 includes only the third hole structure 14, the third hole structure 14 can be a through hole structure or a blind hole structure. In some embodiments, the first hole structure 15, the second hole structure 16, and the third hole structure 14 have the same depth.

[0095] In some embodiments, when a first liquid inlet connection groove 8a and a first liquid outlet connection groove 8b are provided on the side of the first substrate 9 facing the second substrate 10, a fourth hole structure 17 is provided on the hole limiting layer 12 in the area where the first liquid inlet connection groove 8a is to be formed, and a fifth hole structure 18 is provided on the hole limiting layer 12 in the area where the first liquid outlet connection groove 8b is to be formed. The first liquid inlet connection groove 8a includes the fourth hole structure 17, and the first liquid outlet connection groove 8b includes the fifth hole structure 18.

[0096] In some embodiments, when a second liquid inlet connection groove 9a and a second liquid outlet connection groove 9b are provided on the side of the first substrate 9 facing the second substrate 10, a sixth hole structure 19 is provided on the hole limiting layer 12 in the area where the second liquid inlet connection groove 9a is to be formed, and a seventh hole structure 20 is provided on the hole limiting layer 12 in the area where the second liquid outlet connection groove 9b is to be formed. The second liquid inlet connection groove 9a includes the sixth hole structure 19, and the second liquid outlet connection groove 9b includes the seventh hole structure 20.

[0097] Figure 6 for Figure 1 Another cross-sectional diagram along the A-A' direction. Figure 7 for Figure 1 Another cross-sectional diagram along the B-B' direction. (See diagram below.) Figure 6 and Figure 7As shown, in some embodiments, a heating electrode 23 is provided between the substrate 11 and the hole defining layer 12, and the heating electrode 23 is configured to heat the area where the reaction tank 4 is located.

[0098] During PCR, the double-stranded structure of a DNA fragment denatures at high temperatures to form a single-stranded structure. At low temperatures, primers bind to the single strands according to the complementary base pairing principle. Base binding and extension occur at the optimal temperature for DNA polymerase. This process is known as the denaturation-annealing-extension temperature cycle. Through multiple temperature cycles of denaturation-annealing-extension, DNA fragments can be replicated in large quantities. To achieve this temperature cycle, a series of external devices are typically required to heat and cool the detection chip, resulting in bulky equipment, complex operation, and high cost. Furthermore, the overall temperature of the detection chip changes during heating and cooling, causing temperature variations in other structures and components besides the microcavity containing the DNA fragment, increasing the risk of damage to components such as circuitry. Typical dPCR products often use droplet preparation systems, making the detection chips expensive and complex to manufacture.

[0099] To overcome the above-mentioned technical problems, the present invention provides a heating electrode 23 in the first substrate 9 to effectively control the temperature of the micro-reaction chamber. This effectively controls the temperature of the reaction tank 4 of the detection chip, achieves temperature cycling without the need for droplet driving operation, and eliminates the need for external heating equipment. It features high integration, simple operation, low production cost, and effective sample introduction.

[0100] The heating electrode 23 can receive electrical signals, thereby generating heat when current flows through it. This heat is conducted to at least a portion of the microreactor cavity to regulate its temperature. The heating electrode can be made of a conductive material with high resistivity, allowing it to generate more heat with a smaller electrical signal, thus improving energy conversion efficiency. In some embodiments, the heating electrode 23 can be made of a transparent conductive material, such as indium tin oxide (ITO) or tin oxide, or other suitable materials, such as metals. The embodiments of this disclosure are not limited in this regard.

[0101] In this embodiment, the heating electrode 23 can be a planar electrode, for example, uniformly formed on the substrate 11 using a conductive material, so that the multiple micro-reaction chambers are heated uniformly. Of course, the embodiments of this disclosure are not limited to this, and the heating electrode 23 can also have a specific shape or pattern, such as a broken line shape, an arc shape, etc., which can be determined according to the distribution of the multiple reaction tanks 4.

[0102] In some embodiments, a control electrode 21 is disposed between the heating electrode 23 and the substrate 11, and a first insulating layer 22 is disposed between the control electrode 21 and the heating electrode 23. The control electrode 21 is connected to the heating electrode 23 through a via on the first insulating layer 22, and the control electrode 21 is configured to transmit an external electrical signal to the heating electrode.

[0103] The number of control electrodes 21 can be one or more, and the embodiments of this disclosure are not limited in this respect. When multiple control electrodes 21 are used to apply electrical signals to the heating electrode 23, different parts of the heating electrode 23 can receive the electrical signals simultaneously, thereby making the heating of the heating electrode 23 more uniform. For example, when there are multiple control electrodes 21, the first insulating layer 22 may include multiple vias, each of which exposes a portion of the control electrode 21, so that the heating electrode 23 is electrically connected to the multiple control electrodes 21 through the multiple vias respectively. For example, the multiple control electrodes 21 and the multiple vias correspond one-to-one. As another example, the number of multiple vias may also be greater than the number of multiple control electrodes 21, with each control electrode being electrically connected to the heating electrode 23 through one or more vias.

[0104] The control electrode 21 can be made of a material with low resistivity, thereby reducing energy loss on the control electrode 21. The control electrode 21 can be made of a metallic material, such as copper or copper alloy, aluminum or aluminum alloy, etc., and can be a single metal layer or a composite metal layer. The embodiments disclosed herein are not limited in this regard.

[0105] In some embodiments of this disclosure, the heating electrode 23 is made of indium tin oxide (ITO) or tin oxide, and the control electrode 21 is made of a metallic material. Since ITO is not easily oxidized, partial oxidation of the heating electrode exposed to air can be prevented, thereby avoiding problems such as uneven heating or increased power consumption caused by oxidation of the heating electrode 23. The control electrode is covered by an insulating layer, so even if it is made of a metallic material, oxidation is not easily encountered.

[0106] To facilitate electrical connection between the control electrode 21 and an external electrical signal supply device for receiving electrical signals, the control electrode 21 may further include a contact portion 21a, which extends to the edge of the substrate 11 and is not covered by the first insulating layer 22. For example, the contact portion 21a is a relatively large square shape. Figure 1 and Figure 2The example diagram shows four contact portions, which facilitates easy connection with probes or electrodes in an electrical signal supply device. The large contact area ensures stable signal reception. This allows the detection chip to be plug-and-play, simple to operate, and convenient to use. For example, when the control electrode is made of a metallic material, the contact portions can be treated with electroplating, thermal spraying, or vacuum plating to form a protective layer on the surface of the contact portions 21a, preventing oxidation without affecting their conductivity.

[0107] In some embodiments, a second insulating layer 24 is disposed between the heating electrode 23 and the hole defining layer 12, and a light-shielding layer 25 is disposed between the second insulating layer 24 and the hole defining layer 12. A perforated structure 30 is provided on the light-shielding layer 25 in the area where the reaction tank 4 is to be formed; the reaction tank 4 also includes the perforated structure 30. Generally, after the PRC reaction is completed in the reaction tank 4, optical inspection of the reaction tank 4 is required to obtain a fluorescence image. By setting a light-shielding layer to block areas other than the area where the reaction tank 4 is located, interference from external light within the reaction tank 4 can be avoided, which helps improve the accuracy of optical inspection.

[0108] It should be noted that when the reaction tank 4 includes a hollow structure, the third hole structure 14 is a through hole structure to ensure communication with the hollow structure 30.

[0109] Figure 8 for Figure 1 Another cross-sectional diagram along the A-A' direction. Figure 9 for Figure 1 Another cross-sectional diagram along the B-B' direction. Figure 10 This is a schematic diagram of a structure of a portion of the detection chip in an embodiment of this disclosure. Figure 11 for Figure 10 A top view of a partial area of ​​the intermediate substrate 11. (See diagram below.) Figures 8 to 11 As shown, in some embodiments, a first receiving groove 27 is provided on the side of the substrate 11 facing the hole defining layer 12 and in the area where the reaction tank 4 is to be formed, and the reaction tank 4 further includes the first receiving groove 27. In the embodiments of this disclosure, by providing the first receiving groove 27 on the substrate 11, the first receiving groove 27, as part of the reaction tank 4, can effectively increase the depth of the reaction tank 4, so that more sample solution can be injected into the reaction tank 4, making it easier to perform detection. It should be noted that when the reaction tank 4 includes the first receiving groove 27, the third hole structure 14 is a through hole structure to ensure communication with the first receiving groove 27.

[0110] In some embodiments, the first flow channel 5 includes: a first portion 501, a second portion 502, and a third portion 503 connected sequentially along a first direction X. The first portion 501 and the third portion 503 are respectively connected to two adjacent reaction channels 4. The second portion 502 is connected to the second flow channel 6 (the second portion 502 is located on the flow path of the second flow channel 3). A second receiving channel 28 is provided on the substrate 11 in the area where the first portion 501 is to be formed. A third receiving channel 29 is provided on the substrate 11 in the area where the third portion 503 is to be formed. Both the second receiving channel 28 and the third receiving channel 29 are connected to the corresponding first receiving channel 27. The first flow channel 5 also includes the second receiving channel 28 and the third receiving channel 29.

[0111] In some embodiments, the depths of the first portion 501 and the third portion 503 are the same as the depth of the reaction tank 4 and greater than the depth of the second connecting tank 6, and the depth of the second portion is the same as the depth of the second connecting tank 6.

[0112] In some embodiments, a light-shielding layer 25 is disposed between the substrate 11 and the hole defining layer 12, and a perforation structure 30 is disposed on the light-shielding layer 25 in the area where the reaction tank 4 is to be formed; the reaction tank 4 also includes the perforation structure 30. When the material of the light-shielding layer 25 is a black resin material and the substrate 11 is a glass substrate, since the black resin material is easy to peel off from the surface of the glass substrate, in order to increase the bonding strength between the two, an auxiliary layer can be disposed between the substrate 11 and the light-shielding layer. The material of the auxiliary layer includes inorganic insulating materials, such as silicon oxide, silicon nitride, or a laminated structure composed of both.

[0113] In some embodiments, the second substrate 10 includes a cover plate 13 and a heating electrode 23 located on the side of the cover plate 13 facing the first substrate 9, the heating electrode 23 being configured to heat the region where the reaction tank 4 is located. The cover plate 13 may be a glass cover plate or a rigid plastic cover plate.

[0114] In some embodiments, a first protective layer 26 is provided on the side of the heating electrode 23 facing away from the cover plate 13 to prevent the heating electrode from directly contacting the sample solution or oil phase.

[0115] In some embodiments, a control electrode 21 is provided between the heating electrode 23 and the cover plate 13, and a first insulating layer 22 is provided between the control electrode 21 and the heating electrode 23. The control electrode 21 is connected to the heating electrode 23 through a through hole on the first insulating layer 22, and the control electrode 21 is configured to apply an electrical signal to the heating electrode 23.

[0116] See Figures 4 to 9 As shown, in some embodiments, the material of the hole defining layer 12 includes photoresist; in this case, the corresponding hole structures can be formed by exposing and developing the photoresist.

[0117] In some embodiments, a hydrophilic layer (not shown) is provided at the bottom of the reaction tank 4, the sidewall of the reaction tank 4, the bottom of the first flow channel 5, and / or the sidewall of the first flow channel 5. Providing a hydrophilic layer at at least one of the following locations—the bottom of the reaction tank 4, the sidewall of the reaction tank 4, the bottom of the first flow channel 5, and the sidewall of the first flow channel 5—facilitates confining the sample solution within the first flow channel 2. Specifically, providing a hydrophilic layer at the bottom of the reaction tank 4 and / or the sidewall of the reaction tank 4 facilitates the entry of the sample solution into the reaction tank 4.

[0118] In some embodiments, a hydrophobic layer (not shown) is provided at the bottom of the second flow channel 6 and / or on the sidewall of the second flow channel 6, which facilitates better adsorption of the oil phase for liquid sealing within the second flow channel 3.

[0119] In some embodiments, a second protective layer (not shown) is further provided on the side of the hole defining layer 12 facing away from the substrate 11 to prevent the hole defining layer 12 from directly contacting the sample solution or oil phase. It should be noted that when the above-mentioned hydrophilic / hydrophobic layer and the second protective layer are provided in the detection chip, the hydrophilic / hydrophobic layer is located at a corresponding position on the side of the second protective layer facing away from the substrate 11.

[0120] Of course, the second protective layer can also be reused as a hydrophilic layer and a hydrophobic layer. In this case, there is no need to set an additional hydrophilic or hydrophobic layer when a second protective layer is provided. For example, the material of the second protective layer is silicon oxide. The untreated silicon oxide film itself is hydrophilic. Then, the surface of the silicon oxide film in the area where a hydrophobic layer needs to be set is treated (e.g., plasma treatment) to reduce the surface energy of the corresponding area and make it hydrophobic.

[0121] Figure 12 This is another schematic diagram of the detection chip provided in an embodiment of this disclosure. For example... Figure 12 As shown, unlike the previous embodiments, the number of functional areas 1 in this embodiment is multiple. Figure 12 The example shows four functional areas, meaning that the detection chip has multiple independent reaction areas 101 to meet the detection needs of different application scenarios.

[0122] Based on the same inventive concept, this disclosure also provides a method for preparing a detection chip, which can be used to prepare the detection chip provided in the above embodiments.

[0123] Figure 13 A flowchart illustrating a method for fabricating a detection chip according to an embodiment of this disclosure is shown below. Figure 13 As shown, the preparation method includes:

[0124] Step S101: Prepare the first substrate and the second substrate respectively.

[0125] The first substrate has a plurality of reaction cells arranged in an array along a first direction and a second direction on one side. A first flow channel connected to the two reaction cells is provided between two adjacent reaction cells in the first direction. The first flow channel extends along the first direction. A second flow channel connected to the two first flow channels is provided between two adjacent first flow channels in the second direction. The second flow channel extends along the second direction. The first direction and the second direction intersect.

[0126] To prepare Figure 4 and Figure 5 The first and second substrates shown are examples. The process of preparing the first substrate is as follows: First, a substrate is provided; then, a hole-defining layer is prepared on the substrate. The substrate can be a glass substrate. The process of preparing the hole-defining layer is as follows: First, photoresist is spin-coated at 300 rpm for 10 seconds and baked at 90°C for 2 minutes; then, the photoresist is spin-coated once and the above process is repeated to obtain a photoresist layer; then, the photoresist layer is exposed through a mask; then, the exposed photoresist layer is developed with a developer for 45 seconds, and then cured at 230°C for 30 minutes to obtain the hole-defining layer. The process of preparing the second substrate is as follows: First, a cover plate is provided; then, a first liquid inlet / outlet and a second liquid inlet / outlet are formed on the cover plate. The cover plate can be a glass cover plate or a rigid plastic cover plate; the first liquid inlet / outlet and the second liquid inlet / outlet can be formed on the cover plate by laser drilling or etching.

[0127] It should be noted that if the first substrate and the second substrate are adopted Figure 6 and Figure 7 As shown, in the process of fabricating the first substrate, before the step of fabricating the hole defining layer, the steps of fabricating a control electrode, fabricating a first insulating layer, fabricating a heating electrode, fabricating a second insulating layer, and fabricating a light-shielding layer are included. Optionally, after fabricating the hole defining layer, the steps of fabricating a second protective layer and fabricating a hydrophilic / hydrophobic layer may also be included.

[0128] The control electrode can be made of a metallic material, such as a molybdenum-aluminum-neodymium-molybdenum (Mo-AlNd-Mo) laminate; wherein the thickness of the lower molybdenum layer can be [missing information]. The thickness of aluminum neodymium can be The thickness of the upper molybdenum layer can be The material of the first insulating layer can be silicon oxide (SiO2), and its thickness can be [missing information]. The heating electrode can be made of indium tin oxide (ITO) and its thickness can be [missing information]. The second insulating layer can be a laminated structure of silicon oxide and silicon nitride, wherein the thickness of the silicon oxide can be [missing information]. The thickness of silicon nitride (SiNx) can be... The light-shielding layer can be made of black resin. The second protective layer can be made of silicon dioxide, and its thickness can be [missing information]. At this point, the second protective layer serves as both a hydrophilic and a hydrophobic layer. Specifically, the portion of the second protective layer (silicon oxide) covering the bottom of the reaction tank, the sidewalls of the reaction tank, the bottom of the first flow tank, and the sidewalls of the first flow tank is hydrophilic to serve as a hydrophilic layer. The portion of the second protective layer (silicon oxide) covering the bottom of the second flow tank and the sidewalls of the second flow tank undergoes surface treatment (e.g., plasma treatment) to reduce the surface energy of the second protective layer at the corresponding locations, making it hydrophobic to serve as a hydrophobic layer. Of course, the hydrophilic / hydrophobic layer can also have a different structure from the second protective layer.

[0129] It should be noted that if the first substrate and the second substrate are adopted Figure 8 and Figure 9 As shown, in the process of fabricating the first substrate, before the step of fabricating the hole defining layer, there are steps of fabricating an auxiliary layer and a light-shielding layer; in the process of fabricating the second substrate, after the steps of forming the first liquid inlet / outlet and the second liquid inlet / outlet on the cover plate, there are steps of fabricating a control electrode, fabricating a first insulating layer, fabricating a heating electrode, and fabricating a first protective layer. For the steps of fabricating the light-shielding layer, fabricating the control electrode, fabricating the first insulating layer, and fabricating the heating electrode, please refer to the preceding content, and they will not be repeated here.

[0130] The auxiliary layer can be a laminated structure of silicon oxide and silicon nitride, wherein the thickness of the silicon oxide can be [missing information]. The thickness of silicon nitride can be The first protective layer can be made of silicon oxide, and its thickness can be [missing information].

[0131] It should be noted that when the cover plate is provided with structures such as control electrodes, first insulating layer, first insulating layer, and first protective layer, these structures on the cover plate will not cover the first inlet / outlet and the second inlet / outlet, so as to ensure that the first inlet / outlet and the second inlet / outlet can communicate with the corresponding connecting grooves on the first substrate.

[0132] Step S102: The side of the first substrate with the reaction tank, the first flow tank and the second flow tank is positioned opposite to the second substrate, and the first substrate and the second substrate are packaged.

[0133] In step S102, a pressure-sensitive adhesive film can be attached to the side of the second substrate opposite to the first substrate. Then, the first and second substrates are joined together and rolled under pressure to complete chip packaging.

[0134] Based on the same inventive concept, this disclosure also provides a sampling method for a detection chip, which is based on the detection chip provided in the above embodiments.

[0135] Figure 14 This is a flowchart illustrating a sample introduction method for a detection chip provided in an embodiment of this disclosure. Figure 14 As shown, the preparation method includes:

[0136] Step S201: Inject the sample solution into the reaction tank through the first flow channel.

[0137] Step S202: Inject liquid seal oil phase into the second flow channel to isolate the oil phase in each reaction tank.

[0138] In some embodiments, before sample injection begins, rubber caps can be used to secure the first inlet, first outlet, second inlet, and second outlet to ensure that all three outlets are sealed. During sample injection, two metal needles are inserted into the rubber caps at the first inlet and first outlet respectively to release the seals. Then, the pre-mixed sample solution is forced into the first inlet through the metal needle at the first inlet (simultaneously, a certain negative pressure can be applied at the first outlet). The sample solution flows within the first flow channel, filling each reaction cell sequentially. After all reaction cells are filled, the metal needles at the first inlet and first outlet are removed to reseal the first inlet and first outlet. Next, two metal needles are inserted into the rubber caps at the second inlet and the second outlet to release the seals. Then, the liquid seal oil phase is injected into the second inlet through the metal needle (at the same time, a certain negative pressure can be applied at the second outlet). The oil phase flows in the second flow channel and cuts the sample solution in the first flow cell into two parts (the second part in the first flow cell contains the oil phase, and the first and third parts in the first flow cell contain the sample solution), thereby isolating each reaction cell with the oil phase. The injection of the oil phase is stopped when no more air bubbles are expelled from the second outlet, and the sample injection is completed.

[0139] In some embodiments, when the first substrate includes a heating electrode, an electrical signal can be provided to the heating electrode as needed during the sample injection process and during the PCR reaction after the sample injection is completed, so as to adjust the temperature of the reaction tank.

[0140] It is understood that the above embodiments are merely exemplary embodiments used to illustrate the principles of this disclosure, and this disclosure is not limited thereto. For those skilled in the art, various modifications and improvements can be made without departing from the spirit and substance of this disclosure, and these modifications and improvements are also considered to be within the scope of protection of this disclosure.

Claims

1. A detection chip, wherein, The chip is divided into at least one functional area, the functional area including: a reaction area and a non-reaction area surrounding the reaction area, the detection chip including: a first substrate and a second substrate disposed opposite to each other, the first substrate having a plurality of reaction slots arranged in an array along a first direction and a second direction on the side facing the second substrate and located in the reaction area. A first flow channel is provided between two adjacent reaction channels in the first direction, and the first flow channel extends along the first direction. A second flow groove connected to the two first flow grooves is provided between two adjacent first flow grooves in the second direction, and the second flow groove extends along the second direction. The first direction intersects with the second direction; The first substrate includes: a substrate substrate and a hole defining layer located on the side of the substrate substrate facing the second substrate; the side of the substrate substrate facing the hole defining layer and having a first receiving groove in the region where the reaction groove is to be formed; the reaction groove includes the first receiving groove; The first flow channel includes: a first part, a second part, and a third part connected sequentially along a first direction, wherein the first part and the third part are respectively connected to two adjacent reaction channels, and the second part is connected to the second flow channel; A second receiving groove is provided on the substrate in the region where the first part is to be formed, and a third receiving groove is provided on the substrate in the region where the third part is to be formed. Both the second receiving groove and the third receiving groove are connected to the corresponding first receiving groove. The first flow channel further includes the second receiving channel and the third receiving channel; A light-shielding layer is provided between the substrate and the hole-defining layer, and a hollow structure is provided on the light-shielding layer in the area where the reaction tank is to be formed. The reaction tank also includes the hollow structure.

2. The detection chip according to claim 1, wherein, The width of the first flow channel is greater than the width of the second flow channel.

3. The detection chip according to claim 1 or 2, wherein, The depth of the reaction tank is greater than or equal to the depth of the second flow tank.

4. The detection chip according to any one of claims 1 to 3, wherein, The first flow channel includes: a first part, a second part, and a third part connected sequentially along a first direction, wherein the first part and the third part are respectively connected to two adjacent reaction channels, and the second part is connected to the second flow channel; The depth of the first part and the depth of the third part are both greater than or equal to the depth of the second flow channel; The depth of the second part is equal to the depth of the second flow channel.

5. The detection chip according to any one of claims 1 to 4, wherein, A first liquid inlet and a first liquid outlet penetrating the second substrate are provided on the second substrate and located in the non-reactive region. A plurality of reaction tanks and a plurality of first flow tanks arranged alternately in a first direction constitute a first flow channel, and the two ends of the first flow channel are respectively connected to the first liquid inlet and the first liquid outlet.

6. The detection chip according to claim 5, wherein, The first inlet and the first outlet are located on opposite sides of the reaction zone in the first direction.

7. The detection chip according to claim 6, wherein, The line connecting the center of the first inlet and the center of the first outlet extends along the first direction and passes through the center of the reaction region.

8. The detection chip according to any one of claims 5 to 7, wherein, A first liquid inlet connection groove and a first liquid outlet connection groove corresponding to the first flow channel are also provided on the side of the first substrate facing the second substrate. One end of the first liquid inlet connection groove is connected to one end of the corresponding first flow channel, and the other end of the first liquid inlet connection groove extends to the non-reaction area and is connected to the first liquid inlet. One end of the first liquid outlet connection groove is connected to the other end of the corresponding first flow channel, and the other end of the first liquid inlet connection groove extends to the non-reaction area and is connected to the first liquid outlet.

9. The detection chip according to any one of claims 1 to 8, wherein, A second liquid inlet and a second liquid outlet are formed on the second substrate and located in the non-reactive region, penetrating the second substrate. The second flow channels arranged in the second direction constitute a second flow channel, and the two ends of the second flow channel are respectively connected to the second liquid inlet and the second liquid outlet.

10. The detection chip according to claim 9, wherein, The second inlet and the second outlet are located on opposite sides of the reaction zone in the second direction.

11. The detection chip according to claim 10, wherein, The line connecting the center of the second inlet and the center of the second outlet extends along the second direction and passes through the center of the reaction region.

12. The detection chip according to any one of claims 9 to 11, wherein, A second liquid inlet connection groove and a second liquid outlet connection groove corresponding to the second flow channel are also provided on the side of the first substrate facing the second substrate. One end of the second liquid inlet connection groove is connected to one end of the corresponding second flow channel, and the other end of the second liquid inlet connection groove extends to the non-reaction area and is connected to the second liquid inlet. One end of the second liquid outlet connection groove is connected to the other end of the corresponding second flow channel, and the other end of the second liquid inlet connection groove extends to the non-reaction area and is connected to the second liquid outlet.

13. The detection chip according to any one of claims 1 to 12, wherein, The width of the first flow channel ranges from 20µm to 30µm; The width of the second flow channel ranges from 10µm to 20µm.

14. The detection chip according to any one of claims 1 to 13, wherein, The hole limiting layer has a first hole structure in the area where the first flow groove is to be formed, and the first flow groove includes the first hole structure. The hole limiting layer has a second hole structure in the area where the second flow groove is to be formed, and the second flow groove includes the second hole structure. A third hole structure is provided on the hole-defining layer in the area where the reaction tank is to be formed, and the reaction tank includes the third hole structure.

15. The detection chip according to claim 14, wherein, When a first liquid inlet connection groove and a first liquid outlet connection groove are provided on the side of the first substrate facing the second substrate, a fourth hole structure is provided on the hole limiting layer in the area where the first liquid inlet connection groove is to be formed, and a fifth hole structure is provided on the hole limiting layer in the area where the first liquid outlet connection groove is to be formed. The first liquid inlet connection groove includes the fourth hole structure, and the first liquid outlet connection groove includes the fifth hole structure.

16. The detection chip according to claim 14, wherein, When a second liquid inlet connection groove and a second liquid outlet connection groove are provided on the side of the first substrate facing the second substrate, a sixth hole structure is provided on the hole limiting layer in the area where the second liquid inlet connection groove is to be formed, and a seventh hole structure is provided on the hole limiting layer in the area where the second liquid outlet connection groove is to be formed. The second liquid inlet connection groove includes the sixth hole structure, and the second liquid outlet connection groove includes the seventh hole structure.

17. The detection chip according to any one of claims 14 to 16, wherein, A heating electrode is disposed between the substrate and the hole defining layer, and the heating electrode is configured to heat the region where the reaction tank is located.

18. The detection chip according to claim 17, wherein, A control electrode is disposed between the heating electrode and the substrate, and a first insulating layer is disposed between the control electrode and the heating electrode. The control electrode is connected to the heating electrode through a via on the first insulating layer, and the control electrode is configured to apply an electrical signal to the heating electrode.

19. The detection chip according to claim 17 or 18, wherein, A second insulating layer is provided between the heating electrode and the hole limiting layer, and a light-shielding layer is provided between the second insulating layer and the hole limiting layer. A hollow structure is provided on the light-shielding layer in the area where the reaction tank is to be formed. The reaction tank also includes the hollow structure.

20. The detection chip according to claim 1, wherein, The second substrate includes a cover plate and a heating electrode located on the side of the cover plate facing the first substrate, the heating electrode being configured to heat the area where the reaction tank is located.

21. The detection chip according to claim 20, wherein, A first protective layer is provided on the side of the heating electrode facing away from the cover plate.

22. The detection chip according to claim 20 or 21, wherein, A control electrode is disposed between the heating electrode and the cover plate, and a first insulating layer is disposed between the control electrode and the heating electrode. The control electrode is connected to the heating electrode through a through hole in the first insulating layer, and the control electrode is configured to apply an electrical signal to the heating electrode.

23. The detection chip according to any one of claims 14 to 22, wherein, The material of the hole-defining layer includes photoresist.

24. The detection chip according to any one of claims 1 to 23, wherein, A hydrophilic layer is provided on the bottom of the reaction tank, the side wall of the reaction tank, the bottom of the first flow tank, and / or the side wall of the first flow tank.

25. The detection chip according to any one of claims 1 to 24, wherein, A hydrophobic layer is provided at the bottom of the second flow channel and / or on the sidewall of the second flow channel.

26. The detection chip according to any one of claims 1 to 25, wherein, There are multiple functional areas.

27. A method for preparing a detection chip as described in any one of claims 1 to 26, wherein, The detection chip is divided into at least one functional region, the functional region including: a reactive region and a non-reactive region surrounding the reactive region, the fabrication method including: A first substrate and a second substrate are prepared respectively. A plurality of reaction cells are arranged in an array along a first direction and a second direction on one side of the first substrate. A first flow channel connected to the two reaction cells is provided between two adjacent reaction cells in the first direction. The first flow channel extends along the first direction. A second flow channel connected to the two first flow channels is provided between two adjacent first flow channels in the second direction. The second flow channel extends along the second direction. The first direction and the second direction intersect. The first substrate, which has the reaction tank, the first flow tank and the second flow tank, is positioned opposite the second substrate, and the first substrate and the second substrate are then encapsulated.

28. A sample introduction method for a detection chip as described in any one of claims 1 to 26, wherein, include: The sample solution is injected into the reaction tank through the first flow channel; An oil phase is injected into the second flow channel to isolate the reaction tanks from the oil phase.

Images