Microfluidic chips, nucleic acid detection cartridges and nucleic acid extraction cartridges

Abstract

This application discloses a microfluidic chip, a nucleic acid detection cartridge, and a nucleic acid extraction cartridge, comprising a chip body and thin films respectively disposed on two surfaces of the chip body. The chip body is provided with n first microfluidic channels and m second microfluidic channels. The first microfluidic channels include m sequentially connected first reaction chambers, and the second microfluidic channels include n sequentially connected second reaction chambers. The first and second reaction chambers are connected in a one-to-one correspondence, and a phase change material isolator is disposed at the connection point. m and n are both natural numbers not less than 2. One of the first and second microfluidic channels is used to inject a template-free nucleic acid amplification system, and the other is used to inject the sample to be tested. This application can achieve simultaneous detection of multiple nucleic acid targets in multiple samples to be tested through simple sample addition, greatly simplifying the operation process for technicians and reducing consumable costs.

Description

Technical Field

[0001] This application relates to the field of nucleic acid detection technology, and more specifically, to a microfluidic chip, a nucleic acid detection cartridge, and a nucleic acid extraction cartridge. Background Technology

[0002] In vitro diagnostics, such as cell detection, biochemical detection, immunoassay, and nucleic acid amplification, often require simultaneous detection and analysis of the same indicator in multiple samples or multiple indicators in the same sample. Microfluidics is a technology that uses structures such as microchannels and microcavities to control microfluidics to complete various biological and chemical processes. The use of microfluidic chips to perform multi-indicator biochemical reactions or detections offers advantages such as ease of automation, low reagent consumption, and low overall detection cost, making it highly suitable for research and applications in in vitro diagnostics.

[0003] Faced with the ever-increasing demand for pathogen detection, it is often difficult to balance sample throughput with the number of detection targets. Current technologies can only meet the needs of detecting multiple pathogens in the same sample, or only meet the needs of detecting one pathogen in multiple samples.

[0004] Therefore, how to simultaneously detect multiple nucleic acid targets in multiple test samples is a problem that urgently needs to be solved by those skilled in the art. Summary of the Invention

[0005] In view of this, the purpose of this application is to provide a microfluidic chip that can simultaneously detect multiple nucleic acid targets in multiple test samples;

[0006] Another objective of this application is to provide a nucleic acid detection cartridge and a nucleic acid extraction cartridge for use with the aforementioned microfluidic chip.

[0007] To achieve the above objectives, this application provides the following technical solution:

[0008] The first aspect of this application provides a microfluidic chip, including a chip body and thin films respectively disposed on a first surface and a second surface of the chip body, at least one of the thin films being a transparent thin film, and the chip body being provided with n first microfluidic channels extending along a first direction and m second microfluidic channels extending along a second direction, wherein the first direction and the second direction intersect.

[0009] The first microfluidic channel includes m first reaction chambers that are sequentially connected through a first connecting portion, and the second microfluidic channel includes n second reaction chambers that are sequentially connected through a second connecting portion. The first connecting portion and the second connecting portion are connected to at least one side surface of the chip body.

[0010] Along the thickness direction of the chip body, the first reaction chamber and the second reaction chamber are connected in a one-to-one correspondence, and a phase change material isolator is provided at the connection point, where m and n are both natural numbers not less than 2;

[0011] One of the first microfluidic conduit and the second microfluidic conduit is used to inject the nucleic acid amplification system, and the other is used to inject the sample to be tested.

[0012] In one possible implementation, the opening side of the first reaction chamber is located on the first surface, and the opening side of the second reaction chamber is located on the second surface.

[0013] In one possible implementation, the thin film disposed on the first surface is a first thin film, and the thin film disposed on the second surface is a second thin film.

[0014] The first film is adhered to the first surface, and the second film is adhered to the second surface.

[0015] In one possible implementation, the chip body is made of a polymer material, the first thin film is a transparent polymer film, and the second thin film is a polymer film or a metal film.

[0016] In one possible implementation, the phase change material insulator is made of one or both of paraffin and organic gel.

[0017] In one possible implementation, the organic gel includes at least one of a first organic gel and a second organic gel;

[0018] The first organic gel is a pressed oil organic gel containing 0.1%-10%wt dodecyl stearic acid;

[0019] The second organic gel is a pressed oil organic gel containing 0.1%-10%wt octacosanol.

[0020] In one possible implementation, the first connecting portion includes a first connecting pipe and a first recess. The first inlet and the first outlet of the first microfluidic pipe are both connected to the adjacent first reaction chamber through the corresponding first connecting pipe. Any two adjacent first reaction chambers are connected through the first recess between them. The opening side of the first recess is connected to at least one side surface of the chip body.

[0021] In one possible implementation, the second connecting portion includes a second connecting pipe and a second recess. The second inlet and the second outlet of the second microfluidic pipe are both connected to the adjacent second reaction chamber through the corresponding second connecting pipe. Any two adjacent second reaction chambers are connected through the second recess between them. The opening side of the second recess is connected to at least one side surface of the chip body.

[0022] In one possible implementation, the opening side of the first recess and the opening side of the second recess are connected to the same side surface of the chip body.

[0023] In one possible implementation, the opening side of the first recess and the opening side of the second recess are both connected to a side surface of the chip body adjacent to the second reaction chamber.

[0024] In one possible implementation, the opening side of the first recess, the opening side of the second recess, and the opening side of the second reaction chamber are connected to the same side surface of the chip body.

[0025] In one possible implementation, the opening side of the first recess, the opening side of the second recess, and the opening side of the second reaction chamber are all connected to the second surface, and the opening side of the first reaction chamber is connected to the first surface.

[0026] A connecting hole is provided on the outer side of the first recess. One end of the connecting hole is connected to the first recess, and the other end extends toward the first surface and is connected to the corresponding first reaction chamber through a corresponding first connecting pipe.

[0027] In one possible implementation, the opening side of the first recess and the opening side of the second recess are respectively connected to different side surfaces of the chip body.

[0028] In one possible implementation, the first connecting portion includes a first connecting pipe, and any two adjacent ones of the first inlet of the first microfluidic pipe, each of the first reaction chambers and the first outlet of the first microfluidic pipe are connected through the corresponding first connecting pipe, and each of the first connecting pipes is connected to the first surface.

[0029] The second connecting portion includes a second connecting pipe. Any two adjacent pairs of the second inlet of the second microfluidic pipe, each of the second reaction chambers, and the second outlet of the second microfluidic pipe are connected through a corresponding second connecting pipe. Each of the second connecting pipes is connected to the second surface of the chip body.

[0030] In use, the microfluidic chip provided in this application first fixes a phase change material isolator at the connection between the first and second reaction chambers, separating them at room temperature. Then, two thin films are fixed to the first and second surfaces of the chip body. Next, different nucleic acid amplification systems and test samples are added using a pipette from the inlets of the first and second microfluidic channels, respectively. Because the first and second reaction chambers are blocked by the phase change material isolator, a temporary physical isolation is achieved between the nucleic acid amplification system and the test sample. The inlets and outlets of the first and second microfluidic channels can be sealed with single-sided adhesive tape. Then, using an external device, the first connecting portion connecting each first reaction chamber and the second connecting portion connecting each second reaction chamber are pressed, causing the thin films to be squeezed into the corresponding first and second connecting portions, thus completely separating the first and second reaction chambers.

[0031] Microfluidic chips can be used to break down the phase change material separator through centrifugation, ultrasound, or heating, thereby connecting the first reaction chamber with the corresponding second reaction chamber. This allows the test sample to be mixed with the nucleic acid amplification system, creating a mixture of different nucleic acid amplification systems and different test samples. Furthermore, the test sample in each reaction chamber (composed of the connected first and second reaction chambers) is mixed with only one primer probe. This enables nucleic acid detection of at least two different targets targeting two different test samples to be performed on a single microfluidic chip. Finally, the microfluidic chip can perform a nucleic acid amplification reaction, which can be LAMP or PCR.

[0032] The microfluidic chip disclosed in this application can simultaneously detect multiple nucleic acid targets in multiple test samples through simple sample addition, which greatly simplifies the operation process for technicians, reduces the cost of consumables, and is of great significance for clinical scenarios such as high-throughput rapid nucleic acid screening.

[0033] A second aspect of this application provides a nucleic acid detection cartridge for use with the microfluidic chip described above, comprising:

[0034] A heating device is used to heat the microfluidic chip;

[0035] A pressing mechanism is used to press the microfluidic chip onto the heating device;

[0036] The micro-pillars are pressed to pass through the heating device and correspond one-to-one with each of the first and second recesses.

[0037] In one possible implementation, the nucleic acid test kit also includes:

[0038] The base has each of the pressing micro-pillars disposed on it. The base has a first positioning post, and an elastic reset member is sleeved on the first positioning post. The heating device has a first positioning hole that slides with each of the first positioning posts. One end of the elastic reset member abuts against the base, and the other end abuts against the heating device.

[0039] A support member is disposed on the base;

[0040] The top cover is hinged to the support member at one end. The pressing mechanism is disposed on the top cover. When the top cover is closed, the pressing mechanism presses the microfluidic chip onto the heating device. When the top cover is open, the pressing mechanism disengages from the microfluidic chip.

[0041] In one possible implementation, the pressing mechanism includes:

[0042] The first connecting rod has its first end hinged to the upper cover;

[0043] The second link and the third link are both hinged to the second end of the first link;

[0044] A support rod is fixed to the support member, and a sliding sleeve is provided on the support rod;

[0045] The fourth link slides with the sliding sleeve. One end of the fourth link is hinged to the second end of the third link, and the other end is used to press the microfluidic chip. The second end of the second link is hinged to the support rod.

[0046] In one possible implementation, chip pressing portions are provided at both ends of the chip body, and there are two pressing mechanisms, each of which is used to press on the chip pressing portion.

[0047] In one possible implementation, a second positioning post is provided on the base, and the heating device has a heating device positioning hole that slides with the second positioning post.

[0048] The microfluidic chip has a chip positioning hole that slides with the second positioning post.

[0049] In one possible implementation, the heating device includes a heating film and a heat transfer plate, the heating film being adhered to the heat transfer plate, and the heat transfer plate being arranged closer to the microfluidic chip than the heating film.

[0050] In one possible implementation, an imaging device is also included, which is used to acquire fluorescence signals or color signals that change over time.

[0051] The nucleic acid test kit provided in this application is used in conjunction with the aforementioned microfluidic chip for testing, and therefore possesses all the technical effects of the aforementioned microfluidic chip, which will not be elaborated upon here.

[0052] A third aspect of this application provides a nucleic acid extraction cartridge for use with a microfluidic chip as described in any of the preceding claims, comprising:

[0053] The nucleic acid extraction cartridge body has a lysis chamber, a washing chamber and an elution chamber connected sequentially from top to bottom. The nucleic acid extraction cartridge body also has a sample chamber at the same height as the lysis chamber. The sample chamber and the lysis chamber are connected through a sample injection hole at the bottom. A vent is provided at the upper part of the lysis chamber.

[0054] The sample piston push rod and the fluid piston push rod are provided, wherein the sample piston push rod is used to cooperate with the sample cavity, and the fluid piston push rod is used to cooperate with the lysis cavity;

[0055] A soft plug is used to seal the elution chamber;

[0056] A hollow needle is inserted into the soft plug from the lower side, with the upper end of the hollow needle located below the upper surface of the soft plug. The lower end of the hollow needle is used to communicate with the inlet of one of the first microfluidic channel and the second microfluidic channel.

[0057] In one possible implementation, the vent is sealed by a hydrophobic and breathable membrane;

[0058] And / or,

[0059] When the sample piston push rod is not engaged with the sample cavity, the top opening of the sample cavity is sealed by a sealing layer.

[0060] In one possible implementation, the pyrolysis chamber and the washing chamber are connected through a first through hole, and the washing chamber and the elution chamber are connected through a second through hole;

[0061] The diameter of the first through hole is smaller than the diameter of the pyrolysis chamber and the cleaning chamber, and the diameter of the second through hole is smaller than the diameter of the cleaning chamber and the elution chamber. The first through hole and the second through hole are filled with organic reagents that are incompatible with the aqueous phase reagents.

[0062] In one possible implementation, the organic reagent is mineral oil, silicone oil, phase change paraffin, or organic gel.

[0063] The nucleic acid extraction cartridge provided in this application is used in conjunction with the aforementioned microfluidic chip, and therefore possesses all the technical effects of the aforementioned microfluidic chip, which will not be elaborated upon here. Attached Figure Description

[0064] To more clearly illustrate the technical solutions in the embodiments of this application or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0065] Figure 1 This is an exploded view of the microfluidic chip disclosed in the embodiments of this application;

[0066] Figure 2 This is a perspective view of the chip body disclosed in the embodiments of this application;

[0067] Figure 3 This is a schematic diagram of the structure of the first surface of the chip body disclosed in an embodiment of this application;

[0068] Figure 4 This is a schematic diagram of the structure of the second surface of the chip body disclosed in the embodiments of this application;

[0069] Figure 5 This is a top view of the first surface of the chip body disclosed in the embodiments of this application;

[0070] Figure 6 for Figure 5 Sectional view along AA;

[0071] Figure 7 for Figure 5 Sectional view along BB;

[0072] Figure 8 This is a cross-sectional view of the chip body at the first and second reaction chambers disclosed in the embodiments of this application;

[0073] Figure 9 A cross-sectional view of the connection between the first reaction chamber and the second reaction chamber disclosed in the embodiments of this application, which is filled with a phase change material insulator.

[0074] Figure 10 This is a schematic diagram of the structure of the first surface of the chip body disclosed in another embodiment of this application;

[0075] Figure 11 This is a schematic diagram of the structure of the second surface of the chip body disclosed in another embodiment of this application;

[0076] Figure 12 This is a perspective view of the chip body disclosed in another embodiment of this application;

[0077] Figure 13 This is a schematic diagram of the structure of the first surface of the chip body disclosed in another embodiment of this application;

[0078] Figure 14 This is a schematic diagram of the structure of the second surface of the chip body disclosed in another embodiment of this application;

[0079] Figure 15 This is a perspective view of the chip body disclosed in another embodiment of this application;

[0080] Figure 16 This is a schematic diagram of the structure in which the reaction chamber of the first surface of the microfluidic chip disclosed in the embodiments of this application is blocked;

[0081] Figure 17 This is a schematic diagram of the structure in which the reaction chamber on the second surface of the microfluidic chip disclosed in the embodiments of this application is blocked;

[0082] Figure 18 This is a perspective view showing the reaction chamber of the microfluidic chip disclosed in the embodiments of this application being blocked;

[0083] Figure 19 This is a schematic diagram illustrating the working mode of multi-sample, multi-index nucleic acid detection using a microfluidic chip disclosed in this application.

[0084] Figure 20 for Figure 19 Sectional view along AA;

[0085] Figure 21 for Figure 19 Sectional view along BB;

[0086] Figure 22 This is a schematic diagram of the structure of the nucleic acid test card box disclosed in this application when the top cover is in the closed state;

[0087] Figure 23 This is a schematic diagram of the structure of the nucleic acid test card box disclosed in this application when the top cover is in the open state;

[0088] Figure 24 This is a schematic diagram of the internal structure of the nucleic acid test card box disclosed in this application when the top cover is in the open state;

[0089] Figure 25 This is a schematic diagram of the internal structure of the nucleic acid test card box disclosed in this application when the top cover is in the closed state;

[0090] Figure 26 This is an exploded view of the internal components of the nucleic acid test kit disclosed in the embodiments of this application;

[0091] Figure 27 This is a side view of the pressing mechanism disclosed in the embodiments of this application;

[0092] Figure 28This is an exploded view of a microfluidic chip disclosed in another embodiment of this application;

[0093] Figure 29 This is a schematic diagram of the structure of the nucleic acid extraction cartridge and the microfluidic chip disclosed in the embodiments of this application after being combined;

[0094] Figure 30 This is an exploded view of the nucleic acid extraction cartridge disclosed in the embodiments of this application;

[0095] Figure 31 This is a top view of the nucleic acid extraction cartridge and microfluidic chip disclosed in the embodiments of this application after being combined;

[0096] Figure 32 for Figure 31 Sectional view along CC;

[0097] Figure 33 This is a diagram illustrating the process of extracting nucleic acid using the nucleic acid extraction cartridge disclosed in this application.

[0098] The meanings of the various reference numerals in the figure are as follows:

[0099] 100 - Chip body; 110 - First surface; 111 - First microfluidic channel; 1111 - First inlet; 1112 - First reaction chamber; 1113 - First outlet; 1114 - First connecting channel; 1115 - Connecting hole; 1116 - First recess; 120 - Second surface; 121 - Second microfluidic channel; 1211 - Second inlet; 1212 - Second reaction chamber; 1213 - Second outlet; 1214 - Second connecting channel; 1215 - Second recess; 130 - Chip positioning hole; 140 - Chamber connecting hole; 150 - Chip pressing part;

[0100] 200 - First thin film body;

[0101] 300 - Second thin film body;

[0102] 400-Nucleic acid test cartridge; 410-Top cover; 411-Imaging device; 412-Hinge; 420-Base; 421-First positioning post; 4211-Elastic reset component; 4212-Second positioning post; 422-Pressing micro-post; 423-Heating film; 4231-First heating device positioning hole; 4232-Heating film through hole; 424-Heat transfer plate; 4241-First positioning hole; 4242-Second heating device positioning hole; 4243-Heat transfer plate through hole; 431-Mounting part; 432-First connecting rod; 433-Second connecting rod; 434-Fourth connecting rod; 4341-Pressing seat; 435-Third connecting rod; 436-Support rod; 4361-Sliding sleeve; 440-Support component;

[0103] 500 - Nucleic acid extraction cartridge; 510 - Nucleic acid extraction cartridge body; 511 - Lysis chamber; 512 - Sample chamber; 513 - Vent; 514 - Sample injection port; 515 - First through hole; 516 - Washing chamber; 517 - Second through hole; 518 - Elution chamber; 520 - Sample piston push rod; 530 - Fluid piston push rod; 540 - Soft stopper; 550 - Hollow needle;

[0104] 600-Magnet. Detailed Implementation

[0105] This application discloses a microfluidic chip capable of simultaneously detecting multiple nucleic acid targets in multiple test samples;

[0106] This application also discloses a nucleic acid detection cartridge and a nucleic acid extraction cartridge for use with the above-mentioned microfluidic chip.

[0107] Hereinafter, embodiments will be described with reference to the accompanying drawings. Furthermore, the embodiments shown below do not limit the scope of the application as described in the claims. Additionally, the complete composition represented in the embodiments below is not limited to what is necessary as the solution to the application described in the claims. It should be noted that, for ease of description, only the parts relevant to the application are shown in the drawings. Unless otherwise specified, the embodiments and features described in this application can be combined with each other.

[0108] like Figure 1 As shown, the microfluidic chip disclosed in this application includes a chip body 100 and thin films respectively disposed on a first surface 110 and a second surface 120 of the chip body 100. The first surface 110 and the second surface 120 of the chip body 100 are two surfaces arranged opposite to each other. Since fluorescence reaction needs to be detected during detection, at least one thin film needs to be a transparent film to facilitate observation and detection.

[0109] like Figures 2-4 As shown, the chip body 100 is provided with n first microfluidic channels 111 extending along a first direction and m second microfluidic channels 121 extending along a second direction, the first and second directions intersecting. For example, the first and second directions can be designed to be perpendicular. It should be noted that the first and second directions can also be designed to be at other angles, as long as they can intersect each other.

[0110] One of the first microfluidic conduit 111 and the second microfluidic conduit 121 is used to inject a template-free nucleic acid amplification system, and the other is used to inject the sample to be tested. The nucleic acid amplification system includes primers, polymerases, etc., required for the amplification process, as well as fluorescent probes, etc., required for detection. Those skilled in the art can determine the specific substances included in the nucleic acid amplification system according to their detection needs.

[0111] Taking the injection of a test sample into the first microfluidic channel 111 and the injection of a nucleic acid amplification system into the second microfluidic channel 121 as an example, n first microfluidic channels 111 can each inject n different test samples, and m second microfluidic channels 121 can each inject m different nucleic acid amplification systems. Those skilled in the art will understand that in specific operations, the number of test samples may be less than n, and the corresponding number of nucleic acid amplification systems may be less than m. That is, depending on the number of test samples and the different types and numbers of nucleic acid targets, some first microfluidic channels 111 and second microfluidic channels 121 may be left unused. n and m represent the upper limits of the number of test samples and the number of nucleic acid targets, respectively.

[0112] Both m and n are natural numbers not less than 2, ensuring that the microfluidic chip can perform nucleic acid detection on at least two samples for two different targets. Of course, those skilled in the art can choose the actual values ​​of m and n according to their needs. For example, depending on the values ​​of m and n, the microfluidic chip can perform nucleic acid detection on eight samples for eight different targets. It can also perform nucleic acid detection on four samples for eight targets, six samples for eight targets, sixteen samples for sixteen targets, and so on. This embodiment does not limit the specific values ​​of m and n.

[0113] The first microfluidic conduit 111 includes m first reaction chambers 1112 that are sequentially connected through a first connecting portion. The second microfluidic conduit 121 includes n second reaction chambers 1212 that are sequentially connected through a second connecting portion.

[0114] Those skilled in the art will understand that the first microfluidic channel 111 has an inlet and an outlet at each end, and the second microfluidic channel 121 also has an inlet and an outlet at each end. The inlet is the injection port, used to inject a solution (such as a sample to be tested or a nucleic acid amplification system). Under the action of microfluidics, the solution enters through the inlet and fills the entire microfluidic channel (first microfluidic channel 111, second microfluidic channel 121). The outlet is for venting, allowing the solution to flow.

[0115] For ease of understanding, the inlet and outlet of the first microfluidic channel 111 and the second microfluidic channel 121 are distinguished. The inlet and outlet of the first microfluidic channel 111 are defined as the first inlet 1111 and the first outlet 1113, respectively; the inlet and outlet of the second microfluidic channel 121 are defined as the second inlet 1211 and the second outlet 1213, respectively.

[0116] like Figures 5-9As shown, along the thickness direction of the chip body 100, the first reaction chamber 1112 and the second reaction chamber 1212 are connected in a one-to-one correspondence, and the first reaction chamber 1112 and the second reaction chamber 1212 are arranged vertically along the thickness direction of the chip body 100. The first reaction chamber 1112 and the second reaction chamber 1212 are interconnected to form a reaction chamber. It should be noted that, except for the interconnection between the first reaction chamber 1112 and the second reaction chamber 1212, the other parts of the first microfluidic channel 111 and the second microfluidic channel 121 are isolated from each other.

[0117] Taking n=4 and m=4 as an example, the different first reaction chambers 1112 and the different second reaction chambers 1212 are distinguished by letters. The first reaction chambers 1112 of the four first microfluidic channels 111 are designated as first reaction chamber a, first reaction chamber b, first reaction chamber c, and first reaction chamber d, with a total of four first reaction chambers 1112 for each first microfluidic channel 111; the second reaction chambers 1212 of the four second microfluidic channels 121 are designated as second reaction chamber a, second reaction chamber b, second reaction chamber c, and second reaction chamber d, with a total of four first reaction chambers 1112 for each second microfluidic channel 121.

[0118] Four first reaction chambers 'a' are combined with second reaction chambers 'a', 'b', 'c', and 'd' to form four reaction chambers; four first reaction chambers 'b' are combined with second reaction chambers 'a', 'b', 'c', and 'd' to form four reaction chambers; four first reaction chambers 'c' are combined with second reaction chambers 'a', 'b', 'c', and 'd' to form four reaction chambers; and four first reaction chambers 'd' are combined with second reaction chambers 'a', 'b', 'c', and 'd' to form four reaction chambers. This results in a total of 16 reaction chambers in different combinations. Therefore, based on four test samples and four nucleic acid amplification systems, 16 different combinations can be formed, allowing for simultaneous nucleic acid detection of four targets on four test samples.

[0119] When n and m take other values, other numbers of reaction chambers can be formed according to the above example, which will not be elaborated on in this article.

[0120] A phase change material isolator is disposed at the connection between the first reaction chamber 1112 and the second reaction chamber 1212. The phase change material isolator can isolate the first reaction chamber 1112 and the second reaction chamber 1212 at room temperature, preventing them from communicating with each other and preventing the sample to be tested from mixing with the nucleic acid amplification system. The first reaction chamber 1112 and the second reaction chamber 1212 can be connected through a chamber communication hole 140. The diameter of the chamber communication hole 140 is smaller than the diameter of the first reaction chamber 1112 and the second reaction chamber 1212, making it easier to fix the phase change material isolator within the chamber communication hole 140.

[0121] Those skilled in the art will understand that each reaction chamber can form a unique sample-target pair, meaning that there are no two reaction chambers with completely identical sample-target pairs. When injecting the test sample and nucleic acid amplification system, it is necessary to ensure that the test sample and nucleic acid amplification system can fill the entire microfluidic channel (first microfluidic channel 111, second microfluidic channel 121) through the corresponding inlet. That is, all the first reaction chambers 1112 of any first microfluidic channel 111 need to be in a connected state, and correspondingly, all the second reaction chambers 1212 of any second microfluidic channel 121 need to be in a connected state.

[0122] The connection between the first reaction chamber 1112 and the second reaction chamber 1212 is isolated by a phase change material isolator, which can prevent the sample to be tested in a certain first reaction chamber 1112 from passing through its corresponding second reaction chamber 1212, and then through the second reaction chamber 1212 into other second reaction chambers 1212 located in the same second microfluidic channel 121, and then through other second reaction chambers 1212 into the corresponding first reaction chamber 1112, so as to prevent the samples to be tested from being mixed and contaminated.

[0123] Similarly, the connection between the first reaction chamber 1112 and the second reaction chamber 1212 is isolated by a phase change material isolator, which can also prevent the nucleic acid amplification system in a certain second reaction chamber 1212 from passing through its corresponding first reaction chamber 1112, and then through the first reaction chamber 1112 into other first reaction chambers 1112 located in the same first microfluidic channel 111, and then through other first reaction chambers 1112 into the corresponding second reaction chamber 1212, thus preventing mixed contamination of the various nucleic acid amplification systems.

[0124] The first and second connecting portions are connected to at least one side surface of the chip body 100, so that by pressing down the thin film body on the first surface 110 and the second surface 120, the thin film body can be embedded in the first and second connecting portions and cut off the first and second connecting portions, thereby cutting off the communication between each reaction chamber and other reaction chambers and preventing the solutions in each reaction chamber from mixing and contaminating each other.

[0125] like Figures 19-21As shown, in the microfluidic chip disclosed in this application embodiment, during use, a phase change material isolator is first fixed at the connection between the first reaction chamber 1112 and the second reaction chamber 1212, separating the first reaction chamber 1112 and the second reaction chamber 1212 at room temperature. Then, two thin films are fixed to the first surface 110 and the second surface 120 of the chip body 100. Afterwards, nucleic acid amplification systems containing different primer probes and test samples are added using a pipette from the inlets of the first microfluidic channel 111 and the second microfluidic channel 121, respectively. Vertically arranged dashed lines represent multiple microfluidic channels for adding nucleic acid amplification systems, with different colors representing nucleic acid amplification systems containing different primer probes; each microfluidic channel contains a different target for its added nucleic acid amplification system. Horizontally arranged solid lines represent multiple microfluidic channels for adding test samples, with different colors representing different test samples.

[0126] Because the first reaction chamber 1112 and the second reaction chamber 1212 are blocked by the phase change material separator, a temporary physical isolation is achieved between the nucleic acid amplification system and the sample to be tested. The inlet and outlet of the first microfluidic channel 111 and the second microfluidic channel 121 can be sealed with single-sided adhesive. Then, using an external device, the first connecting portion connecting each of the first reaction chambers 1112 and the second connecting portion connecting each of the second reaction chambers 1212 are pressed, causing the film to be squeezed into the corresponding first and second connecting portions, thus completely separating each of the first reaction chambers 1112 and each of the second reaction chambers 1212.

[0127] The microfluidic chip can be subjected to centrifugation, ultrasonication, heating, etc., to destroy the phase change material insulator, thereby connecting the first reaction chamber 1112 with the corresponding second reaction chamber 1212. This allows the sample to be tested to be mixed with the nucleic acid amplification system, resulting in a mixture of different nucleic acid amplification systems and different samples to be tested. Furthermore, the sample to be tested within each reaction chamber (composed of the connected first reaction chamber 1112 and second reaction chamber 1212) is mixed with only one primer probe, enabling nucleic acid detection of at least two different targets for two different samples to be tested on a single microfluidic chip. Finally, the microfluidic chip can perform a nucleic acid amplification reaction, which can be LAMP (Loop-mediated isothermal amplification) or PCR (Polymerase Chain Reaction).

[0128] The microfluidic chip disclosed in this application can simultaneously detect multiple nucleic acid targets in multiple test samples through simple sample addition, which greatly simplifies the operation process for technicians, reduces the cost of consumables, and is of great significance for clinical scenarios such as high-throughput rapid nucleic acid screening.

[0129] In one specific embodiment of this application, the opening side of the first reaction chamber 1112 is located on the first surface 110, and the opening side of the second reaction chamber 1212 is located on the second surface 120. This allows the first reaction chamber 1112 and the second reaction chamber 1212 to be processed from both sides of the chip body 100, facilitating the processing of the chip body 100. The first reaction chamber 1112 and the second reaction chamber 1212, which are in communication with the atmosphere, are sealed by a thin film body, ensuring that liquid can flow within the first microfluidic channel 111 and the second microfluidic channel 121.

[0130] For ease of understanding, the thin film disposed on the first surface 110 is defined as the first thin film 200, and the thin film disposed on the second surface 120 is defined as the second thin film 300. The first thin film 200 is adhered to the first surface 110, and the second thin film 300 is adhered to the second surface 120. For example, the first thin film 200 and the second thin film 300 can be adhered to the two surfaces of the chip body 100 using single-sided adhesive.

[0131] The chip body 100 can be made of polymer materials, such as PMMA (Poly methyl methacrylate), PC (Polycarbonate), COP (Cyclo olefin polymer), COC (copolymers of cycloolefin), PP (Polypropylene), etc.

[0132] The first film body 200 is a transparent polymer film, for example, a transparent polymer film with good biocompatibility and containing a single-sided adhesive. The second film body 300 is a polymer film or a metal film, for example, a polymer film or a metal film with good biocompatibility and good ductility containing a single-sided adhesive.

[0133] In one specific embodiment of this application, the phase change material separator is made of one or both of paraffin wax and organic gel. The organic gel may include at least one of a first organic gel and a second organic gel. The first organic gel is a pressed oil organic gel containing 0.1%-10%wt dodecyl stearic acid; the second organic gel is a pressed oil organic gel containing 0.1%-10%wt octacosanol.

[0134] Paraffin wax has a melting point between 30-85℃. The organic gel can be a coconut oil organic gel containing 0.1%-10%wt dodecyl stearic acid or a coconut oil organic gel containing 0.1%-10%wt octacosanol. Coconut oil can also be pressed oils such as peanut oil, corn oil, and olive oil. At the same time, the organic gel can also be mixed with paraffin wax, such as a mixture of pressed oil organic gel containing 0.1%-10%wt octacosanol and paraffin wax containing 20%-40%wt with a melting point between 35-45℃. It should be noted that the material of the phase change material separator can be any material that is insoluble in water with a melting point between 30-85℃, and is not limited to the types disclosed in the above examples.

[0135] like Figures 3-12 As shown, in a specific embodiment of this application, the first connecting portion may include a first connecting pipe 1114 and a first recess 1116. It should be noted that the specific number of the first connecting pipe 1114 and the first recess 1116 in the first connecting portion should be determined according to the actual situation, or needs to be determined according to the specific number of the first reaction chambers 1112.

[0136] The first inlet 1111 of the first microfluidic conduit 111 and a first reaction chamber 1112 adjacent to the first inlet 1111 are connected by one of the first connecting conduits 1114. The first outlet 1113 of the first microfluidic conduit 111 and a first reaction chamber 1112 adjacent to the first outlet 1113 are connected by one of the first connecting conduits 1114.

[0137] Any two adjacent first reaction chambers 1112 are connected by a first recess 1116 between them. That is, in this embodiment, at least one first recess 1116 is provided in the flow path between any two adjacent first reaction chambers 1112. The opening side of the first recess 1116 is connected to at least one side surface of the chip body 100. Therefore, the thin film can be pressed into the first recess 1116 by the corresponding side surface of the chip body 100, and the first recess 1116 can be blocked by the thin film, thereby blocking the flow path between two adjacent first reaction chambers 1112 and completely separating each first reaction chamber 1112.

[0138] In one specific embodiment of this application, the second connecting portion includes a second connecting pipe 1214 and a second recess 1215. It should be noted that the specific number of the second connecting pipe 1214 and the second recess 1215 in the second connecting portion should be determined according to the actual situation, or needs to be determined according to the specific number of the second reaction chambers 1212.

[0139] The second inlet 1211 of the second microfluidic conduit 121 and a second reaction chamber 1212 adjacent to the second inlet 1211 are connected by one of the second connecting conduits 1214. The second outlet 1213 of the second microfluidic conduit 121 and a second reaction chamber 1212 adjacent to the second outlet 1213 are connected by one of the second connecting conduits 1214.

[0140] Any two adjacent second reaction chambers 1212 are connected by a second recess 1215 between them. That is, in this embodiment, at least one second recess 1215 is provided in the flow path between any two adjacent second reaction chambers 1212. The opening side of the second recess 1215 is connected to at least one side surface of the chip body 100. Therefore, the thin film can be pressed into the second recess 1215 by the corresponding side surface of the chip body 100, and the second recess 1215 can be blocked by the thin film, thereby blocking the flow path between two adjacent second reaction chambers 1212 and completely separating each second reaction chamber 1212.

[0141] Furthermore, such as Figures 5-7 As shown, the opening side of the first recess 1116 and the opening side of the second recess 1215 can be connected to the same side surface of the chip body 100. This arrangement allows the film to be pressed simultaneously into the first recess 1116 and the second recess 1215 on one side surface of the chip body 100, simplifying the structure of the corresponding nucleic acid detection cartridge. It should be noted that since the first recess 1116 and the second recess 1215 are arranged on the same side surface of the chip body 100, they should be spaced apart to prevent them from communicating with each other.

[0142] like Figures 10-12 As shown, in this embodiment, the opening side of the first recess 1116 and the opening side of the second recess 1215 can also be connected to different side surfaces of the chip body 100, respectively. With this configuration, the film can be pressed into the first recess 1116 and the second recess 1215 simultaneously on both sides of the chip body 100. A structure with pressing recesses (first recess 1116 and second recess 1215) needs to be provided both above and below the nucleic acid detection card cartridge, so that the first recess 1116 and the second recess 1215 can be blocked on the upper and lower sides of the chip body 100, respectively.

[0143] When the opening side of the first recess 1116 and the opening side of the second recess 1215 are connected to the same side surface of the chip body 100, the opening side of the first recess 1116 and the opening side of the second recess 1215 are both connected to the side surface of the chip body 100 adjacent to the second reaction chamber 1212. During nucleic acid detection, the side surface of the chip body 100 adjacent to the second reaction chamber 1212 can be arranged on the lower side.

[0144] The opening sides of the first recess 1116, the second recess 1215, and the second reaction chamber 1212 are connected to the same side surface of the chip body 100. For example, the first reaction chamber 1112 may be connected to the first surface 110, and the first recess 1116, the second recess 1215, and the second reaction chamber 1212 may be connected to the second surface 120. During nucleic acid detection, the second surface is positioned on the lower side, which facilitates the compression and sealing of the first recess 1116 and the second recess 1215 from below.

[0145] When the opening side of the first recess 1116 and the opening side of the second recess 1215 are connected to the same side surface of the chip body 100, the first recess 1116 can be connected to the corresponding first reaction chamber 1112 through the connecting hole 1115 and the first connecting pipe 1114.

[0146] Since the first recess 1116 is located on the same side as the second recess 1215 and the second reaction chamber 1212, there is a certain height difference between the first recess 1116 and the first reaction chamber 1112 on the other side. Furthermore, since the first recess 1116 requires external sealing, its depth cannot be too deep, otherwise the sealing effect will be affected. This necessitates that the first recess 1116 be connected to the first reaction chamber 1112 using some other structure.

[0147] In this embodiment, as Figure 8 and Figure 9 As shown, a connecting hole 1115 is provided on the outer side of the first recess 1116. One end of the connecting hole 1115 is connected to the first recess 1116, and the other end extends towards the first surface 110 and is connected to the corresponding first reaction chamber 1112 through a corresponding first connecting pipe 1114. The connecting hole 1115 can penetrate the wall thickness of the chip body 100. The connecting hole 1115 can be arranged adjacent to the first recess 1116 and at a certain distance from the first reaction chamber 1112. The connection between the connecting hole 1115 and the first reaction chamber 1112 can be achieved through the first connecting pipe 1114. The connecting holes 1115 can be symmetrically arranged on both sides of the first recess 1116 so that the two connecting holes 1115 can be connected to the two first reaction chambers 1112 on both sides of the first recess 1116 respectively, thereby achieving the connection between the two first reaction chambers 1112.

[0148] Figures 10-12In the illustrated scheme, since the first recess 1116 and the second recess 1215 are located on the same side of their respective first reaction chambers 1112 and 1212, the connecting hole 1115 disclosed in the above embodiment can be eliminated. The first recess 1116 can be directly connected to the corresponding first reaction chamber 1112 through the corresponding first connecting pipe 1114, and the second recess 1215 can be directly connected to the corresponding second reaction chamber 1212 through the corresponding second connecting pipe 1214.

[0149] like Figures 13-15 As shown, the first connecting part may also include only the first connecting pipe 1114. Any two adjacent ones among the first inlet 1111 of the first microfluidic pipe 111, each of the first reaction chambers 1112 and the first outlet 1113 of the first microfluidic pipe 111 are connected through the corresponding first connecting pipe 1114, and each of the first connecting pipes 1114 is connected to the first surface 110.

[0150] Correspondingly, the second connecting part may only include the second connecting pipe 1214. Any two adjacent ones among the second inlet 1211 of the second microfluidic pipe 121, each of the second reaction chambers 1212 and the second outlet 1213 of the second microfluidic pipe 121 are connected through the corresponding second connecting pipe 1214. Each second connecting pipe 1214 is connected to the second surface 120 of the chip body 100.

[0151] The chip body 100 disclosed in this embodiment differs from those in the above embodiments in that it does not require the first recess 1116 and the second recess 1215. The material of the chip body 100 disclosed in this embodiment can be the same as that of the chip body 100 disclosed in the above embodiments, and will not be described again here; the corresponding materials of the first thin film 200 and the second thin film 300 can also be the same, and will not be described again here.

[0152] like Figures 16-18 As shown, the chip body 100 is bonded to the first thin film 200 and the second thin film 300 by thermocompression bonding. The blocking between the reaction chambers can also be achieved by thermocompression bonding, i.e., according to... Figure 18 At the locations marked by the black dashed and solid lines, a heated cutting tip presses against the microfluidic chip, causing the thin film above the first and second connecting channels 1114 and 1214 to thermally bond to their substrates, thus achieving physical isolation between the different reaction chambers. Finally, the phase change material separator is destroyed through ultrasound, centrifugation, and heating, connecting the first reaction chamber 1112 and the second reaction chamber 1212, enabling specific mixing of different test samples with different primers and probes. Ultimately, multi-sample, multi-target nucleic acid detection is achieved on a single microfluidic chip.

[0153] like Figures 22-27 As shown in the embodiments of this application, a nucleic acid detection cartridge 400 is also disclosed. The nucleic acid detection cartridge 400 can be used in conjunction with the microfluidic chip disclosed in some of the above embodiments for detection, for example, it can be used to detect the chip body 100 with a first pit 1116 and a second pit 1215.

[0154] The nucleic acid detection cartridge 400 includes a heating device, a pressing mechanism, and a pressing micropillar 422. The heating device heats the microfluidic chip to meet the temperature requirements of LAMP or PCR amplification processes. It should be noted that when used in PCR nucleic acid amplification reactions, a corresponding cooling device is also required to lower the temperature of the microfluidic chip. This cooling device can be an air-cooled device or a liquid-cooled device; this embodiment does not limit the specific cooling method.

[0155] The pressing mechanism is used to press the microfluidic chip firmly onto the heating device. The pressing mechanism applies pressure to the microfluidic chip, making the microfluidic chip adhere tightly to the heating device, thereby ensuring that the heat from the heating device can be conducted to the microfluidic chip, increasing the temperature of the microfluidic chip, and ensuring that the temperature environment of the reaction chamber of the microfluidic chip meets the temperature requirements of the nucleic acid amplification reaction.

[0156] The pressing micropillar 422 is used to pass through the heating device and corresponds one-to-one with each of the first recesses 1116 and each of the second recesses 1215. The pressing micropillar 422 can press the film body into the first recesses 1116 and the second recesses 1215, thereby blocking each reaction chamber by sealing the first recesses 1116 and the second recesses 1215.

[0157] When the first recess 1116 and each of the second recesses 1215 are located on the same side of the chip body 100, the pressing micropillars 422 only need to be arranged on one side of the chip body 100, for example, on the lower part of the chip body 100. When the first recess 1116 and each of the second recesses 1215 are located on both sides of the chip body 100, the pressing micropillars 422 need to be arranged on both the upper and lower sides of the chip body 100 to seal the first recess 1116 and each of the second recesses 1215 respectively.

[0158] Because the nucleic acid test cartridge is equipped with a heating device, the microfluidic chip can complete the amplification process within the nucleic acid test cartridge 400. The pressing micropillars 422 can isolate the various reaction chambers, preventing cross-contamination. The pressure applied by the pressing mechanism not only ensures the adhesion between the heating device and the microfluidic chip, guaranteeing heat transfer from the heating device to the microfluidic chip, but also allows the pressing micropillars 422 to press the film into the first recess 1116 and each of the second recesses 1215, thus sealing the first recess 1116 and each of the second recesses 1215.

[0159] When testing needs to be completed within the nucleic acid detection cartridge 400, the nucleic acid detection cartridge 400 may further include an imaging device 411, the imaging range of which can cover the entire microfluidic chip. The imaging device 411 is used to acquire fluorescence signals or color signals that change over time, thereby determining the nucleic acid amplification results in each reaction chamber. Those skilled in the art will understand that the nucleic acid detection cartridge 400 should also include necessary modules such as a power supply module and a microcontroller control module, which will not be elaborated upon here.

[0160] In one specific embodiment of this application, the nucleic acid test kit 400 may further include a base 420, a support 440, and a top cover 410. Each of the pressing micropillars 422 is disposed on the base 420. To facilitate the installation of each pressing micropillar 422, the pressing micropillars 422 can be integrated onto a base plate, and the base plate and its pressing micropillars 422 can be installed together onto the base 420.

[0161] To facilitate the positioning of the heating device, this embodiment includes a first positioning post 421 on the base 420. An elastic reset member 4211 is fitted onto the first positioning post 421. The heating device has first positioning holes 4241 that slide in conjunction with each of the first positioning posts 421. The number of first positioning posts 421 can be designed according to requirements. Figure 26 Taking the illustrated scheme as an example, four first positioning posts 421 are set at the four corners of the heating device to ensure the stability of the heating device when it moves up and down.

[0162] One end of the elastic reset member 4211 abuts against the base 420, and the other end abuts against the heating device. During testing, the microfluidic chip is placed above the heating device. When the pressing mechanism presses the microfluidic chip onto the heating device, the microfluidic chip and the heating device compress the elastic reset member 4211 downwards, causing the elastic reset member 4211 to store energy and generate an upward supporting force. This clamps the microfluidic chip and the heating device between the pressing mechanism and the elastic reset member 4211, ensuring a flexible and tight fit between them. Furthermore, when the downward pressing force of the pressing mechanism acts on the microfluidic chip and the heating device, the microfluidic chip and the heating device can compress the elastic reset member 4211 downwards for cushioning, preventing the microfluidic chip and the heating device from being damaged by the pressing mechanism.

[0163] In this embodiment, the heating device may include a heating film 423 and a heat transfer plate 424, with the heating film 423 adhered to the heat transfer plate 424. Specifically, the heating film 423 can be attached to the lower surface of the heat transfer plate 424 using double-sided adhesive tape, and the heat transfer plate 424 is positioned closer to the microfluidic chip than the heating film 423. During use, the microfluidic chip is in contact with the heat transfer plate 424, and the heat from the heating film 423 is conducted to the microfluidic chip through the heat transfer plate 424.

[0164] It should be noted that the area of ​​the heating film 423 can be smaller than the area of ​​the heat transfer plate 424. Since four first positioning posts 421 are provided at the four corners of the heating device, the heating film 423 can be located within the quadrilateral formed by the four first positioning posts 421. The heating film 423 does not need to have positioning holes that mate with the first positioning posts 421. It is only necessary to have first positioning holes that slide and mate with the first positioning posts 421 at the corresponding positions on the heat transfer plate 424.

[0165] Furthermore, since the pressing micropillar 422 needs to pass through the heating device to cooperate with the microfluidic chip, holes need to be made on both the heating film 423 and the heat transfer plate 424 to cooperate with the pressing micropillar 422. For example, heating film through holes 4232 need to be made on the heating film 423, and heat transfer plate through holes 4243 need to be made on the heat transfer plate 424. When attaching the heating film 423 to the heat transfer plate 424, it should be ensured that each heating film through hole 4232 and each heat transfer plate through hole 4243 are connected in a one-to-one correspondence.

[0166] The support member 440 is disposed on the base 420, and one end of the top cover 410 is hinged to the support member 440. The base 420, the support member 440 and the top cover 410 together form the outer shell of the nucleic acid test card box 400, and when the top cover 410 is closed, it can surround the microfluidic chip and heating device and other components in the cavity of the outer shell.

[0167] The pressing mechanism is located on the upper cover 410. When the upper cover 410 rotates along its hinge axis, it drives the pressing mechanism to move, thereby changing the position of the pressing mechanism. When the upper cover 410 is in the closed state, the pressing mechanism is in the pressed position, that is, the pressing mechanism presses the microfluidic chip onto the heating device. When the upper cover 410 is in the open state, the pressing mechanism is in the released position, that is, the pressing mechanism disengages from the microfluidic chip, facilitating the placement and removal of the microfluidic chip.

[0168] In one specific embodiment of this application, the pressing mechanism can be a linkage structure, specifically including a first linkage 432, a second linkage 433, a third linkage 435, a support rod 436, and a fourth linkage 434.

[0169] The first end of the first connecting rod 432 is hinged to the upper cover 410. Specifically, a mounting part 431 can be provided on the inner side of the upper cover 410, and the first end of the first connecting rod 432 is hinged to the mounting part 431.

[0170] The first ends of the second link 433 and the third link 435 are both hinged to the second end of the first link 432. This means that the first ends of the second link 433, the third link 435, and the first link 432 can be hinged to the same hinge axis.

[0171] Support rod 436 is fixed to support member 440, and a sliding sleeve 4361 is provided on support rod 436. Fourth link 434 is slidably engaged with sliding sleeve 4361. One end of fourth link 434 is hinged to the second end of third link 435, and the other end is used to press the microfluidic chip. The second end of second link 433 is hinged to support rod 436.

[0172] It should be noted that the first and second ends of the aforementioned connecting rods, as well as one and the other ends of the support rod 436, do not refer only to the ends. In the absence of interference, the hinge point can be set at any feasible position other than the end face.

[0173] like Figure 27 As shown, when the top cover 410 is lifted, that is, when the top cover 410 moves counterclockwise around the hinge 412, it pulls the first link 432 to move upward with the top cover 410. At the same time, the first link 432 swings along the hinge end with the top cover 410. The first link 432 pulls the second link 433 and the third link 435 to follow the movement. At this time, the third link 435 swings around the hinge end with the support rod 436. Since the fourth link 434 is slidably engaged with the sliding sleeve 4361, the fourth link 434 only has the freedom of up and down movement. It is pulled upward by the second link 433 and the third link 435, thereby causing the fourth link 434 to lift up and release the pressure on the microfluidic chip.

[0174] When the top cover 410 is lowered, it moves clockwise around the hinge 412, pushing the first link 432 downwards with the top cover 410. Simultaneously, the first link 432 swings along its hinged end with the top cover 410. The first link 432 pushes the second link 433 and the third link 435 to follow suit, at which point the third link 435 swings around its hinged end with the support rod 436. Because the fourth link 434 is slidably engaged with the sliding sleeve 4361, the fourth link 434 only has the freedom to move up and down. It is pushed downwards by the second link 433 and the third link 435, causing the fourth link 434 to press down, applying pressure to the microfluidic chip.

[0175] The imaging device 411 can be installed on the inner wall of the upper cover 410. When the upper cover 410 is lowered, the imaging range of the imaging device 411 can cover the entire microfluidic chip.

[0176] like Figure 28As shown, to facilitate the pressing mechanism applying pressure to the microfluidic chip, in this embodiment, chip pressing portions 150 are provided at both ends of the chip body 100, and there are two pressing mechanisms, each used to press against the chip pressing portion 150. In this embodiment, by providing chip pressing portions 150 at both ends of the chip body 100 and cooperating with the pressing mechanism with the chip pressing portion 150, it is possible to prevent the chip pressing portion 150 from applying pressure to the microfluidic channel, thus avoiding affecting the performance of the microfluidic chip.

[0177] like Figure 27 As shown, a pressing seat 4341 can be provided at the lower end of the fourth link 434. The cross-sectional area of ​​the pressing seat 4341 is larger than that of the fourth link 434, so that the contact area with the microfluidic chip is larger, reducing the pressure acting on the microfluidic chip and preventing damage to the microfluidic chip.

[0178] like Figure 26 As shown, to facilitate the positioning of the microfluidic chip, a second positioning post 4212 is provided on the base 420. The second positioning post 4212 is mainly used to position the microfluidic chip. Similarly, the pressing micro-posts 422 also need to have a corresponding positional relationship with the microfluidic chip. Therefore, to improve installation accuracy, all pressing micro-posts 422 and all second positioning posts 4212 can be integrated onto a single base plate, easily ensuring the positional relationship between the second positioning posts 4212 and the pressing micro-posts 422. During installation, simply install the base plate onto the base 420; the second positioning posts 4212 and the pressing micro-posts 422 will then have a defined positional relationship. Only the positional relationship between the base plate and the first positioning post 421 needs to be adjusted. It should be noted that the base 420, as well as the first positioning post 421, second positioning post 4212, and pressing micro-post 422 on the base 420, can also be made into a single integrated structure.

[0179] The heating device has a heating device positioning hole that slides with the second positioning post 4212. The heating device positioning hole includes a first heating device positioning hole 4231 on the heating film 423 and a second heating device positioning hole 4242 on the heat transfer plate 424.

[0180] The microfluidic chip has a chip positioning hole 130 that slides with the second positioning post 4212. The second positioning post 4212 can ensure the positional relationship between the microfluidic chip and the heating device, and at the same time determine the positional relationship between the first recess 1116 and the second recess 1215 of the microfluidic chip and the pressing micropost 422.

[0181] In summary, the detection process using the nucleic acid test kit 400 is as follows:

[0182] After the sample to be tested and the nucleic acid amplification system are added to the microfluidic chip, the inlet and outlet of the first microfluidic channel 111 and the second microfluidic channel 121 are sealed with single-sided adhesive tape.

[0183] The microfluidic chip is placed in the nucleic acid detection card cartridge 400, wherein the chip positioning hole 130 on the microfluidic chip cooperates with the second positioning post 4212, so that the microfluidic chip is accurately fixed on the upper surface of the heat transfer plate 424;

[0184] Then, the top cover 410 is lowered, and the microfluidic chip is pressed against the upper surface of the heat transfer plate 424 by the fourth link 434 of the top cover 410, making close contact with it. At this time, the microfluidic chip and the heating device are pressed and move downward together. The pressing micro-pillars 422 on the base 420 make close contact with the thin film on the lower surface of the microfluidic chip through the heat transfer plate 424, and make it contact the first recess 1116 and the second recess 1215 provided on the lower surface of the microfluidic chip, thus completing the isolation of each reaction chamber.

[0185] Finally, under the control of the microcontroller, the heating film 423 generates heat, the microfluidic chip is heated, and the phase change material insulator at the chamber connection hole 140 melts, thereby connecting the first reaction chamber 1112 and the second reaction chamber 1212 on both sides of the chip body 100. The sample to be tested is mixed with the nucleic acid amplification system, and the sample in each reaction chamber is mixed with only one nucleic acid amplification system. Different reaction chambers of the same sample are mixed with different primers and probes, thus realizing portable and rapid multi-sample multi-index nucleic acid detection. The imaging device 411 located on the top cover 410 can collect fluorescence signals or color signals that change over time to determine the nucleic acid amplification results of each reaction chamber.

[0186] like Figures 29-32 As shown in the embodiments, this application also discloses a nucleic acid extraction cartridge 500, which is used in conjunction with the microfluidic chip disclosed in the above embodiments, so that the microfluidic chip and the nucleic acid extraction cartridge 500 are combined to form a fully integrated nucleic acid analysis scheme design of "sample in - result out".

[0187] The nucleic acid extraction cartridge 500 includes a nucleic acid extraction cartridge body 510, a sample piston pusher 520, a fluid piston pusher 530, a soft stopper 540, and a hollow needle 550. The nucleic acid extraction cartridge body 510 has a lysis chamber 511, a washing chamber 516, and an elution chamber 518 arranged sequentially from top to bottom. A vent 513 is provided at the upper part of the lysis chamber 511, allowing the lysis chamber 511 to communicate with the outside environment.

[0188] The nucleic acid extraction cartridge body 510 also includes a sample chamber 512 at the same height as the lysis chamber 511, and the sample chamber 512 and lysis chamber 511 are connected through a sample injection hole 514 at the bottom. Those skilled in the art will understand that both the sample chamber 512 and the lysis chamber 511 contain a certain amount of lysis buffer. The sample chamber 512 is used to hold the original biological sample. The lysis chamber 511 uses its lysis buffer to lyse the cells in the sample. The lysis chamber 511 uses a lysis buffer (such as one containing detergents, proteases, etc.) to disrupt the cell membranes, nuclear membranes, and other structures in the sample, releasing intracellular nucleic acids (DNA or RNA) into the system, preparing for subsequent extraction steps.

[0189] When nucleic acids are released from cells, they contain many impurities, such as proteins, polysaccharides, lipids, and reagent residues from the lysis process. The purpose of the washing chamber 516 is to remove these impurities. A suitable washing solution is added to the washing chamber 516, and through multiple washing operations, the nucleic acids are separated from the impurities, thus improving the purity of the nucleic acids.

[0190] After the preceding steps, the nucleic acid has been preliminarily separated and purified, but it still needs to be eluted from the binding carrier (if a nucleic acid binding carrier such as a silica membrane or magnetic beads was used during extraction) to form a nucleic acid solution that can be used for subsequent experiments. Elution chamber 518 is the part that performs this function. By adding a specific elution buffer, the interaction between the nucleic acid and the binding carrier is disrupted, allowing the nucleic acid to dissolve in the elution solution, ultimately obtaining a solution containing high-purity nucleic acid, thus yielding the sample to be tested.

[0191] The sample piston pusher 520 engages with the sample chamber 512, and the fluid piston pusher 530 engages with the lysis chamber 511. By controlling the sliding of the sample piston pusher 520 within the sample chamber 512, the pressure in the sample chamber 512 can be changed, thereby driving the flow of the sample within the sample chamber 512. Similarly, by controlling the sliding of the fluid piston pusher 530 within the lysis chamber 511, the pressure in the lysis chamber 511 can be changed, thereby driving the flow of the solution within the lysis chamber 511.

[0192] A soft stopper 540 is sealed within the elution chamber 518. One end of a hollow needle 550 is inserted into the soft stopper 540 from its lower side (the soft stopper 540 can be a rubber stopper or any other soft material, as long as it can be pierced by the hollow needle 550). The upper end of the hollow needle 550 is located below the upper surface of the soft stopper 540, meaning the hollow needle 550 does not penetrate the soft stopper 540, and thus the upper end of the hollow needle 550 is not connected to the upper cavity of the elution chamber 518. The lower end of the hollow needle 550 is used to connect to the inlet of one of the first microfluidic channel 111 and the second microfluidic channel 121, for injecting the nucleic acid extracted sample solution into one of the first microfluidic channel 111 and the second microfluidic channel 121 through the hollow needle 550. The other of the first microfluidic channel 111 and the second microfluidic channel 121 is then used to inject the nucleic acid amplification system.

[0193] To avoid aerosol contamination, a hydrophobic and breathable membrane can be used to seal the vent 513. In the initial state, when the sample piston push rod 520 is not engaged with the sample cavity 512, the top opening of the sample cavity 512 is sealed by a sealing layer, for example, by sealing the top opening of the sample cavity 512 with aluminum foil.

[0194] Furthermore, the pyrolysis chamber 511 and the washing chamber 516 are connected through a first through-hole 515, and the washing chamber 516 and the elution chamber 518 are connected through a second through-hole 517. The diameter of the first through-hole 515 is smaller than the diameters of the pyrolysis chamber 511 and the washing chamber 516, and the diameter of the second through-hole 517 is smaller than the diameters of the washing chamber 516 and the elution chamber 518. The first through-hole 515 and the second through-hole 517 are filled with organic reagents that are incompatible with the aqueous phase reagents, so that the solutions in the pyrolysis chamber 511, the washing chamber 516, and the elution chamber 518 can exist stably. The organic reagents can be mineral oil, silicone oil, phase change paraffin, or organic gels, etc.

[0195] In summary, since the sample chamber 512 and the lysis chamber 511 are connected through the sample injection hole 514, both the sample chamber 512 and the lysis chamber 511 contain a certain amount of lysis solution under the principle of communicating vessels. The washing chamber 516 is filled with washing solution, and the elution chamber 518 is filled with elution solution. The first through hole 515 and the second through hole 517, which serve as connecting holes between the upstream and downstream chambers, are filled with organic reagents that are incompatible with the aqueous phase reagents, such as mineral oil, silicone oil, phase change paraffin, and organic gels, so that the reagents in the lysis chamber 511, washing chamber 516, and elution chamber 518 can exist stably. In the initial state, the vent hole 513 is covered by a hydrophobic and breathable membrane, the sample piston push rod 520 has not yet been added to the sample chamber 512, and the top opening of the sample chamber 512 is sealed with aluminum foil.

[0196] It should be noted that a sample piston pusher 520, fluid piston pusher 530, soft stopper 540, and hollow needle 550 form a group to serve nucleic acid extraction for one sample. Those skilled in the art can set up multiple groups as needed to extract nucleic acids from multiple samples simultaneously. The same number of groups can also be designed based on the number of sample inlets on the microfluidic chip (e.g., the inlet of the first microfluidic channel 111); of course, the number can be more or less than the number of sample inlets on the microfluidic chip.

[0197] Furthermore, a sample chamber 512, a lysis chamber 511, a washing chamber 516, and an elution chamber 518 are used in conjunction with each set of sample piston push rods 520, fluid piston push rods 530, soft plugs 540, and hollow needles 550. When the number of sets of sample piston push rods 520, fluid piston push rods 530, soft plugs 540, and hollow needles 550 is n, then correspondingly n sets of sample chambers 512, lysis chambers 511, washing chambers 516, and elution chambers 518 need to be configured.

[0198] like Figure 33 As shown, for the sake of simplicity, the images will be... Figure 33 All reference markers in the attached drawings have been removed. For more information on reference markers, please refer to [link / reference]. Figure 32 .

[0199] like Figure 33 As shown in Figure a, after the sample and magnetic beads are added to the sample cavity 512, due to the principle of communicating vessels, the surfaces of both the sample cavity 512 and the lysis cavity 511 will rise. At the same time, since the compression of the liquid is negligible and the oil-water interface formed at the first through hole 515 provides stable separation, the newly added sample aqueous solution will not affect the solutions in the washing cavity 516 and the elution cavity 518 below.

[0200] like Figure 33 As shown in b, the sample piston pusher 520 is added to the sample chamber 512. Due to the compression of the air in the sample chamber 512 by the sample piston pusher 520, some of the liquid in the sample chamber 512 is forced into the lysis chamber 511. The sample piston pusher 520 can be repeatedly pulled in and out, causing the lysis fluid to mix with the newly added sample solution and magnetic beads. Excess gas will enter and exit through the vent 513.

[0201] like Figure 33 As shown in Figure c, magnetic beads are enriched on the outside of the nucleic acid extraction cartridge body 510 using a magnet 600. The specific process is as follows: First, the sample piston push rod 520 is pressed all the way down, so that all the liquid enters the lysis chamber 511, and excess gas is discharged from the vent 513. At this time, a magnet 600 is used to adhere tightly to the nucleic acid extraction cartridge body 510, and the magnetic beads in the lysis chamber 511 that have adsorbed nucleic acid will adhere tightly to the magnet area.

[0202] like Figure 33As shown in d, a magnet 600 is used to transfer magnetic beads adsorbed with nucleic acids from the lysis chamber 511 to the washing chamber 516 for cleaning. Since the first through-hole 515 connecting the lysis chamber 511 and the washing chamber 516 is filled with organic reagents incompatible with the aqueous phase, such as mineral oil, silicone oil, phase change paraffin, and organic gel, the transfer of the magnetic beads will not disrupt the oil-water interface. Therefore, it can be ensured that the reagents in the upper lysis chamber 511 will not enter the washing chamber 516. After the magnetic beads adsorbed with nucleic acids enter the washing chamber 516, they will be cleaned by the washing solution therein.

[0203] like Figure 33 As shown in section e, a magnet 600 is used to transfer magnetic beads adsorbed with nucleic acids from the washing chamber 516 to the elution chamber 518. The nucleic acids adsorbed on the magnetic beads are eluted by the elution buffer in the elution chamber 518. The elution buffer here can be a PCR or LAMP reaction system, etc.

[0204] like Figure 33 As shown in f, the reaction system (elution buffer) mixed with nucleic acid is injected into the microfluidic chip. Under the action of the fluid piston pusher 530, the fluid in the nucleic acid extraction cartridge body 510 moves downward. Due to the poor compressibility of the fluid, the soft stopper 540 eventually moves downward, causing the hollow needle 550 to completely penetrate the soft stopper 540. The reaction system (elution buffer) mixed with nucleic acid enters the flow channel of the microfluidic chip through the hollow needle 550. The subsequent operation of the microfluidic chip is similar to the operation process described above. Under heating conditions, the first reaction chamber 1112 and the second reaction chamber 1212 on the upper and lower surfaces of the microfluidic chip are connected, allowing the primer probe to mix with the nucleic acid amplification system, realizing fully integrated nucleic acid monitoring of multiple samples and multiple indicators.

[0205] As indicated in this application and claims, unless the context clearly indicates otherwise, the words "a," "an," "a," and / or "the" are not specifically singular and may include the plural. Generally, the terms "comprising" and "including" only indicate the inclusion of expressly identified steps and elements, which do not constitute an exclusive list, and the method or apparatus may also include other steps or elements. An element defined by the phrase "comprising an..." does not exclude the presence of other identical elements in the process, method, product, or apparatus that includes the element.

[0206] In the description of this application, unless otherwise expressly defined, terms such as "setup," "installation," and "connection" should be interpreted broadly, and those skilled in the art can reasonably determine the specific meaning of the above terms in this application in conjunction with the specific content of the technical solution.

[0207] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on the differences from other embodiments. The same or similar parts between the various embodiments can be referred to each other.

[0208] This document uses specific examples to illustrate the principles and implementation methods of this application. The descriptions of the embodiments above are only for the purpose of helping to understand the method and core ideas of this application. It should be noted that those skilled in the art can make several improvements and modifications to this application without departing from the principles of this application, and these improvements and modifications also fall within the protection scope of the claims of this application.

Claims

1. A microfluidic chip, characterized in that, The chip body includes a chip body (100) and thin films respectively disposed on a first surface (110) and a second surface (120) of the chip body (100), at least one of the thin films being a transparent film. The chip body (100) is provided with n first microfluidic channels (111) extending along a first direction and m second microfluidic channels (121) extending along a second direction, the first direction and the second direction intersecting. The first microfluidic channel (111) includes m first reaction chambers (1112) connected in sequence through a first connecting portion, and the second microfluidic channel (121) includes n second reaction chambers (1212) connected in sequence through a second connecting portion. The first connecting portion and the second connecting portion are connected to at least one side surface of the chip body (100). Along the thickness direction of the chip body (100), the first reaction chamber (1112) and the second reaction chamber (1212) are connected in a one-to-one correspondence, and a phase change material isolator is provided at the connection point. m and n are both natural numbers not less than 2. The thin film can be squeezed into the first connection part and the second connection part to completely separate each of the first reaction chambers (1112) and each of the second reaction chambers (1212). One of the first microfluidic conduit (111) and the second microfluidic conduit (121) is used to inject the nucleic acid amplification system, and the other is used to inject the sample to be tested; The opening side of the first reaction chamber (1112) is located on the first surface (110), and the opening side of the second reaction chamber (1212) is located on the second surface (120). The thin film disposed on the first surface (110) is the first thin film (200), and the thin film disposed on the second surface (120) is the second thin film (300). The first film body (200) is adhered to the first surface (110), and the second film body (300) is adhered to the second surface (120).

2. The microfluidic chip as described in claim 1, characterized in that, The phase change material insulator is made of one or both of paraffin and organic gel.

3. The microfluidic chip as described in claim 2, characterized in that, The organic gel includes at least one of a first organic gel and a second organic gel; The first organic gel is a pressed oil organic gel containing 0.1%-10%wt dodecyl stearic acid; The second organic gel is a pressed oil organic gel containing 0.1%-10%wt octacosanol.

4. The microfluidic chip according to any one of claims 1-3, characterized in that, The first connecting portion includes a first connecting pipe (1114) and a first recess (1116). The first inlet (1111) and the first outlet (1113) of the first microfluidic pipe (111) are connected to the adjacent first reaction chamber (1112) through the corresponding first connecting pipe (1114). Any two adjacent first reaction chambers (1112) are connected through the first recess (1116) between them. The opening side of the first recess (1116) is connected to at least one side surface of the chip body (100). The second connecting portion includes a second connecting pipe (1214) and a second recess (1215). The second inlet (1211) and the second outlet (1213) of the second microfluidic pipe (121) are connected to the adjacent second reaction chamber (1212) through the corresponding second connecting pipe (1214). Any two adjacent second reaction chambers (1212) are connected through the second recess (1215) between them. The opening side of the second recess (1215) is connected to at least one side surface of the chip body (100).

5. The microfluidic chip as described in claim 4, characterized in that, The opening side of the first recess (1116) and the opening side of the second recess (1215) are connected to the same side surface of the chip body (100); or, The opening side of the first recess (1116) and the opening side of the second recess (1215) are respectively connected to different side surfaces of the chip body (100).

6. The microfluidic chip according to any one of claims 1-3, characterized in that, The first connecting part includes a first connecting pipe (1114). Any two adjacent ones of the first inlet (1111) of the first microfluidic pipe (111), each of the first reaction chambers (1112) and the first outlet (1113) of the first microfluidic pipe (111) are connected through the corresponding first connecting pipe (1114). Each of the first connecting pipes (1114) is connected to the first surface (110). The second connecting portion includes a second connecting pipe (1214). Any two adjacent pairs of the second inlet (1211) of the second microfluidic pipe (121), each of the second reaction chambers (1212) and the second outlet (1213) of the second microfluidic pipe (121) are connected through the corresponding second connecting pipe (1214). Each of the second connecting pipes (1214) is connected to the second surface (120) of the chip body (100).

7. A nucleic acid testing kit, characterized in that, For use with the microfluidic chip as described in claim 4 or 5, comprising: A heating device is used to heat the microfluidic chip; A pressing mechanism is used to press the microfluidic chip onto the heating device; The micro-pillars (422) are pressed to pass through the heating device and correspond one-to-one with each of the first recesses (1116) and each of the second recesses (1215).

8. The nucleic acid detection cartridge as described in claim 7, characterized in that, Also includes: The base (420) has each of the pressing micro-pillars (422) disposed on the base (420). The base (420) has a first positioning post (421) disposed on the first positioning post (421) and an elastic reset member (4211) sleeved on the first positioning post (421). The heating device has a first positioning hole (4241) that slides with each of the first positioning posts (421). One end of the elastic reset member (4211) abuts against the base (420) and the other end abuts against the heating device. A support member (440) is disposed on the base (420); The top cover (410) is hinged at one end to the support member (440). The pressing mechanism is disposed on the top cover (410). When the top cover (410) is in the closed state, the pressing mechanism presses the microfluidic chip onto the heating device. When the top cover (410) is in the open state, the pressing mechanism disengages from the microfluidic chip.

9. The nucleic acid detection cartridge as described in claim 8, characterized in that, The pressing mechanism includes: The first link (432) is hinged at its first end to the upper cover (410); The second link (433) and the third link (435) are hinged to the second end of the first link (432); A support rod (436) is fixed to the support member (440), and a sliding sleeve is provided on the support rod (436); The fourth link (434) is slidably engaged with the sliding sleeve. One end of the fourth link (434) is hinged to the second end of the third link (435), and the other end is used to press the microfluidic chip. The second end of the second link (433) is hinged to the support rod (436).

10. The nucleic acid detection cartridge as described in claim 8, characterized in that, The base (420) is provided with a second positioning post (4212), and the heating device is provided with a heating device positioning hole that slides with the second positioning post (4212); The microfluidic chip has a chip positioning hole (130) that slides with the second positioning post (4212). And / or, The heating device includes a heating film (423) and a heat transfer plate (424). The heating film (423) is bonded to the heat transfer plate (424), and the heat transfer plate (424) is arranged closer to the microfluidic chip than the heating film (423). And / or, The nucleic acid detection cartridge also includes an imaging device (411), which is used to collect fluorescence signals or color signals that change over time.

11. A nucleic acid extraction cartridge, characterized in that, Including the microfluidic chip as described in any one of claims 1-6, further comprising: The nucleic acid extraction cartridge body (510) is provided with a lysis chamber (511), a washing chamber (516) and an elution chamber (518) connected sequentially from top to bottom. The nucleic acid extraction cartridge body (510) is also provided with a sample chamber (512) at the same height as the lysis chamber (511). The sample chamber (512) and the lysis chamber (511) are connected through a sample injection hole (514) at the bottom. A vent hole (513) is provided at the upper part of the lysis chamber (511). The sample piston push rod (520) and the fluid piston push rod (530) are provided, wherein the sample piston push rod (520) is used to cooperate with the sample chamber (512) and the fluid piston push rod (530) is used to cooperate with the lysis chamber (511); A soft plug (540) is sealed in the elution chamber (518); A hollow needle (550) is inserted into the soft plug (540) from the lower side, and the upper end of the hollow needle (550) is located below the upper surface of the soft plug (540). The lower end of the hollow needle (550) is used to communicate with the inlet of one of the first microfluidic channel (111) and the second microfluidic channel (121).

12. The nucleic acid extraction cartridge as described in claim 11, characterized in that, The vent (513) is sealed by a hydrophobic and breathable membrane; And / or, When the sample piston push rod (520) is not engaged with the sample cavity (512), the top opening of the sample cavity (512) is sealed by the sealing layer; And / or, The pyrolysis chamber (511) and the cleaning chamber (516) are connected through a first through hole (515), and the cleaning chamber (516) and the elution chamber (518) are connected through a second through hole (517). The diameter of the first through hole (515) is smaller than the diameter of the pyrolysis chamber (511) and the cleaning chamber (516), and the diameter of the second through hole (517) is smaller than the diameter of the cleaning chamber (516) and the elution chamber (518). The first through hole (515) and the second through hole (517) are filled with organic reagents that are incompatible with the aqueous phase reagents. The organic reagents are mineral oil, silicone oil, phase change paraffin, or organic gel.

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