A closed microfluidic chip for nucleic acid amplification detection

Abstract

The present invention provides a closed microfluidic chip for nucleic acid amplification detection, comprising a gland and at least one chip; the first surface of the gland is provided with at least one mounting groove, and the second surface is provided with at least one gland inlet and gland outlet corresponding to each mounting groove; at least one amplification unit is formed inside the chip, and the chip is installed in the mounting groove; an elastic tape is provided at the bottom of the mounting groove, and the elastic tape fixes the chip in the mounting groove; at least two microcavities are formed inside the elastic tape, and the at least two microcavities seal the connection between the gland inlet and the amplification unit, and the connection between the gland outlet and the amplification unit; the second surface of the gland is provided with at least one sealing mechanism, and the sealing mechanism is used to seal the gland inlet and the gland outlet. During the nucleic acid amplification detection process, leakage of liquid samples can be prevented, thereby achieving a fully enclosed PCR environment. The manufacturing process is simple, the cost is low, and it is easy to apply and promote on a large scale.

Description

Technical Field

[0001] The present invention relates to the field of in vitro diagnostic nucleic acid detection, and in particular to a closed microfluidic chip for nucleic acid amplification detection. Background Art

[0002] As one of the primary testing methods in the field of in vitro diagnostics, nucleic acid testing is the most direct, reliable, and sensitive method for early, rapid, and specific detection of pathogens. It can rapidly detect pathogen nucleic acids in test samples, providing a scientific basis for confirming infection. Nucleic acid amplification testing involves the amplification of nucleic acid sequences using enzymes. PCR (polymerase chain reaction) technology is the most widely used due to its high specificity and low cost. PCR consists of three basic reaction steps: denaturation, annealing, and extension. Reaction time has always been a limiting factor in the application of PCR technology. Instruments that improve PCR reaction time have emerged, such as patent document CN 111269825 A, which discloses a rapid nucleic acid instant detector with a rapid heating and cooling module. However, current PCR reaction vessels have low thermal conductivity. For PCR reactions that require cyclic heating and cooling, this low thermal conductivity results in slow heating and cooling speeds, resulting in a lengthy process.

[0003] Microfluidic chip technology integrates traditional biochemical analysis onto a chip measuring a few square centimeters or even smaller, performing detection and analysis within the chip's micro- and nanoscale channels and microchambers. However, because microfluidic chips require suitable thermally conductive materials for nucleic acid detection, manufacturing difficulty and cost are significantly increased, and achieving a fully enclosed PCR amplification environment is difficult, limiting their further clinical application. Therefore, there is an urgent need for a closed, highly thermally conductive microfluidic chip that can improve the chip's thermal conductivity, reduce detection time, achieve a fully enclosed PCR amplification environment, and reduce chip manufacturing costs. Summary of the Invention

[0004] In view of all or part of the above-mentioned deficiencies in the prior art, the present invention provides a closed microfluidic chip for nucleic acid amplification detection, which can prevent leakage of liquid samples during the nucleic acid amplification detection process, thereby realizing a fully closed PCR environment. Its manufacturing process is simple, the cost is low, and it is easy to apply and promote on a large scale.

[0005] In order to achieve the above-mentioned purpose of the invention, the present invention provides the following technical solutions: a closed microfluidic chip for nucleic acid amplification detection, comprising a pressure cover and at least one chip; the first surface of the pressure cover is provided with at least one mounting groove, and the second surface is provided with at least one pressure cover sample inlet and pressure cover air outlet corresponding to each mounting groove; at least one amplification part is formed inside the chip, and the chip is installed in the mounting groove; an elastic tape is provided at the bottom of the mounting groove, and the elastic tape fixes the chip in the mounting groove; at least two microcavities are formed inside the elastic tape, and at least two of the microcavities seal the connection between the pressure cover sample inlet and the amplification part, and the connection between the pressure cover air outlet and the amplification part; the second surface of the pressure cover is provided with at least one sealing mechanism, and the sealing mechanism is used to seal the pressure cover sample inlet and the pressure cover air outlet.

[0006] The elastic tape has an elastic buffering effect. On the one hand, the chip can be fixed in place by sticking the chip in the mounting groove with the sticky tape. On the other hand, when the microfluidic chip is used together with an external device (such as a PCR device), the elastic buffering effect of the elastic tape can be used to seal the connection between the gland inlet and the chip amplification part, and the connection between the gland outlet and the chip amplification part. The liquid sample enters the chip amplification part from the gland inlet, and the gas in the chip is discharged from the chip amplification part to the gland outlet. The sealing effect of the elastic tape can prevent the sample from leaking. The sealing of the gland inlet and the gland outlet by the sealing mechanism can achieve a fully enclosed PCR amplification environment. There can be one or more sealing mechanisms, and one sealing mechanism can be used to seal one or more gland inlets and gland outlets.

[0007] The first surface of the gland is provided with four mounting slots, and the second surface is provided with a gland sample inlet and a gland air outlet corresponding to each mounting slot. A chip is mounted in each mounting slot, and an amplification unit is formed within each chip. Each amplification unit can test one sample. When one amplification unit is present in a chip, one chip can test one sample. One chip can optionally be mounted in one mounting slot. When a gland has four mounting slots, four samples can be tested simultaneously.

[0008] The first surface of the gland is provided with at least one iron sheet, which interacts with a magnet on an external device to generate suction. The iron sheet can be formed on the gland by an embedding process during the gland's injection molding process. The provided iron sheet interacts with a magnet mounted on the external device to generate suction, causing the elastic tape within the mounting slot to be squeezed by a force (the elastic tape and chip must have a certain thickness to allow the elastic tape to be squeezed when the iron sheet on the gland generates suction with the magnet on the external device). The suction between the iron sheet and the magnet further enhances the sealing effect of the elastic tape, preventing the reaction liquid from leaking into the air during the temperature increase and decrease process, thereby preventing aerosol contamination.

[0009] Two iron sheets are provided on the first surface of the gland, symmetrically positioned on either side of the gland and around the periphery of the mounting slot. These two iron sheets ensure that the chip is uniformly affected in multiple directions by the attraction between the surrounding iron sheets and the magnets, thereby exerting multi-directional pressure on the elastic tape and further enhancing the sealing effect.

[0010] The sealing mechanism includes a fixed structure fixed on the gland, and a plurality of sealing plugs connected to the fixed structure, and the sealing plugs are used to seal the gland inlet and the gland outlet. The plurality of sealing plugs can be in a row and as a whole, and a row of sealing plugs can be connected to a fixed structure to seal multiple gland inlets and gland outlets at the same time. The sealing plug is connected to the gland through the fixed structure, which can facilitate the sealing of the gland inlet and the gland outlet. The material of the sealing plug is silicone or an elastic substrate coated with a rubber layer. The use of silicone sealing plugs can prevent the sealing material from reacting with the sample.

[0011] The elastic tape is a high-temperature-resistant foam tape, but is not limited to this material. The scope of protection of this patent includes polyacrylate, silicone, and other elastic and high-temperature-resistant materials as the primary chemical materials. High-temperature resistance refers to the ability to maintain a temperature of 100°C for more than 30 minutes. The nucleic acid amplification process involves multiple heating and cooling stages. The use of high-temperature-resistant foam tape prevents the reaction liquid from leaking into the air during the rapid heating and cooling process, causing aerosol contamination, thereby achieving a fully enclosed PCR environment.

[0012] The chip comprises a first material layer and a second material layer, with the first layer positioned between the elastic tape and the second layer. The first layer serves as the sample flow path, while the second layer serves as the amplification layer. The second layer is the primary component of the amplification unit, where the sample undergoes amplification reactions, while the first layer serves as a channel for sample inlet and outlet. The composite dual-layer chip structure simplifies chip processing.

[0013] The chip also includes a heat conductive sheet, which is located on the side surface of the second material layer facing away from the first material layer, and the heat conductive sheet is connected to an external heat source. The heat conductive sheet can be a non-metallic heat conductive sheet or a metal heat conductive sheet. The non-metallic heat conductive sheet includes a heat conductive silicone sheet, a heat conductive sheet made of a heat conductive phase change material, or a heat conductive graphite sheet, etc. The metal heat conductive sheet includes a metal heat conductive sheet such as copper or aluminum. An amplification part is formed inside the second material layer, and during the nucleic acid amplification process, there are multiple temperature rise and fall processes. The heat conductive sheet is directly contacted with the second material layer formed with the amplification part, and heat is transferred to the heat conductive sheet through an external heat source (for example, the heat conductive sheet is placed on the heat transfer seat of the PCR device and the heat transfer seat is connected to the external heat source), and the heat conductive sheet then transfers the heat to the second material layer of the chip.

[0014] At least one sample inlet and an air outlet are formed inside the first material layer, and at least one liquid inlet microchannel, an amplification chamber, and a liquid outlet microchannel are formed inside the second material layer; the pressure cap sample inlet, the sample inlet, the liquid inlet microchannel, and the amplification chamber are connected in sequence, and the pressure cap air outlet, the air outlet, the liquid outlet microchannel, and the amplification chamber are connected in sequence. The microcavities formed inside the elastic tape are respectively located between the pressure cap sample inlet and the sample inlet, and between the pressure cap air outlet and the air outlet, and seal the pressure cap sample inlet and the sample inlet, and between the pressure cap air outlet and the air outlet. The formation of the amplification chamber, the liquid inlet microchannel, and the liquid outlet microchannel can be completed by an etching process, forming a ditch-like structure in the second material layer, and using the first material layer to cooperate to form the amplification chamber, the liquid inlet microchannel, and the liquid outlet microchannel. The process is simple and the chip preparation cost is low.

[0015] The amplification chamber is a serpentine tube amplification chamber. The serpentine shape of the amplification chamber allows for a longer reaction chamber within a smaller space, extending the sample flow path and thereby improving the detection sensitivity and reliability of the microfluidic chip. Arranging the amplification chamber into multiple reciprocating, curved chamber structures, such as serpentines, can reduce the size of the chip.

[0016] The first material layer is a glass sheet, and the second material layer is a silicon wafer. Using a multi-material composite chip allows the properties of different materials to be utilized to meet the needs of the target chip, and a silicon-based chip can be obtained through a wafer etching process. The silicon-based chip is made of a material with high thermal conductivity. This high thermal conductivity material allows the temperature rise and fall of the amplification chamber within the chip to be nearly synchronized with the temperature rise and fall of the heat transfer base of the external device, resulting in faster heat conduction and rapid temperature rise and fall of the microfluidic chip.

[0017] The pressure cover is provided with at least one positioning hole, through which the pressure cover is positioned on the external device. The positioning hole allows the part of the chip that needs to be heated to be accurately positioned at the heat source of the external device, thereby improving the heating and cooling efficiency.

[0018] The two sides of the gland are symmetrically provided with hand-held structures, which facilitates the operator to use the microfluidic chip and is safer and more convenient during the use of nucleic acid amplification detection.

[0019] The beneficial effects of the technical solution of the present invention mainly include: the closed microfluidic chip for nucleic acid amplification detection provided by the present invention is fixed by the chip by arranging a layer of elastic tape between the mounting groove and the chip, and the elastic buffering effect of the elastic tape is used to seal the pressure cover sample inlet, the pressure cover air outlet and the chip. The pressure cover sample inlet and the pressure cover air outlet are sealed by a sealing mechanism, and the elastic tape is further squeezed by the suction effect of the iron sheet and the magnet, thereby improving the sealing effect of the elastic tape and realizing a fully enclosed PCR environment. The chip formed of a high thermal conductivity material is further heated by a heat conductive sheet, so that the amplification part can be quickly heated and cooled, greatly shortening the time consumption of nucleic acid detection. The manufacturing process of the present invention is simple, the cost is low, and it is convenient for large-scale application and promotion. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] In order to more clearly illustrate the technical solutions in the specific embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0021] Figure 1 This is a front perspective view of the microfluidic chip according to Example 1 of the present invention.

[0022] Figure 2 This is a schematic diagram of the three-dimensional structure of the gland according to Example 1 of the present invention.

[0023] Figure 3 This is a top view of the silicon-based chip of Example 1 of the present invention.

[0024] Figure 4 This is a main view of the silicon-based chip of Example 1 of the present invention.

[0025] Figure 5 This is a front perspective view of the silicon-based chip of Example 1 of the present invention.

[0026] Figure 6 This is a perspective structural diagram of the silicon-based chip and foam tape according to Example 1 of the present invention.

[0027] Figure 7 This is a side perspective view of the microfluidic chip according to Example 1 of the present invention.

[0028] Figure 8 This is a top perspective view of the microfluidic chip (including the heat transfer seat) according to Example 1 of the present invention.

[0029] Figure 9 Schematic diagram of the three-dimensional structure of the microfluidic chip according to Example 1 of the present invention.

[0030] Figure 10 This is a rear view (second side, back side) of the microfluidic chip according to Example 1 of the present invention.

[0031] Figure 11 This is a main view (first side, front) of the microfluidic chip (including the heat transfer seat) according to Example 1 of the present invention.

[0032] Figure markings: 1-pressure cover; 101-pressure cover sample inlet; 102-pressure cover air outlet; 103-pressure cover liquid inlet micropore; 104-pressure cover liquid outlet micropore; 2-sealing mechanism; 201-fixing structure; 202-sealing plug; 301, 302-iron sheets; 4-positioning hole; 5-chip; 501-amplification chamber; 502-liquid inlet microchannel; 503-liquid outlet microchannel; 504-sample injection hole; 505-air outlet; 506-first material layer; 507-second material layer; 508-heat conductive sheet; 6-elastic tape; 601-liquid inlet hole; 602-liquid outlet hole; 7-heat transfer seat; 8-mounting slot; 110-handheld structure. DETAILED DESCRIPTION

[0033] The following is a clear and complete description of the technical solutions in the specific embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0034] Example 1

[0035] This embodiment provides a closed microfluidic chip for nucleic acid amplification detection, such as Figure 1 and Figure 2 As shown, it includes a pressure cover 1, an elastic tape 6 and a chip 5. The pressure cover 1 is a plastic pressure cover, specifically a polycarbonate material (PC plastic) made by an injection molding process. PC plastic is a colorless and transparent amorphous thermoplastic material with excellent physical and mechanical properties such as heat resistance, good transparency and extremely high impact resistance, and has the characteristics of high light transmittance, refractive index and easy processing and molding. In other embodiments, the substrate of the pressure cover 1 can also be made of other polymer materials such as polystyrene (PS), polypropylene (PP), cycloolefin polymer (COP), polyvinyl chloride (PVC) and other heat-resistant materials. The pressure cover 1 includes a first side (front side, the front side of which serves as the main view of the microfluidic chip) and a second side (back side, the back side of which serves as the rear view of the microfluidic chip, and the back side can be facing up when in use). The substrate of the pressure cover 1 used in this embodiment is colorless and transparent. Figure 1Shown is a front perspective view of the microfluidic chip, which simultaneously shows partial structures of the first and second sides of the gland 1.

[0036] On the first side (front side) of the gland 1, four mounting grooves 8 are formed by an injection molding process for mounting the chip 5. The four mounting grooves 8 are distributed in the middle of the gland 1 and can be distributed in a straight line parallel manner or in a cross shape. In this embodiment, the four mounting grooves 8 are distributed in a cross shape. On the second side (back side) of the gland 1 corresponding to the mounting grooves 8, a gland inlet 101 and a gland outlet 102 are formed. In the mounting grooves 8, gland inlet microholes 103 communicating with the gland inlet 101 and gland outlet microholes 104 communicating with the gland outlet 102 are correspondingly formed.

[0037] At least one chip 5 can be installed in each mounting groove 8, and one chip 5 can be used to detect at least one sample. Therefore, by providing multiple mounting grooves 8, multiple samples can be detected simultaneously. The corresponding number of chips 5 is placed in the mounting grooves 8 as needed, and the chips 5 can be selectively assembled. In other embodiments, the gland 1 may also be provided with more or fewer than four mounting grooves 8, for example, only one or two mounting grooves 8 may be provided. Of course, eight mounting grooves 8 may also be provided to achieve simultaneous detection of more samples.

[0038] As Figure 3 shown, the chip 5 is a silicon-based chip, which is composed of a first material layer 506, a second material layer 507, and a heat conducting sheet 508. The first material layer 506 is a glass sheet, and the second material layer 507 is a silicon wafer. The glass sheet, silicon wafer, and heat conducting sheet 508 are stacked and connected in sequence to form a composite silicon-based chip 5. The second material layer 507 is selected as a silicon wafer. On the one hand, it is convenient to form an amplification part inside the silicon wafer through a wafer etching process. On the other hand, the temperature rise and fall efficiency of the PCR reagent can be almost the same as that of the heat source on the external device by utilizing the semiconductor heat conduction property of the silicon wafer. In other embodiments, the materials of the first material layer 506 and the second material layer 507 may also be selected from other materials, such as high thermal conductivity materials like aluminum nitride and silicon carbide. The heat conducting sheet 508 is preferably an elastic heat conducting material, which can reduce the air cavity generated between the chip 5 and the heat transfer seat of the external device.

[0039] As <000009^{8}>[[ID=I6]]shown, a serpentine tube amplification cavity 501, a liquid inlet microchannel 502, and a liquid outlet microchannel 503 are formed in the second material layer 507 (silicon wafer) through a wafer etching process. The liquid inlet microchannel 502, amplification cavity 501, and liquid outlet microchannel 503 are connected in sequence. Combining Figure 3 and Figure 4An injection hole 504 is formed in the first material layer 506 (glass sheet), and the injection hole 504 is connected to the liquid inlet microchannel 502; an outlet hole 505 is formed in the first material layer 506 (glass sheet), and the outlet hole 505 is connected to the liquid outlet microchannel 503.

[0040] like Figure 6 and Figure 7 As shown, a layer of elastic tape 6 is attached to the side of the first material layer 506 (glass sheet) of the chip 5 facing away from the second material layer 507 (silicon wafer). Two microcavities are formed within the elastic tape 6: one is a liquid inlet 601, and the other is a liquid outlet 602. The liquid inlet 601 of the elastic tape 6 communicates with the sample inlet 504 of the chip 5, while the liquid outlet 602 of the elastic tape 6 communicates with the gas outlet 505 of the chip 5. The other side of the elastic tape 6 is attached to the mounting slot 8 using a mating fixture. The liquid inlet 601 covers the outer side of the gland liquid inlet micropore 103, while the liquid outlet 602 covers the outer side of the gland liquid outlet micropore 104. This ensures that the gland liquid inlet micropore 103 is sealed and connected to the sample inlet 504 of the chip 5, and the gland liquid outlet micropore 104 is sealed and connected to the gas outlet 505 of the chip 5. Thus, the elastic tape 6 achieves a sealing effect between the pressure-covered sample inlet 101 , the pressure-covered air outlet 102 and the chip 5 .

[0041] The elastic tape 6 refers to an elastic tape, which can be a tape with sticky one side or a double-sided tape, and can be used after tearing off the release film on the surface. The elastic tape 6 is made of high-temperature resistant foam tape. In the PCR nucleic acid amplification test, there are multiple heating and cooling stages. The use of high-temperature resistant foam tape can prevent the reaction liquid from leaking into the air and causing aerosol contamination during the rapid heating and cooling process, thereby achieving a fully enclosed PCR environment. In other embodiments, other high-temperature resistant elastic tapes 6 can also be selected, such as a high-temperature resistant organic silicone layer, an acrylic adhesive layer, etc., and elastic and high-temperature resistant materials such as polyacrylate material and silicone material can also be selected as the main chemical material. The elastic tape 6 is sticky, and one side of the foam tape is attached to the glass sheet. The shape and size of the foam tape match the mounting groove 8. The other side of the foam tape is attached to the mounting groove 8 of the pressure cover 1 through tool matching, thereby fixing the chip 5 in the mounting groove 8.

[0042] Reference Figure 1 、 Figures 7 to 9 Two sealing mechanisms 2 are formed on the second surface (back surface) of the gland 1. The two sealing mechanisms 2 are arranged on both sides of the gland 1. One of the sealing mechanisms 2 is taken as an example for explanation. Figure 7 is a side perspective view of the microfluidic chip. Figure 1The AA line in the middle represents the direction of observation). The sealing mechanism 2 includes a fixed structure 201 fixed on the gland 1, and a group of sealing plugs 202 connected to the fixed structure 201. The sealing plugs 202 are silicone sealing plugs. A group of sealing plugs 202 has four silicone plugs in total, and the four silicone plugs are connected to the fixed structure 201 as a whole. The four silicone plugs are distributed on one side of two of the mounting grooves 8 on the gland 1, in groups of two, and each group corresponds to the gland inlet 101 and the gland outlet 102 of one mounting groove 8. One sealing mechanism 2 can seal two chips 5, i.e., the two gland inlets 101 and the two gland outlets 102 corresponding to the two mounting grooves 8. The material of the sealing plug 202 can also be an elastic substrate of other rubber coating layers.

[0043] Reference Figure 1 and Figure 2 , two symmetrical iron sheets 301 and 302 are embedded in the gland 1 through the injection molding process, and the two iron sheets 301 and 302 are symmetrically arranged on both sides of the gland 1. In this embodiment, the four mounting grooves 8 are distributed in the middle of the gland 1 in a field shape, and the iron sheets 301 and 302 are distributed on the outside of the four mounting grooves 8 on the gland 1. The two iron sheets 301 and 302 are used to generate suction with the magnets on the external device, and this suction is used to generate pressure on the chip 5 in the four mounting grooves 8 between the iron sheets 301 and 302, thereby squeezing the elastic tape 6 between the chip 5 and the mounting groove 8. By squeezing the elastic tape 6, the elastic buffering effect of the elastic tape 6 is exerted, and the sealing between the chip 5 and the mounting groove 8 is increased, thereby improving the sealing of the entire detection process.

[0044] Specifically, under the effect of suction between the magnet and the iron sheet 301, 302, the elastic buffer between the glass sheet and the mounting groove 8 is realized by using the elastic tape 6, so as to further improve the sealing performance between the injection hole 504 of the chip 5 and the gland liquid inlet micropore 103, and between the air outlet 505 of the chip 5 and the gland liquid outlet micropore 104. In addition, the foam tape adopted has high temperature resistance, which can prevent the reaction liquid from leaking into the air and causing aerosol pollution during rapid temperature increase and decrease. During the extrusion process, the buffering effect of the elastic tape 6 can also be used to adjust the deviation caused by the height difference of the chip 5 in the mounting groove 8, so that each chip can obtain good sealing performance. In other embodiments, the number and distribution of the iron sheet 5 can also be adjusted according to the number and distribution of the mounting groove 8. For example, four iron sheets can be symmetrically placed in pairs outside the mounting groove 8, or can also be distributed between each mounting groove 8.

[0045] like Figure 10 and Figure 11As shown, during the injection molding process, four positioning holes 4 are symmetrically provided at the four corners of the gland 1. These four positioning holes 4 are used to position the gland 1 and chip 5 on an external device. A heat conducting plate 508 is connected to an external heat transfer base 7, which is connected to a heat source. The four positioning holes 4 precisely position the heat conducting plate 508 (connected to the heat transfer base 7) on the chip 5 relative to the external heat source. This allows external heat to be transferred through the heat transfer base 7 to the heat conducting plate 508, which then transfers the heat to the second material layer 507, i.e., the silicon wafer layer. This effectively ensures that the temperature rise and fall of the amplification chamber 501 and the temperature rise and fall of the heat transfer base 7 are nearly synchronized, achieving an average temperature rise and fall of over 15°C / s. This reduces the 60-minute cycle time required for 40 PCR amplification cycles on the market to just 5 minutes. This shortens the amplification time, further improving detection efficiency and reducing testing costs.

[0046] like Figure 9 As shown, two hand-held structures 110 are symmetrically provided on both sides of the pressure cover 1, which facilitates the operator to use the microfluidic chip and makes it safer and more convenient to use it during nucleic acid amplification detection.

[0047] The structure of the microfluidic chip provided in this embodiment is as follows:

[0048] The plastic gland 1 comprises two symmetrically arranged gripping structures 110, two symmetrically arranged sealing mechanisms 2, two symmetrically arranged iron plates 301 and 302, four symmetrically arranged positioning holes 4, and four symmetrically arranged mounting slots 8. Each mounting slot 8 houses a chip 5, for a total of four chips 5. A layer of elastic tape 6 is placed between each chip 5 and the mounting slot 8, securing the chip 5 within the slot.

[0049] Each chip 5 has an amplification chamber 501 formed inside. The microfluidic chip provided in this embodiment has a total of four chips 5, i.e., four amplification chambers 501, which can detect four samples simultaneously. Taking one chip 5 as an example, the chip 5 is formed by stacking and connecting three layers of materials in sequence. The first layer is a glass sheet, the second layer is a silicon sheet, and the third layer is a heat conductive sheet 508. The glass sheet has an injection hole 504 and an air outlet 505 formed therein, and the silicon sheet has an amplification chamber 501, a liquid inlet microchannel 502, and a liquid outlet microchannel 503 formed therein. The mounting groove 8 is formed with a pressure-capped injection port 101, a pressure-capped liquid inlet micropore 103, a pressure-capped air outlet 102, and a pressure-capped liquid outlet micropore 104. The elastic tape 6 is formed with a liquid inlet hole 601 and a liquid outlet hole 602.

[0050] The gland sample inlet 101, gland liquid inlet micropore 103 (externally covered with liquid inlet hole 601), sample inlet 504, liquid inlet microchannel 502, and amplification chamber 501 are sequentially connected. The gland gas outlet 102, gland liquid outlet micropore 104 (externally covered with liquid outlet hole 602), gas outlet hole 505, liquid outlet microchannel 503, and amplification chamber 501 are sequentially connected. Each sealing mechanism 2 has four corresponding silicone plugs, two in a group, each group corresponding to one gland sample inlet 101 and one gland gas outlet 102. During use, the gland sample inlet 101 and gland gas outlet 102 are sealed with the sealing plug 202 on the sealing mechanism 2, thereby forming a fully enclosed PCR nucleic acid amplification detection environment.

[0051] The working principle of this embodiment is as follows: the sample enters the gland sample inlet 101 on the gland 1 into the gland liquid inlet micropore 103, and then enters the sample inlet 504 of the chip 5 from the gland liquid inlet micropore 103 (the gland liquid inlet micropore 103 and the sample inlet 504 are sealed by a foam tape provided with a liquid inlet hole 601), flows from the sample inlet 504 into the liquid inlet microchannel 502 of the chip 5, and then flows from the liquid inlet microchannel 502 into the serpentine tube amplification chamber 501. The gas originally trapped in the serpentine tube amplification chamber 501 is discharged from the liquid outlet microchannel 503 and the air outlet 505 of the chip 5 to the gland liquid outlet micropore 104, and finally discharged from the gland air outlet 102. At this time, the silicone plugs on the sealing plug 202 block the gland sample inlet 101 and the gland air outlet 102, thereby completing the sealing of the chip 5, forming a fully enclosed PCR reaction environment, and starting the reaction. The heat transfer base 7 heats the heat conducting plate 508, thereby cyclically raising and lowering the temperature of the amplification chamber 501. The position corresponding to the chip 5 in the mounting slot 8 serves as the fluorescence detection area for external equipment. After the sample completes the amplification reaction in the serpentine amplification chamber 501 of the chip 5, fluorescence detection of the sample can be performed using external equipment.

[0052] The above embodiment describes a case in which four silicon-based chips 5 are assembled with a pressure cap 1, but the technical solution protected by the present invention is not limited thereto. The microfluidic chip design provided by the present invention has great amplification, and the silicon-based chip 5 can be selectively assembled to achieve 1 to 4 sample detection. In other embodiments, more amplification cavities 501 can be set in the silicon wafer of each chip 5, such as four serpentine tube amplification cavities 501, which can simultaneously achieve amplification detection of 1 to 16 samples. Even more samples can be detected, that is, the design of the silicon wafer and the pressure cap 1 is diverse.

[0053] Most of the existing microfluidic chip consumables are made of a single material, such as pure glass material or pure PC material. In one embodiment provided by the present invention, the consumables of the microfluidic chip use a variety of materials such as PC plastic, elastic foam glue material, glass, Si material, silica gel, iron sheet, etc., and a multi-material combination method is used to prepare the target microfluidic chip, and a fully enclosed PCR amplification environment is achieved. The present invention provides a closed chip consumable that combines silicon-based materials and polymer materials. The extracted nucleic acid or the nucleic acid that does not require extraction is added to the amplification cavity 501 of the chip 5 for amplification and detection, thereby realizing a closed-tube nucleic acid amplification detection. The microfluidic chip is prepared by adopting a low-cost wafer etching process and an injection molding production cap. The production cost of the product is low, suitable for large-scale application and promotion, and the product of the present invention has completed mass production.

[0054] In one embodiment of the present invention, a silicone sealing plug is used to seal the gland sample inlet 101 and the gland air outlet 102, and the elastic buffering performance of the foam tape is used to seal the gap between the gland 1 (installation slot 8) and the chip 5. An iron sheet is embedded in the gland 1 and a magnet assembled on the PCR device generates suction to squeeze the foam tape between the chip 5 and the gland 1, thereby achieving the sealing of the chip 5 cavity. The microfluidic chip provided by the present invention does not require a separate cover plate to be provided on the gland 1, and can achieve a fully enclosed PCR amplification environment, so that the reaction liquid will not leak into the air during the rapid heating and cooling process to cause aerosol contamination. The structure that achieves a closed PCR amplification environment completely isolates the interior of the chip 5 from the external environment, so it will not be affected by external contamination and has a wide range of applicable scenarios.

[0055] In one embodiment of the present invention, the positioning hole 4 is used to position the heat conducting sheet 508 and the heat transfer seat 7 in contact with the heat conducting sheet 508, and the heat conducting sheet 508 is positioned at the heat source of the external device. The heat conducting sheet 508 is in direct contact with the silicon chip formed with the serpentine tube amplification cavity 501, and covers the serpentine tube amplification cavity 501 in the silicon chip. The rapid semiconductor thermal technology of the device can be combined to provide a fast heating and cooling heat source, and then the heat is transferred to the amplification cavity 501 of the chip 5 through the heat transfer seat 7 with high thermal conductivity, such as a copper sheet. Since the amplification cavity 501 is a three-dimensional silicon material serpentine tube, the heating and cooling efficiency of the PCR reagent can be almost synchronized with the heating and cooling frequency of the heat transfer seat 7, thereby improving the heat conduction efficiency. Forty thermal cycle amplifications can be completed within five minutes, achieving rapid heating and cooling. The 60 minutes required for 40 PCR amplification cycles in the previous market can be reduced to 5 minutes, greatly shortening the time required for PCR testing. The elastic buffer design of the foam tape and the heat conducting sheet 508 can be used for double buffering so that the silicon chip can effectively contact the copper heat transfer seat 7 on the PCR device without generating an air cavity, thereby improving the heat transfer effect.

[0056] The above embodiments are only intended to help understand the method and core concept of the present invention. It should be noted that, without departing from the principles of the present invention, a number of improvements and modifications may be made to the present invention by those skilled in the art, and such improvements and modifications also fall within the scope of protection of the claims of the present invention.

Claims

1. A closed microfluidic chip for nucleic acid amplification detection, characterized by: The present invention comprises a pressure cover and at least one chip; the first surface of the pressure cover is provided with at least one mounting groove, and the second surface is provided with at least one pressure cover sampling port and pressure cover air outlet corresponding to each mounting groove; at least one amplification part is formed inside the chip, and the chip is installed in the mounting groove; an elastic tape is provided at the bottom of the mounting groove, and the elastic tape fixes the chip in the mounting groove; at least two microcavities are formed inside the elastic tape, and the at least two microcavities seal the connection between the pressure cover sampling port and the amplification part, and the connection between the pressure cover air outlet and the amplification part; the second surface of the pressure cover is provided with at least one sealing mechanism, and the sealing mechanism is used to seal the pressure cover sampling port and the pressure cover air outlet; The first surface of the gland is provided with four mounting grooves, and the second surface is provided with a gland inlet and a gland outlet corresponding to each mounting groove; a chip is installed in each mounting groove, and an amplification part is formed inside each chip; The first surface of the gland is provided with at least one iron sheet, and the iron sheet interacts with a magnet on an external device to generate suction; The chip includes a first material layer and a second material layer, wherein the first material layer is located between the elastic tape and the second material layer; the first material layer is a sample inlet and outlet circulation layer, and the second material layer is an amplification layer; The chip further includes a heat conducting sheet, the heat conducting sheet is located on a surface of the second material layer facing away from the first material layer, and the heat conducting sheet is connected to an external heat source; At least one sample inlet and an air outlet are formed inside the first material layer, and at least one liquid inlet microchannel, an amplification chamber and a liquid outlet microchannel are formed inside the second material layer; the pressure cover sample inlet, sample inlet, liquid inlet microchannel and amplification chamber are connected in sequence, and the pressure cover air outlet, air outlet, liquid outlet microchannel and amplification chamber are connected in sequence.

2. A closed microfluidic chip for nucleic acid amplification detection according to claim 1, characterized in that: There are two iron sheets arranged on the first surface of the gland, and the two iron sheets are symmetrically arranged on both sides of the gland and the periphery of the mounting groove.

3. The closed microfluidic chip for nucleic acid amplification detection according to claim 1, characterized in that: The sealing mechanism comprises a fixing structure fixed on the gland and a plurality of sealing plugs connected to the fixing structure, wherein the sealing plugs are used to seal the gland inlet and the gland outlet.

4. A closed microfluidic chip for nucleic acid amplification detection according to claim 3, characterized in that: The material of the sealing plug is silicone or an elastic base material with a rubber coating.

5. The closed microfluidic chip for nucleic acid amplification detection according to claim 1, characterized in that: The elastic tape is a high-temperature resistant foam tape.

6. The closed microfluidic chip for nucleic acid amplification detection according to claim 1, characterized in that: The amplification chamber is a serpentine tube amplification chamber.

7. The closed microfluidic chip for nucleic acid amplification detection according to claim 6, characterized in that: The first material layer is a glass sheet, and the second material layer is a silicon sheet.

8. The closed microfluidic chip for nucleic acid amplification detection according to claim 1, characterized in that: The pressure cover is provided with at least one positioning hole, and the pressure cover is positioned on the external device through the positioning hole.

9. The closed microfluidic chip for nucleic acid amplification detection according to claim 1, characterized in that: Hand-held structures are symmetrically arranged on both sides of the pressure cover.

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