Fluid cassette suitable for portable nucleic acid detection devices

Abstract

To provide a fluid cassette that is highly stable, simple, and can be operated easily and be used in portable nucleic acid detection devices.SOLUTION: A fluid cassette 2 comprises a cap 20, a main body 21, an inner tube 25, and a detection box 24. The main body has an outer tube 22 and a shield cover 23, and the outer tube is connected to the shield cover. The inner tube is disposed correspondingly within the outer tube of the main body. The detection box is correspondingly disposed under the shield cover and has an injection chamber, a plurality of channels and a plurality of reaction chambers, and the plurality of reaction chambers are communicated with the injection chamber via the plurality of channels. When a sample is placed in the inner tube, it mixes with a liquid in the inner tube to form a test liquid, which is sealed to the outer tube of the main body via the cap, and is punctured by pushing the inner tube downward. When the structure of the inner tube pierces a sealing film 27a of the detection box downward, the test liquid flows into the injection chamber, passes through the plurality of channels, flows into the plurality of reaction chambers, and multiplexed nucleic acid amplification is performed.SELECTED DRAWING: Figure 2B

Description

[Technical Field]

[0001] The present invention relates to a fluidic cassette, particularly a fluidic cassette suitable for a portable nucleic acid detection device for rapid multiplex nucleic acid amplification and detection. [Background technology]

[0002] To obtain large quantities of specific DNA fragments for various needs, scientists have strived to find efficient methods to achieve this goal. Polymerase chain reaction (PCR) is one of the most economical and rapid techniques, capable of generating billions of copies of a specific DNA fragment in a short period of time. PCR technology can be used in a variety of fields, including selective DNA isolation for gene identification, forensic analysis of ancient DNA in archaeology, medical applications such as genetic testing and tissue typing, rapid and accurate diagnosis of infectious diseases in hospitals and research institutions, environmental hazard testing for food safety, and genetic fingerprinting detection for criminal investigations. PCR uses a small amount of DNA sample extracted from blood or tissue. Simply by adding a fluorescent dye to the nucleic acid solution, the amplified DNA fragments can be detected via fluorescent molecules.

[0003] Infectious diseases are caused by pathogens, including bacteria, viruses, and parasites. Effective prevention of infectious disease outbreaks requires rapid, low-cost, and accurate on-site diagnostics. Therefore, there is an urgent need to develop technologies that can perform low-cost, rapid point-of-care testing (POCT) of infectious pathogens outside of hospitals and laboratories.

[0004] Current state-of-the-art rapid diagnostic kits do not offer low-cost multiplex testing. For example, Cue Health, Lucira, and HealthPod, all currently available, are single-disease diagnostics, offering only single-target tests with varying sensitivity; they cannot offer multiplex testing. Other companies that offer multiplex testing use more expensive technology and complex fluid control systems, resulting in higher machine and test costs. The price and demand for such complex mechanical valve systems make them out of reach of the general public. For example, if the general public wants to perform a multiplex PCR test, they must send their samples to a large laboratory and have the multiplex PCR performed via a complex mechanical valve system. This not only requires expensive testing equipment and complex testing procedures, but also requires significant labor and time costs.

[0005] Furthermore, conventional PCR microfluidic detection procedures present many challenges. For example, the presence of air bubbles in the reaction chamber can affect detection results and lead to erroneous results. Therefore, a separate exhaust system is required to expel air bubbles, but installing an exhaust system to remove air bubbles can cause the detection solution to evaporate, potentially affecting detection accuracy. Furthermore, during the detection process, it is necessary to consider whether there is a risk of biomolecules leaking from the reaction chamber or contaminating the surrounding environment through aerosols or evaporation of water vapor. Furthermore, when applying PCR microfluidic detection programs to portable devices, can the problems of vibration and tilt caused by detection in unstable environments be overcome to obtain accurate detection results?

[0006] Therefore, to improve the shortcomings of conventional PCR microfluidic detection procedures, it is necessary to develop a fluid cassette that is highly stable, simple and easy to operate, and can be used in portable nucleic acid detection devices, so that users can use and understand it easily with minimal training, and test results can be obtained quickly.

[0007] One of the objectives of the present invention is to provide a fluid cassette suitable for a portable nucleic acid detection device, in which a cap is sealed to the body and pressed downward, so that the inner tube on the body faces downward. The puncture structure at the end pierces the sealing membrane downward, and the sample in the inner tube and the buffer solution in the outer tube are released downward into the injection chamber of the detection box, and then flow through multiple flow channels to multiple reaction chambers, allowing multiple nucleic acid amplification and detection to be performed quickly in a single reaction chamber. This is easy to carry, and multiple nucleic acid amplification and detection can be performed quickly and easily anytime and anywhere, saving detection time and costs.

[0008] One of the objectives of the present invention is to provide a fluid cassette suitable for a portable nucleic acid detection device, which, through the design of multiple flow paths in the detection box of the fluid cassette and the phase change material contained therein, changes from a solid phase change material to a liquid phase change material during the heating process during multiplex nucleic acid amplification, and then its volume expands, achieving the sealing effect of a one-way valve, preventing the fluid in the flow path from flowing and preventing the fluids in each flow path and each reaction chamber from interfering with each other, thereby allowing multiple samples to be detected simultaneously and improving detection accuracy.

[0009] According to the concept of the present invention, there is provided a fluid cassette suitable for a portable nucleic acid detection device, comprising a cap, a main body, an inner tube, and a detection box. The main body has an outer tube and a shielding cover, and the outer tube is connected to the shielding cover. The inner tube is correspondingly disposed within the outer tube of the main body. The detection box is correspondingly disposed below the shielding cover and has an injection chamber, multiple flow channels, and multiple reaction chambers, the multiple reaction chambers being connected to the injection chamber via multiple flow channels. When a sample is introduced into the inner tube, it mixes with the liquid in the inner tube to form a test liquid, which is sealed to the outer tube of the main body via the cap. When the inner tube is pressed downward, the puncture structure of the inner tube pierces the sealing membrane of the detection box downward, allowing the test liquid to flow into the injection chamber and then through the multiple flow channels to the multiple reaction chambers for multiplex nucleic acid amplification. [Brief explanation of the drawings]

[0010] [Figure 1]1 is a schematic diagram showing an exploded structure of a fluid cassette according to a first preferred embodiment of the present invention and a portable nucleic acid detection device to which the fluid cassette is applied. [Figure 2A] FIG. 2 is a schematic diagram of the structure of the fluidic cassette and swab shown in FIG. 1. [Figure 2B] FIG. 2B is a schematic diagram of an exploded view of the fluidic cassette shown in FIG. 2A. [Figure 2C] FIG. 2B is a diagram of the assembled structure of the fluidic cassette shown in FIG. 2A. [Figure 2D] FIG. 2D is a schematic diagram of the cross-sectional structure of the fluidic cassette shown in FIG. 2C. [Figure 3A] FIG. 2C is a schematic diagram of the structure of the cap of the fluidic cassette shown in FIG. 2B. [Figure 3B] FIG. 3B is a schematic diagram of a cross-sectional structure of the cap shown in FIG. 3A. [Figure 3C] FIG. 2 is a schematic diagram of the structure of the cap of the second preferred embodiment of the present invention. [Figure 3D] FIG. 10 is a schematic diagram of the structure of the cap of the third preferred embodiment of the present invention. [Figure 4A] FIG. 4A is a schematic diagram of the structure of the inner tube of the fluid cassette shown in FIG. 2A. [Figure 4B] FIG. 4C is a schematic diagram of the structure of the inner tube shown in FIG. 4B viewed from above. [Figure 4C] FIG. 4C is a schematic diagram of the cross-sectional structure of the inner tube shown in FIG. 4A. [Figure 5] FIG. 5 is a schematic diagram of the structure of the inner tube of the fluid cassette according to the fourth preferred embodiment of the present invention. [Figure 6] 6A is a schematic diagram of the structure of the outer tube of the fluid cassette shown in FIG. 2B, 6B is a schematic diagram of the cross-sectional structure of the outer tube shown in FIG. 6A, and 6C is a schematic diagram of the top surface structure of the outer tube shown in FIG. 6A. [Figure 7] FIG. 7A is a schematic diagram of the structure of a fluidic cassette according to a fifth preferred embodiment of the present invention, and FIG. 7B is a schematic diagram of the structure of a fluidic cassette according to a sixth preferred embodiment of the present invention. [Figure 8] 8A-C are schematic diagrams of the process of inserting a swab into the fluidic cassette shown in FIG. 2A. [Figure 9] FIG. 9 is a schematic diagram of a cross-sectional structure of a fluidic cassette according to a seventh preferred embodiment of the present invention. [Figure 10A] FIG. 10A is a schematic diagram of the structure of the detection box of the fluidic cassette shown in FIG. 10B. [Figure 10B] FIG. 10B is an exploded view of the detection box shown in FIG. 10A. [Figure 10C] FIG. 10C is a bottom view of the detection box shown in FIG. 10A. [Figure 10D] FIG. 10D is a bottom view of the detection box shown in FIG. 10C from another angle. [Figure 10E] 10B is a schematic diagram of a partial cross-sectional structure of the detection box shown in FIG. 10A. [Figure 11A] FIG. 13 is a bottom view of the detection box of the fluidic cassette according to the eighth preferred embodiment of the present invention. [Figure 11B] FIG. 13 is a bottom view of the detection box of the fluidic cassette according to the ninth preferred embodiment of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0011] Several exemplary embodiments embodying the features and advantages of the present invention are set forth in detail in the following description. It is to be understood that the present invention may be modified in various ways in different aspects without departing from the scope of the present invention, and that the description and illustrations are intended to be illustrative in nature rather than restrictive.

[0012] Please refer to FIG. 1. FIG. 1 is a schematic diagram of the disassembled structure of a fluid cassette according to a first preferred embodiment of the present invention and a portable nucleic acid detection device incorporating the same. As shown in FIG. 1, the fluid cassette 2 of the present invention is suitable for use in the portable nucleic acid detection device 1, providing users with a portable detection device that is easy to carry and capable of amplifying and detecting multiple nucleic acids anytime and anywhere. In this embodiment, the fluid cassette 2 is a disposable consumable item. After a user completes one test, they can replace the fluid cassette 2 with a new one for the next multiplex nucleic acid test. In other words, the user does not need to assemble the fluid cassette 2 and can directly use the fluid cassette 2 with minimal operation steps, achieving simple and rapid multiplex nucleic acid amplification and detection. Please continue to refer to FIG. 1. In addition to the fluid cassette 2, the portable nucleic acid detection device 1 also includes a base 10, a housing 11, a circuit board 12, a heating module (not shown), and an optical module 14. In this embodiment, the heating module and optical module 14 are integrated on the circuit board 12, but this is not limiting. The fluid cassette 2 is correspondingly disposed on the circuit board 12 and its heating module. The base 10 has an accommodating space 100 for accommodating the circuit board 12, the heating module 14 thereon, the optical module 14, and the fluid cassette 2. The housing 11 is correspondingly fitted onto the base 10, sealing the accommodating space 100 of the base 10. The circuit board 12, the heating module 14 thereon, the optical module 14, and the fluid cassette 2 are effectively isolated from the external environment, thereby achieving a protective isolation effect. Furthermore, in this embodiment, the portable nucleic acid detection device 1 further includes a positioning sheet 19, which is disposed correspondingly between the circuit board 12 and the fluid cassette 2 and helps position the detection box body 21 of the fluid cassette 2 on the heating module and the optical module 14 of the circuit board 12. When a sample is accommodated in the fluid cassette 2, it is heated via a heating element (such as, but not limited to, a heating chip) of the heating module and maintained at a constant temperature.Then, a light source is emitted through the light emitter 141 of the optical module 14 to illuminate the sample, and a light sensor 142 receives the illuminated spectrum for detection and comparison. Furthermore, a plurality of indicator lights 143 are disposed on the circuit board 12 to correspondingly display the detection results.

[0013] In this embodiment, a button 101 and a display unit 102 are provided on the outer surface of the base 10. Both the button 101 and the display unit 102 are electrically connected to the circuit board 12. The button 101 is also electrically connected to a switch (or button) (not shown) on the circuit board 12. When a user presses the button 101 on the base 10, it is electrically connected to the switch (or button) on the circuit board 12, and the portable nucleic acid detection device 1 can be turned on or off. The display unit 102 is electrically connected to an indicator light 143 on the circuit board 12 and displays the detection result. In this embodiment, the display unit 102 may be, but is not limited to, a plurality of display windows that display optical signals after detection, allowing the user to quickly and easily understand the detection result. Taking this embodiment as an example, the positions of the indicator lights 143 on the circuit board 12 correspond to the display windows of the display unit 102 on the base 10. Therefore, when the indicator light 143 receives a signal and lights up, the user can quickly obtain the detection result through the lit display window of the display unit 102 on the base 10. However, in other embodiments, the display unit 102 may be, but is not limited to, a liquid crystal display (LCD). In order to display detailed test results, the display unit 102 is mainly used to display the test results, and its type and shape may be changed depending on the implementation situation and are not limited to this embodiment.

[0014] See Figures 2A to 2D. Figure 2A is a schematic diagram of the structure of the fluidic cassette and swab shown in Figure 1. Figure 2B is a schematic diagram showing the disassembled structure of the fluidic cassette shown in Figure 2A. Figure 2C is a diagram showing the assembled structure of the fluidic cassette shown in Figure 2A. Figure 2D is a schematic diagram of the cross-sectional structure of the fluidic cassette shown in Figure 2C. As described above, the fluidic cassette 2 is primarily suitable for use in a portable nucleic acid detection device 1 for sample processing and multiplexed nucleic acid amplification and detection. In this embodiment, it can be applied to PCR detection or isothermal PCR amplification and detection, and multiple samples can be detected simultaneously. As shown in Figure 2A, in some embodiments, a swab 3 can be used as an accessory to the fluidic cassette 2. The swab 3 may be any swab that can be attached to the fluidic cassette 2 and is commonly available commercially. Alternatively, in other embodiments, the sample can be added directly to the fluidic cassette 2 without using a swab 3. As shown in FIG. 2B , the fluid cassette 2 includes a cap 20, a main body 21, an inner tube 25, a detection box 24, etc., where the main body 21 has an outer tube 22 and a shield cover 23, and the outer tube 22 is connected to the shield cover 23. In some embodiments, the cap 20 seals the outer tube 22 through a seal ring 26, but is not limited to this. The inner tube 25 is correspondingly disposed within the outer tube 22 of the main body 21. The detection box 24 is correspondingly disposed below the shield cover 23 and is covered by the shield cover 23 to prevent evaporation of the liquid. When the cap 20 completely seals the outer tube 22 and the shield cover 23 correspondingly covers the detection box 24, the appearance is as shown in FIGS. 2A and 2C . As shown in Figures 2B and 2D, the detection box 24 has an injection chamber 243 (shown in Figure 2D), multiple flow paths 244 (shown in Figure 10C), and multiple reaction chambers 245 (shown in Figure 10C), and the multiple reaction chambers 245 are connected to the injection chamber 243 via the multiple flow paths 244.When a sample is placed in the inner tube 25 of the fluid cassette 2, it mixes with the liquid in the inner tube 25 to become the test liquid, which is then sealed by pressing it against the outer tube 22 of the main body 21. When the cap 20 is pressed downward, the inner tube 25 is also pressed downward, and the puncture structure 25e at the bottom of the inner tube 25 pierces the first sealing membrane 27a of the detection box 24 downward. The test liquid flows into the injection chamber 243 below the first sealing membrane 27a, and then flows through multiple flow paths 244 to each corresponding reaction chamber 245, whereupon the multiplex nucleic acid amplification process is performed.

[0015] Please refer to Figures 2D, 3A, 3B, 3C, and 3D simultaneously. Figure 3A is a schematic diagram of the structure of the cap of the fluid cassette shown in Figure 2B. Figure 3B is a schematic diagram of the cross-sectional structure of the cap shown in Figure 3A. Figure 3C is a schematic diagram of the structure of the cap of a second preferred embodiment of the present invention. Figure 3D is a schematic diagram of the structure of the cap of a third preferred embodiment of the present invention. As shown in Figures 3A and 3B, a cap 20 is used to seal the outer tube 22 of the fluid cassette 2. The cap 20 has a cap body 20a, a cover ring 20b, and a groove 20c. The cover ring 20b is connected to the cap body 20a and protrudes outward from the cap body 20a, and a groove 20c is provided on the cover ring 20b to accommodate a seal ring 26 (shown in Figure 2B). As shown in Figure 2D, when the cap 20 is sealed onto the outer tube 22, the cap body 20a is positioned within the outer tube 22, and the inner flat surface 20d of the cap body 20a of the cap 20 is abutted against the upper end surface 250 of the inner tube 25. Therefore, when the cap 20 is pressed downward, the inner tube 25 is also pressed downward at the same time, and the puncture structure 25e at the bottom of the inner tube 25 breaks through the first sealing membrane 27a of the detection box 24 (as shown in Figure 2B). In other embodiments, as shown in Figures 3C and 3D, the caps 200 and 201 also have cap bodies 200a and 201a and grooves 200b and 201b. In these two embodiments, the cap bodies 200a and 201a are further provided with exhaust structures to allow gas to escape and thereby maintain pressure balance within the fluidic cassette 2. 3C, the cap body 200a of the cap 200 is provided with an exhaust recess 200c as an exhaust structure. In this embodiment, the exhaust recess 200c may be, but is not limited to, an exhaust recess 200c two levels below the surface. In another embodiment, as shown in the 3D diagram, the cap body 201a of the cap 201 is provided with an exhaust groove 201c. The exhaust groove 201c penetrates the cap body 201a in the longitudinal direction and has two perpendicular L-shaped corners. The exhaust groove 201c penetrates the cap body 201a in the longitudinal direction, allowing gas in the fluid cassette 2 to be exhausted upward, thereby achieving an exhaust effect and a pressure balance maintenance effect.Furthermore, the design of the exhaust structure of the caps 200, 201 is not limited to the above two embodiments, and since the main purpose is to increase gas permeability, it can be changed according to the actual implementation situation.

[0016] See Figures 4A, 4B, and 4C. Figure 4A is a schematic diagram of the structure of the inner tube of the fluid cassette shown in Figure 2A. Figure 4B is a top view of the inner tube shown in Figure 4A. Figure 4C is a schematic diagram of the cross-sectional structure of the inner tube shown in Figure 4A. As shown in Figure 4A, the inner tube 25 of the fluid cassette 2 is primarily disposed within the outer tube 22 of the main body 21 and has a tapered, hollow tubular structure, but is not limited to this. In this embodiment, the inner tube 25 is disposed at the upper end of the inner tube 25 and includes a first tube portion 25c having a first opening 25a, a second tube portion 25d, and a piercing structure 25e. The swab 3 is correspondingly inserted into the inner tube 25 through the first opening 25a. In some embodiments, the diameter of the first tube portion 25c is larger than that of the second tube portion 25d, and the diameter of the second tube portion 25d is larger than that of the piercing structure 25e, and the second tube portion 25d is disposed between the first tube portions 25d. 25c and the puncture structure 25e form a space, forming the tapered hollow tubular structure of this embodiment. The puncture structure 25e is provided at the bottom of the inner tube 25 and has a sharp tip that can puncture the first sealing film 27a. In this embodiment, a second opening 25b is further provided at the end of the puncture structure 25e to connect the inner tube 25 and the outer tube 22 and effectively mix the liquids in the inner tube 25 and the outer tube 22. As shown in FIG. 4B, the diameter of the first opening 25a is larger than the diameter of the second opening 25b, but this is not limited thereto.

[0017] As shown in FIGS. 4A and 4B , the outer surface of the first tube portion 25c is provided with a plurality of convex structures 25f, which protrude from the outer surface of the first tube portion 25c. The inner tube 25 is correspondingly disposed within the outer tube 22 of the main body 21, and the convex structures 25f abut against the inner surface of the outer tube 22, thereby helping to support the inner tube 25. In this embodiment, the convex structures 25f are further inclined to accommodate downward pressure, allowing the inner tube 25 to slide downward along the inner surface of the outer tube 22 to a certain position, after which the convex structures 25f can be used. The stress between the convex structures 25f and the inner surface of the outer tube 22 prevents the inner tube 25 from sliding downward. The number and type of the convex structures 25f can be changed according to actual implementation situations and are not limited thereto.

[0018] In this embodiment, as shown in FIGS. 4B and 4C, a plurality of first ribs 25h are further provided on the inner surface of the second tubular portion 25d of the inner tube 25. These first ribs 25h extend vertically. The second tubular portion 25d has a long, slender rib structure that protrudes from the inner surface of the second tubular portion 25d. By providing the first ribs 25h on the inner surface of the second tubular portion 25d, the protruding first ribs 25h can be used to facilitate the removal of the swab 3 when inserting the swab 3 into the inner tube 25. The swab 3 and the first ribs 25h of the inner tube 25 increase the elution of the sample on the swab 3, improving the sample elution efficiency. In some embodiments, as shown in Figures 4A and 4C, the first tube portion 25c of the inner tube 25 further has a plurality of longitudinal grooves 25g on its outer surface adjacent to the second tube portion 25d, and these longitudinal grooves 25g cooperate with second ribs 22d on the inner surface of the outer tube 22 (as shown in Figure 6B), so that the inner tube 25 can pass through the longitudinal grooves 25g along the corresponding second ribs and slide downward.

[0019] Please refer to FIG. 5. FIG. 5 is a schematic diagram of the structure of the inner tube of a fluid cassette according to a fourth preferred embodiment of the present invention. In this embodiment, the external design of the inner tube 45 is slightly different from that of the previous embodiment, but is similarly applicable to the portable nucleic acid detection device 1 and its fluid cassette 2 of the previous embodiment. Like the previous embodiment, the inner tube 45 is also disposed within the outer tube 22 of the fluid cassette 2 and has a tapered hollow conical structure, but is not limited thereto. However, in this embodiment, the inner tube 45 has only a single tapered tube section 45c, a first opening 45a at the top of the tube section 45c, and a puncture structure 45d connected to the bottom end of the tube section 45c, with its sharp end penetrating the first sealing membrane 27a. In this embodiment, a second opening 45b is further provided at the end of the puncture structure 45d to connect the inner tube 45 and the outer tube 22, thereby effectively mixing the liquids in the inner tube 45 and the outer tube 22. As in the previous embodiment, the diameter of the first opening 45a is larger than the diameter of the second opening 45b, but this is not limited thereto. In this embodiment, multiple convex structures 45e are also provided on the outer surface of the upper end of the tubular portion 45c, and these convex structures 45e protrude from the outer surface of the tubular portion 45c. When the convex structures 45e shown in FIG. 45 are positioned on the outer tube 22, the convex structures 45e abut the inner surface of the outer tube 22 to help support the inner tube 45. Similarly, multiple vertical grooves 45f are formed on the inner surface of the tubular portion 45c adjacent to the puncture structure 45d, and these vertical grooves 45f are provided to communicate with the inner surface of the outer tube 45, thereby causing the upper second rib 22d to slide downward. Additionally, multiple vertical first ribs 45g are also provided on the inner surface of the inner tube 45 to improve sample elution efficiency, but this is not limited thereto. From the above two embodiments, it can be seen that the appearance of the inner tube 25, 45, the convex structures 25f, 45e, the longitudinal grooves 25g, 45f, and the positions of the first ribs 25h, 45g can be determined. These can be changed according to the actual implementation situation and are not limited to these. The structural design of the second rib 22d allows the swab 3 to rub against the second rib 22d, improving sample extraction efficiency.

[0020] See Figures 6A, 6B, and 6C. Figure 6A is a schematic diagram of the structure of the outer tube of the fluid cassette shown in Figure 2B. Figure 6B is a schematic diagram of the cross-sectional structure of the outer tube shown in Figure 6A. Figure 6C is a schematic diagram of the structure of the outer tube shown in Figure 6A viewed from above. In this embodiment, a buffer solution (e.g., a lysis buffer solution) is stored in advance in the main body 21 of the fluid cassette 2 to be mixed with the sample. As shown in the figures, the main body 21 of the fluid cassette 2 has an outer tube 22 and a shield cover 23, and the outer tube 22 is connected to the shield cover 23. In this embodiment, the outer tube 22 has a tube body 22a and a head 22e connected to the tube body 22a. The head 22e is located above the tube body 22a, and a third opening 22b is formed at its top. The head 22e and the lower part of the tube body 22a are disposed within the shield cover 23 so that the cap 20 can be sealed therewith, and a fourth opening 22c is formed at its bottom. In this embodiment, the diameter of the third opening 22b is larger than the diameter of the fourth opening 22c. In some embodiments, the shield cover 23 is a circular cover structure disposed around the outer tube 22, correspondingly shielding and covering the detection box 24 to prevent the liquid from evaporating or the liquid / vapor from leaking from the detection box 24 and causing unnecessary contamination to the environment. Covering the detection box 24 with the shield cover 23 forms a completely sealed environment during the reaction process, eliminating the risk of biological contamination of the external environment. Also, as shown in FIGS. 6B and 6C , the inner surface of the outer tube 22 may be provided with a plurality of second ribs 22d, each of which may have a bump 22f at its upper end, but this is not limiting. As shown in the figures, the second ribs 22d protrude from the inner surface of the outer tube 22 and correspond to the longitudinal grooves 25g of the inner tube 25. Therefore, when the inner tube 25 is placed inside the outer tube 22, the longitudinal grooves 25g (shown in FIG. 4A) are slid downward along the corresponding second ribs 22d, and when the ends of the longitudinal grooves 25g resist the bumps 22f, the sliding displacement is stopped, and the inner tube 25 is set. This prevents the inner tube 25 from slipping out of the outer tube 22.

[0021] Please refer to Figures 7A and 7B. Figure 7A is a schematic diagram of the structure of a fluid cassette according to a fifth preferred embodiment of the present invention. Figure 7B is a schematic diagram of the structure of a fluid cassette according to a sixth preferred embodiment of the present invention. As shown in Figure 7A, the external design of the fluid cassettes 5 and 6 is slightly different from that of the previous embodiment, but can also be applied to the portable nucleic acid detection device 1 of the previous embodiment. Similar to the previous embodiment, the fluid cassettes 5 and 6 also have structures such as caps 50 and 60, main bodies 51 and 61, outer tubes 52 and 62, shield covers 53 and 63, and detection boxes 54 and 64. The shield covers 53 and 63 differ in shape and height. For example, the shield cover 53 of the fifth embodiment is taller than the shield cover 23 of the previous embodiment and has a tall, cylindrical shield structure, while the shield cover of the sixth embodiment has a tapered shield structure. In other words, the shielding covers 53, 63 may be of any shape and height, and the purpose of their design is to ensure that the shielding covers 53, 63 have enough space to accommodate the pressure generated by the liquid vapor evaporated from the detection boxes 54, 64 during the heating reaction, without the resulting backpressure affecting the liquid inside the detection boxes 54, 64. In some embodiments, the shielding covers 53, 63 may be made of, but are not limited to, a hard material (e.g., metal, plastic, glass, etc.), a flexible material (e.g., rubber, silicone, etc.), or a mixture of hard and flexible materials (e.g., rubber / plastic, rubber / metal, rubber / glass, silicone / plastic, silicone / metal, silicone / glass, etc.).

[0022] See Figures 8A-8C. Figures 8A-8C are diagrams illustrating the process of inserting a swab into the fluidic cassette shown in Figure 2A. As shown in these figures, in this embodiment, the sample processing and multiplex nucleic acid amplification / detection procedure for the fluidic cassette 2 includes the following steps: First, the user collects a sample to be tested using a swab 3 and peels off the sealing film (not shown) that seals the third opening 22b of the outer tube 22. Next, as shown in Figure 8A, the user inserts the swab 3 into the inner tube 25 and rotates it several times to fully contact the first rib 25h of the inner tube 25, thereby improving the sample elution efficiency. In some embodiments, if the user is using other types of samples (e.g., blood, saliva), the user can add the sample directly to the inner tube 25 without using a swab 3. As shown in FIG. 8B, after the above steps are completed, the user can either remove the swab 3 or, without removing the swab 3, directly press it into the cap 20 corresponding to the third opening 22b of the outer tube 22 to seal the cap. The cap 20 is sealed to the head 22e of the outer tube 22 via the sealing ring 26. At this time, based on the downward pressure of the cap 20, the inner tube 25 also slides downward. As shown in FIG. 8C, the puncture structure 25e at the end of the inner tube 25 punctures the corresponding first sealing membrane 27a downward, and the sample in the inner tube 21 and the buffer solution in the outer tube 22 pass through the second opening 25b at the end of the inner tube 21 and the fourth opening 22c at the end of the outer tube 22, respectively, and are released downward to enter the detection box 24 below.

[0023] In some embodiments, to ensure that the reaction solution is not acidified by bubbles in the reaction chamber 245 during isothermal amplification, it is necessary to prevent a large amount of bubbles from remaining in the reaction chamber 245 after injection. In this embodiment, when the sealing ring 26 is attached, the downward pressure of the cap 20 increases with the speed at which the user presses the cap 20. If the downward pressure is too high, bubbles will form in the reaction chamber 245 in the detection box 24 below. In other embodiments, the sealing rings 26 and 29 may not be installed to allow the buffer solution to enter the reaction chamber 245 more smoothly, stably, and quickly. Even if the sealing rings 26 and 29 are not installed, the cap 20 may generate momentary pressure on the buffer solution, which may mix with air under the first sealing membrane 27a (e.g., an aluminum membrane) at the moment of puncture and push into the injection chamber 243 of the detection box 24. This air may block the flow path 244 connected to the injection chamber 243, preventing one or more reaction chambers 245 from being properly injected. Air bubbles are blown directly from the injection chamber 243 and pushed into the reaction chamber 245. To improve the above-mentioned problem, one solution is to provide an exhaust structure to the caps 200 and 201, as shown in FIG. 3C or 3D. As mentioned above, the exhaust structure of the cap 200 in FIG. 3C is an exhaust recess 200c on the surface, while the exhaust structure of the cap 201 in FIG. 3D is an exhaust groove 201c running vertically through the surface, and the exhaust groove 201c has a structure with two perpendicular L-shaped corners. Comparing the two, the exhaust structure shown in FIG. 3D (exhaust groove 201c) has a higher air permeability per unit time than the exhaust structure shown in FIG. 3C (exhaust recess 200c). The main reason for this is that the exhaust groove 201c is a groove that penetrates the cap body 201a of the cap 201, and therefore the compressed air is discharged upward through the exhaust groove 201c that penetrates the cap body 201a, resisting the cap. If the tolerance is exceeded, the interference between the cap 201 and the shield cover 23 increases, causing a problem that the pressure generated during capping affects the infusion.Furthermore, if the interference of the cap 201 is large and the cap 201 is pressed down quickly, a portion of the reaction chamber 245 may be prematurely immersed in the liquid, potentially damaging the freeze-dried beads 245d disposed therein. Before the air in the beads dissolves, the upper opening 245a is blocked by the liquid, preventing the original air in the reaction chamber 245 and the air remaining after the freeze-dried beads 245d dissolve from escaping through the upper opening 245a. This also means that in other embodiments, even if the sealing rings 26 and 29 are not provided, the tight fit between the cap 201 and the shield 23 still requires an additional exhaust mechanism design to reduce air bubbles, which could potentially block the injection chamber 243. As mentioned above, when the interference is large, the design of the exhaust groove 201c shown in FIG. 3D can successfully achieve bubble-free injection. If the interference between cap 201 and shield cover 23 is small, air can be smoothly expelled from the gap around cap 201, which is relatively loose, when cap 201 is pressed down, achieving the goal of bubble-free injection. In the embodiment shown in the 3D diagram, considering two variables—the force and speed with which the user presses down on cap 201—the success rate of completely bubble-free injection can exceed 80%. These two different exhaust structures allow interference between cap 201 and shield cover 23, preventing liquid leakage even when fluid cassette 2 is shaken up and down or turned upside down. Furthermore, during the heating process in an unsealed state, excessive pressure can be prevented from causing leakage of the pressurizing membrane or first sealing membrane 27a or preventing pressure from being released. Between the shield cover 23 and the detection box 24, and between the cap 20, air expanded by heating can be discharged to the outside of the fluid cassette 2, avoiding excessive imbalance pressure within the fluid cassette 2 and causing liquid to be pushed back from one or more reaction chambers 245 into the injection chamber 243. This design with exhaust channels has also been proven by verification that amplified contaminants are captured by the shield 23 and do not contaminate the machine or the surrounding environment from the fluid cassette 2.

[0024] Please refer to FIG. 9. FIG. 9 is a schematic cross-sectional view of a fluid cassette according to a seventh preferred embodiment of the present invention. As shown in FIG. 9, in this embodiment, the fluid cassette 7 includes a cap 70, a main body 71, an outer tube 72, a shielding cover 73, a detection box 74, an inner tube 75, and other components. The assembly and arrangement of these components are the same as in the previously described embodiments, and therefore will not be described again here. However, in this embodiment, a separate sealing ring is not required when sealing the cap 70 to the outer tube 72, and a small gap can be left. Therefore, if air expands in the outer tube 72 during the heating process of multiple isothermal nucleic acid amplifications, the air can escape to the outside through the gap between the cap 70 and the outer tube 72, thereby maintaining the internal stability of the system. Furthermore, in this embodiment, when the shielding cover 73 covers the detection box 74, a gap 76 may be formed at the joint between the two. Similarly, this small gap 76 allows the air that expands due to heating inside the shield cover 73 to escape to the outside, maintaining pressure stability inside and outside the shield cover 73 without affecting the fluid pressure in the detection box 74, stabilizing the fluid pressure in the detection box 74, and enabling multiple isothermal nucleic acid amplification and detection procedures to be performed under stable conditions.

[0025] See also Figures 10A, 10B, and 10C. Figure 10A is a schematic diagram of the structure of the detection box of the fluid cassette shown in Figure 2B. Figure 10B is a schematic diagram of the exploded structure of the detection box shown in Figure 10A. Figure 10C is a bottom view of the detection box shown in Figure 10A. As shown in Figure 10A, in this embodiment, the detection box 24 has a disk-shaped box structure and includes a box body 240 and three types of sealing films 27a, 27b, and 28. As seen in Figures 10B and 10C, the cassette body 240 has an injection chamber 243, multiple flow channels 244, and multiple reaction chambers 245. In this embodiment, the injection chamber 243 is located in the center of the cassette body 240. The detection box 24 is a chamber that penetrates an upper surface 241 and a corresponding lower surface 242 (as shown in Figure 10C) and is connected to the multiple reaction chambers 245 via multiple flow channels 244. In this embodiment, each flow path 244 is designed to have the same flow resistance, thereby ensuring equal and uniform distribution of liquid from the injection chamber 243 to all reaction chambers 245, and achieving detection accuracy. Also, in this embodiment, the number of reaction chambers 245 and the flow paths 244 connected thereto is five, but is not limited to this.

[0026] As shown in FIGS. 10A and 10B, the first sealing membrane 27a is located at the center of the top surface 241 of the detection box 24, corresponding to the injection chamber 243 for the outer tube 22. The bottom opening 22c is isolated from the detection box 24, preventing the buffer solution in the outer tube 22 from entering the detection box 24 before use and allowing the buffer solution to be continuously stored in the outer tube 22 when unused. The second sealing membrane 27b is also located on the top surface 241 of the detection box 24, corresponding to the reaction chamber 245, and is used to seal the top opening 245a of the reaction chamber 245 in the detection box 24. In this embodiment, the second sealing membrane 27b is a smaller circular membrane piece, and each circular membrane piece covers one reaction chamber 245. Therefore, in this embodiment, there are five second sealing membranes 27b, which cover five reaction chambers 245. In other embodiments, the second sealing membrane 27b may be a large membrane covering the upper surface 241 and all of the reaction chambers 245. Its shape and quantity can be varied depending on the actual implementation, but are not limited to this. In this embodiment, the second sealing membrane 27b may be a single-layer or multi-layer hydrophobic porous membrane, but is not limited to this. Furthermore, the second sealing membrane 27b can also be made of other waterproof (i.e., hydrophobic) and breathable materials to seal the top of the reaction chambers 245, preventing liquid intrusion while allowing air to pass through. This allows the second sealing membrane 27b to be used as an air outlet during the heating process required for the nucleic acid amplification process to release the pressure of the fluid load due to the water inlet pressure (WEP) of the second sealing membrane 27b. Since the WEP is higher than the driving pressure of the pressurized fluid, it may clog the fluid. Furthermore, the hydrophobic surface of the second sealing membrane 27b helps liquid condense on the membrane, thereby slowing the evaporation process. In other words, experiments have also proven that the second sealing membrane 27b can effectively suppress liquid evaporation. The liquid in the reaction chamber 245 is evaporated to facilitate the nucleic acid amplification and detection procedures. In another embodiment, the third sealing membrane 28 is disposed on the bottom of the detection box 24 so as to cover the lower surface 242 of the detection box 24, but is not limited to this.

[0027] See also Figures 10C, 10D, and 10E. Figure 10D is a bottom view of the detection box shown in Figure 10C from another angle. Figure 10E is a schematic diagram of the partial cross-sectional structure of the detection box shown in Figure 10A. As shown in the figure, from the bottom view of the detection box 24, there is an injection chamber 243 located in the center of the detection box 24, five reaction chambers 245 equally spaced around it, and five flow paths 244 for the corresponding injection chambers 243 and reaction chambers 245. As shown in Figure 10E, each reaction chamber 245 includes a chamber 245b and a top structure 245c connected and disposed above the chamber 245b. In this embodiment, the top structure 245c is a roof structure having, but is not limited to, four slopes. The top structure 245c has a circular top opening 245a connected to the upper surface 241 of the detection box 24. The top structure 245c is primarily used to help air completely evacuate from the reaction chamber 245 through the top opening 245a of the roof structure during the fluid loading process. For example, if air bubbles (not shown) are formed during the fluid injection process, they automatically rise to the top of the "roof structure" of the top structure 245c in response to fluid pressure and are then discharged through the top opening 245a. In some embodiments, the top structure 245c also has another function of accommodating air bubbles (not shown) that are generated during the heating process and cannot be discharged. This means that any air bubbles that cannot be discharged will rise. In this way, the "roof structure" of the top structure 245c can keep the chamber 245b below it free of air bubbles, thereby avoiding interference and optical detection errors caused by air bubbles. In this embodiment, the optical detection path is located at the mid-height of the chamber 245b, and there is a certain buffer space between the optical detection path and the top structure 245c to prevent unexpected bubbles from being generated in the chamber 245b. It can be seen that the roof structure of the top structure 245c of the reaction chamber 245 serves as a protection mechanism to ensure that no bubbles or air are present in the fluid, thereby achieving consistent and accurate optical detection.

[0028] Continuing to refer to FIG. 10E, in some embodiments, the reaction chambers 245 are provided with freeze-dried beads 245d made of freeze-dried reagents (e.g., enzymes, buffers, dyes, etc.). In this embodiment, the freeze-dried beads 245e are stored in the chambers 245b and separated from the buffer solution. When the puncture structure 25e at the end of the inner tube 21 of the fluid cassette 2 pierces the first sealing membrane 27a, the sample in the inner tube 21 and the buffer solution in the outer tube 22 are released downward into the injection chamber 243 and flow into the corresponding reaction chambers 245 through the flow channel 244. At the same time, the freeze-dried beads 245d in the chambers 245b are also melted and mixed with the sample by heating using a heating module (not shown). A multiplex nucleic acid amplification process is then performed, followed by nucleic acid detection for the reaction reagents in each reaction chamber 245 via the optical module 14. In another embodiment, freeze-dried beads 245d are placed in the reaction chambers 245, and small pieces of phase-change material, such as wax, are placed in each reaction chamber 245 before the pressure-sensitive bottom film is sealed. The volume of this phase-change material needs only to be sufficient to block the top opening 245a; if too much is placed, expansion and air bubbles may form on the bottom surface, affecting optical detection under the "roof structure." The design purpose of placing additional phase-change material is primarily to allow the phase-change material to melt during the heating process, since its density is lower than that of the liquid, and naturally migrate upward to block the top opening 245a, thereby reducing thermal expansion. When the liquid in the reaction chambers 245 comes into contact with the liquid, the liquid becomes acidic, ensuring more accurate detection.

[0029] See Figures 10C and 10D. As shown, the channels 244 connecting the injection chamber 243 and the reaction chamber 245 can be designed in various ways and can have any shape, as long as each channel 244 has the same fluid resistance from the injection chamber. This allows the liquid to be uniformly distributed into the target reaction chambers 245 simultaneously. In some embodiments, the time difference between fluids entering different reaction chambers 245 is less than 30 seconds. This time difference is primarily due to small differences in channel resistance created during manufacturing, but if the reaction chambers 245 are completely filled, the acceptable time difference range is within 1 minute. In other embodiments, the width and height of the channels 244 range from 0.01 to 20 mm. As shown in Figure 10C, in this embodiment, the channels 244 have two grooves 244a for accommodating corresponding phase transition materials (PTMs), such as paraffin beads or hydrogel, but are not limited thereto. Taking this embodiment as an example, each corresponding groove 244 is used to accommodate paraffin beads 246. During the heating process of nucleic acid amplification, the paraffin beads 246 melt, expanding their volume by 15% during the phase change, thereby filling and sealing the space within the channel 244. In some embodiments, the paraffin beads 246 may have a diameter of 2 mm and a thickness of 0.7 mm, but are not limited thereto, and can be mass-produced using a silicone mold. Because these phase-change materials are liquid, they are impermeable to the liquid within the reaction chamber 245, locking the reaction chamber 245 and preventing backflow or leakage of the fluid within the reaction chamber 245 into the channel 244. One-way fluid transfer to other channels 244 and reaction chambers 245 is achieved. In other words, the channel design with grooves 244a for accommodating the phase-change material can function as a one-way valve, transporting sample and buffer solution in one direction to the reaction chamber 245 for nucleic acid amplification and detection. Backflow does not interfere with the nucleic acid amplification and detection results in other reaction chambers 245.In this way, the five reaction chambers 245 in the detection box 24 can independently amplify and detect nucleic acids without mutual interference or backflow of liquids affecting the detection results, thereby achieving accurate and effective amplification and detection of multiple nucleic acids. Furthermore, in this embodiment, the sealing mechanism of the phase-change material can also prevent the reagents in the reaction chambers 245 from being diluted by the sample and buffer solution from the channel 244 and the injection chamber 243, so the concentration of the reagents in the reaction chambers 245 is not reduced. This ensures reliable sample retention, enables highly accurate and precise production, and produces accurate and sensitive results.

[0030] In this embodiment, as shown in FIG. 10D , the microchannels 244b connected to the front of each reaction chamber 245 further include an extension portion that extends toward the reaction chambers 245. The material is contained therein, thereby reducing the penetration of the molten phase-change material into the reaction chambers 245. The melting time of a phase-change material is the time required for a solid phase-change material to change to a liquid state. In some embodiments, the melting temperature of the phase-change material can be selected to formulate a specific heating process suitable for the reaction. For example, by selecting an appropriate temperature, a solid phase-change material can melt and complete its previous phase within one minute. The sealing mechanism of the material can be changed, but is not limited to this. This mechanism uses the water pressure difference created by the injection of a buffer solution and the phenomenon in which the pressure of the fluid cassette 2 is generally directed toward the five reaction chambers 245 during heating to push the molten phase-change material forward from the grooves 244a. The sealing effect can be achieved by using different channel designs, such as an expanded structure at the front end of two grooves 244a (microchannel 244b shown in FIG. 10D) or a narrowed structure with a single groove 844a (microchannel 844b shown in FIG. 11A). When the fluid cassette 2 is heated and expands, the phase change material automatically melts and expands, automatically transferring less than 1 microliter of phase change material into the sealed structure, achieving the sealing effect. Compared to other conventional blocking methods, the isolation mechanism of these embodiments is less expensive and does not require electrical or mechanical control programs, resulting in significant cost savings. Furthermore, because the paraffin in the phase change material is an insulator, the blocked reaction chamber 245 can be insulated and made non-conductive, allowing electrochemical detection within the reaction chamber 245. In some embodiments, the detection box 24 with the phase change material sealing mechanism completed is used for injection testing. However, even if the solid phase change material changes to a liquid phase change material and the sealing mechanism is completed, the detection box 24 can be tilted at a 60° angle without affecting the process of loading fluid into each reaction chamber 245. Even if the detection box 24 is tilted at 90°, the stability of the fluid during the isothermal heating process is not affected.Furthermore, the sealing mechanism of the phase-change material also provides robust protection against dropping the detection box 24 during or after the constant temperature heating process. Experiments have shown that even when the detection box 24 is dropped from a height of 1.3 meters, the liquid in each reaction chamber 245 remains full and no air bubbles occur that would interfere with optical detection, confirming the robust performance of this microfluidic system.

[0031] Referring to FIG. 8A, the shield cover 23 is attached to the detection box 24. Therefore, when vapor or molecules escape from the upper opening 245a of the reaction chamber 245 of the detection box 24, they are contained in the internal space of the shield cover 23. The phase-change material sealing mechanism can prevent changes in the pressure difference between the shield cover 23, the inner tube 21, and the outer tube 22, thereby maintaining the stability of the fluid. For example, without the phase-change material for sealing, when the fluid cassette undergoes the heating process of isothermal nucleic acid amplification, the pressure generated by the thermal expansion of the air inside the shield cover 23 would be higher than the pressure inside the shield cover 23. The liquid in the reaction chamber 245 could be pushed back along the flow path 244 and the injection chamber 243 into the inner tube 21, resulting in reagent mixing, detection failure, and other consequences. Therefore, the phase-change material sealing mechanism can effectively block the pressure difference between the reaction chamber 245 and the shield cover 23, preventing the pressure difference from affecting the stability of the fluid.

[0032] Please refer to Figures 11A and 11B. Figure 11A is a bottom view of a detection box of a fluid cassette according to an eighth preferred embodiment of the present invention. Figure 11B is a bottom view of a detection box of a fluid cassette according to a ninth preferred embodiment of the present invention. As shown in the figures, in these two embodiments, detection boxes 84 and 94 are also suitable for use with the fluid cassette 2 and are similar to the previous embodiments. The detection boxes 84 and 94 each have an injection chamber 843 and 843, respectively, and are connected to five equally spaced reaction chambers 845 and 945 and five flow channels 844 and 944 corresponding to the injection chambers 843 and 943 and the reaction chambers 845 and 945. However, in these two embodiments, the flow channels 844 and 944 connected to the injection chambers 843 and 943 and the reaction chambers 845 and 945 have only one groove 844a and can accommodate only one paraffin bead 246. Another difference between these two embodiments is that the width and shape of the microchannels 844b, 944b connected to the injection chambers 843, 943 are slightly different. Each microchannel 844b shown in FIG. 11A tapers toward the injection chamber 843. The microchannel 944b connected to the front of each reaction chamber 945 shown in FIG. 11B tapers toward the reaction chamber 945, but is not limited to this. This can help determine the length, width, depth, and other dimensions of the channels 844, 944, as well as whether or not grooves 844a, 944a for phase-change materials need to be provided, and the number of grooves 844a. Implementations may vary depending on the actual implementation situation, and are not limited to this. In this embodiment, liquid flows from the channel 944 to the reaction chamber 945, melting the freeze-dried beads 245d in the reaction chamber 945. In some embodiments, the extended channel structure connecting the paraffin beads 246 and the reaction chamber 245 allows for slight back-diffusion of the buffer solution. During heating, the diffused buffer solution is pushed back into the reaction chamber 245, which inevitably affects the reaction efficiency.

[0033] Looking back at the embodiment of the present invention, as shown in FIG. 10D, in this embodiment, the extended structure of the microchannel 244b connected between the groove 244a and the reaction chamber 245 can accommodate more molten phase-change material, thereby preventing the molten phase-change material from flowing into the reaction chamber 245. Furthermore, in another embodiment, the problem of back-diffusion of liquid is solved by changing the microchannel 244b shown in FIG. 10D from an extended structure to a microchannel 944b between the connecting groove 944a, as shown in FIGS. 11A and 11B. The reaction chamber 945 has a constricted structure. These changes in the channel design reduce the molecular diffusion rate between the reaction chamber 945 and the channel 944, thereby preventing the freeze-dried beads 245d from melting in the reaction chamber 945 and then being diluted out of the channel 944. Furthermore, in the embodiment shown in Figures 11A and 11B, the cross-sectional area of ​​the constriction structure between the reaction chamber 945 and the microchannel 944b is smaller than that of the expansion structure between the reaction chamber 245 and the microchannel 244b in the previous embodiment (i.e., the embodiment shown in Figures 10C and 10D). The expansion structure between the reaction chamber 245 and the microchannel 244b is significantly different, with a difference of approximately nine times. Experimental results show that the constriction structure of the microchannel 944b in Figures 11A and 11B has excellent ability to suppress back-diffusion of the buffer solution. However, to shorten the injection time, the liquid inlet of the microchannel 944b is enlarged, resulting in faster injection and better ability to suppress diffusion from the start of injection to the end of the heating reaction. This optimized design can achieve an optimal and consistent reaction concentration in the reaction chamber 945. Therefore, the microchannel 944b and the reaction chamber 945, combined with the flow channel design of the narrow structure between them, simultaneously achieve the effects of rapid injection and diffusion suppression. In another embodiment, if the reaction chamber 945 does not require electrical insulation, there is no need to place a phase change material in the groove 944a and the microchannel 944b, and the inlet of the microchannel 944b can be designed to be flush with the groove 944a. The groove 944a is flat, and the channel width gradually narrows from 400 microns at the inlet to 200 microns at the constriction structure.

[0034] This streamlined design allows for faster injection, and the simple fluid pipeline design can suppress back-diffusion or cross-contamination of liquids within the reaction chamber 945. This simplified fluid cassette 2 is not only suitable for commercialization, but also simplifies manufacturing costs and labor and supports optical signal detection within the reaction chamber 945. Furthermore, in other embodiments, if the reaction chamber 945 needs to be electrically isolated, the groove 944a can be designed to accommodate a phase-change material. In this case, whether it is a single wax groove (e.g., the groove 944a shown in FIG. 11B) or two wax grooves (e.g., the groove 244a shown in FIG. 10D), a blocking structure capable of being filled with the expanded phase-change material is required. The filled blocking structure is either an expansion structure of the microchannel 244b provided between the groove 244a and the reaction chamber 245 or a constriction structure of the microchannel 944b provided between the groove 944a and the reaction chamber 945. The insulating barrier requires melting the phase-change material, which is forced into the barrier structure by the differential pressure of microfluidics, preventing it from entering the reaction chambers 245 and 945 in large quantities. As shown in FIG. 11B, the expanded liquid inlet of microchannel 944b and the constricted structure at the front of reaction chamber 945 achieve excellent diffusion suppression without the need for electrically insulating phase-change material, and also prevents cross-contamination between reaction chambers 945. This makes the non-electrochemical detection fluid cassette 2 easier and more effective to operate and use, further contributing to its commercialization.

[0035] In summary, the present invention provides a fluid cassette suitable for a portable nucleic acid detection device. When the cap is sealed to the body and pressed downward, the inner tube of the body faces downward. The puncture structure at the end displaces downward to pierce the sealing membrane, releasing the sample in the inner tube and the buffer solution in the outer tube downward into the injection chamber of the detection box, and then through the multiple flow channels to multiple flow channels. Multiplexed nucleic acid amplification and detection can be performed quickly and easily using a single reaction chamber. This allows for easy portability and allows for fast and easy multiplexed nucleic acid amplification and detection anytime, anywhere, thereby saving time and money. Furthermore, the design of the multiple flow channels and the phase change material contained within the detection box of the fluid cassette allows the phase change material to change from solid to liquid during the heating process of multiplexed nucleic acid amplification. This large volume allows the phase change material to expand, creating a one-way valve sealing effect. This prevents the fluid in the flow channels from flowing back into the injection chamber, and prevents the fluids in each flow channel and each reaction chamber from interfering with each other, allowing for simultaneous detection of multiple samples and improving detection accuracy. In addition, the present invention provides a portable nucleic acid detection device in which the stability of the detection box is greatly improved by the sealing mechanism of the phase-change material, so that tilting or vibration does not affect the liquid in the detection box, and the accuracy of the detection results is not affected even if the device is tilted, vibrated, or dropped when being carried. [Explanation of symbols]

[0036] 1: Portable nucleic acid detection device 10: Bass 100: Containment space 101: Button 102: Display section 11: Housing 12: Circuit board 14: Optical module 141: Luminous object 142: Optical sensor 143: Indicator light 19: Positioning sheet 2, 5, 6, 7: Fluid cassette 20, 50, 60, 70, 200, 201: Cap 20a, 200a, 201a: Cap body 20b: Covering 20c, 200b, 201b: groove 20d: Internal plane 200c: Exhaust dent 201c: Exhaust groove 21, 51, 61, 71: Main unit 22, 52, 62, 72: Outer tube 22a: Pipe body 22b: Third opening 22c: 4th opening 22d: Second rib 22e: Head 22f: Bump 23, 53, 63, 73: Shield cover 24, 54, 64, 74, 84, 94: Detection box 240: Box body 241:Top surface 242: Bottom surface 243, 843, 943: Injection room 244, 844, 944: Flow path 244a, 844a, 944a: groove 244b, 844b, 944b: Microchannels 245, 845, 945: Reaction chamber 245a: Upper opening 245c:Top structure 245b: Chamber 245d: freeze-dried beads 246: Paraffin beads 25, 45, 75: Inner tube 250: Upper end surface 25a, 45a: 1st opening 25b, 45b: 2nd opening 25c: 1st pipe section 25d: 2nd pipe section 25e, 45d: Puncture structure 25f, 45e: Convex structure 25g, 45f: Vertical groove 25h, 45g: First rib 26, 29: Seal ring 27a: First sealing film 27b: Second sealing film 28: Third sealing film 3: Swab 45c: Pipe section 76: Gap

Claims

1. A fluid cassette for use in a portable nucleic acid detection device, the fluid cassette comprising: Cap and a main body including an outer tube and a shield cover, the outer tube being connected to the shield cover; an inner tube disposed within the outer tube of the body; a detection box disposed under the shield cover and having an injection chamber, a plurality of flow channels, and a plurality of reaction chambers, the plurality of reaction chambers being in communication with the injection chamber via the plurality of flow channels; When a sample is placed in the inner tube, the sample mixes with the liquid in the inner tube to become a test liquid, and a cap seals the outer tube of the main body and presses the inner tube downward using the cap, causing the puncture structure of the inner tube to move downward and break through the sealing membrane of the detection box, and the test liquid is injected into the injection chamber and flows through multiple flow paths to multiple reaction chambers to perform multiple nucleic acid amplification.

2. The fluid cassette of claim 1 , further comprising a sealing ring disposed between the cap and the outer tube of the body.

3. The fluid cassette of claim 1 , wherein the cap further comprises a vent structure.

4. 2. The fluid cassette according to claim 1, wherein the sealing membrane is a first sealing membrane that covers an upper surface of the detection box and is positioned to correspond to a position of the injection chamber.

5. The fluid cassette of claim 1 , wherein the detection box further comprises a plurality of second sealing membranes covering an upper surface of the detection box, the second sealing membranes being positioned to correspond to the positions of the plurality of reaction chambers.

6. The sealing film is a first sealing film covering an upper surface of the detection box, and is positioned to correspond to the position of the injection chamber; the detection box further includes a plurality of second sealing films covering an upper surface of the detection box, the second sealing films being positioned to correspond to the positions of the plurality of reaction chambers; The fluid cassette of claim 1 , wherein the first sealing membrane and / or the second sealing membrane are made of a waterproof and breathable material.

7. The fluid cassette of claim 1 , wherein the detection box further comprises a plurality of third sealing membranes covering a lower surface of the detection box.

8. The fluid cassette according to claim 1 , wherein the outer tube of the body is provided with a head for sealing the cap.

9. The fluidic cassette of claim 1 , wherein the inner tube is a tapered hollow tubular structure.

10. 2. The fluid cassette of claim 1, wherein the inner tube has a first tube portion, a second tube portion, and a puncture structure, the second tube portion being disposed between the first tube portion and the puncture structure, the diameter of the first tube portion being larger than the diameter of the second tube portion, and the diameter of the second tube portion being larger than the diameter of the puncture structure.

11. The fluidic cassette of claim 10 , wherein the first tube portion of the inner tube has a plurality of convex structures.

12. The fluid cassette of claim 10 , wherein a plurality of first ribs are formed on the inner surface of the second tube portion of the inner tube.

13. 11. The fluid cassette of claim 10, wherein the first tube section of the inner tube has a plurality of longitudinal grooves on an outer surface thereof adjacent the second tube section.

14. 2. The fluid cassette of claim 1, wherein the inner surface of the outer tube of the body is provided with a plurality of second ribs, each having a bump at its top.

15. 2. The fluidic cassette of claim 1, wherein each reaction chamber includes a chamber and a top structure, the top structure being disposed above and in communication with the chamber, and a lyophilized reagent being disposed within the chamber.

16. The fluidic cassette of claim 1 , wherein each channel of the detection box has at least one groove for receiving a phase change material.

17. 17. The fluid cassette of claim 16, wherein each flow path of the detection box has two grooves, and the flow path connected to the front of each reaction chamber further has an extension, the extension facing the reaction chamber.

18. 17. The fluid cassette of claim 16, wherein each flow channel of the detection box has a single groove, and the flow channel connected to the front of each reaction chamber tapers toward the reaction chamber.

19. 20. The fluidic cassette of claim 18, wherein each channel of the detection box comprises a microchannel, each microchannel being connected to the inlet chamber and tapering toward the inlet chamber.

Images