A centrifugal microfluidic chip

Abstract

The present invention provides a centrifugal microfluidic chip, belonging to the technical field of microfluidics, which includes a chip body; an upstream cavity located on the chip body; a downstream cavity located on the chip body, and the far-center end side of the upstream cavity is communicated with the downstream cavity through a flow channel; a repeat switching valve provided on the flow channel, with a phase change material disposed inside, and by controlling heating and rotation speed, the upstream cavity and the downstream cavity can be blocked or communicated repeatedly. The present invention is composed of a three-way path, a phase change material and an energy storage device connected thereto, and can conveniently achieve the controllable batch release and precise quantitative release of the upstream liquid. Its structural composition is simple, which is convenient for reducing the production and manufacturing cost and simplifying the complexity of fluid controlled release.

Description

Technical Field

[0001] This application relates to the field of microfluidic technology, and particularly to a centrifugal microfluidic chip. Background Art

[0002] In recent years, with the in-depth cross-development of multiple disciplines such as physics, material chemistry, mechanical engineering, and biotechnology, the technology for manipulating tiny fluids (abbreviated as "microfluidic technology") has been continuously developed and innovated, keeping pace with the times. Due to characteristics such as miniaturization, integration, and high automation, microfluidic chips have been widely applied in fields such as biomedical detection and synthetic biology. Microfluidic chips for loading and running various tiny-volume fluid reagents are gradually becoming the most effective carrier form for efficient in vitro diagnosis and high-precision biochemical synthesis. Among them, an important research field of high-precision synthetic biology is synthetic genomics, such as the synthesis of oligonucleotides with specific sequences, which has very wide applications in molecular biology fields such as gene sequencing and gene chips.

[0003] In-situ synthesis of oligonucleotides with specific sequences is the high-throughput synthesis of oligonucleotides on a solid chip substrate, with advantages such as high throughput and high precision. Microfluidic chips can flexibly release, transfer, mix, and separate liquid components such as reagents and samples. Therefore, in-situ synthesis of oligonucleotides based on microfluidic chips is more flexible and efficient. The efficient and precise processing of liquid behavior by microfluidic chips relies on various microvalves integrated inside them. These microvalves and microvalve-related components are the key to guiding liquid flow or determining the liquid movement mode. For a microfluidic chip used for in-situ synthesis of oligonucleotides with specific sequences (taking the most commonly used phosphoramidite chemistry method as an example), it needs to achieve the repeated cyclic release of liquid reagents such as deprotecting agents, active nucleotide monomers, oxidants, and capping agents in terms of function. Therefore, it has higher requirements for the selection of microvalve types and the implementation of valve control methods. The valve control methods used in current synthesis process methods (such as mechanical valves, pneumatic valves, etc.) generally face problems such as complex component compositions, bulky external devices, and high costs. As a most typical and commonly used phase change valve, the paraffin valve is characterized by a simple structure and convenient opening and closing. Generally, no additional complex component modules are required except for a simple heating module, and it hardly affects the activity of various biochemical reagents, the detection, and the occurrence of synthesis reactions. It is an ideal microfluidic valve.

[0004] However, there are still some problems in the application of paraffin valves in the current microfluidic chip field. For example, the inability to achieve repeated opening and closing leads to limited applications, the cavities generated by the cooling and contraction of paraffin affect the tightness and reliability of the valve body, and the structure of the paraffin valve components is too complex. These problems often limit the functionality in the chip design process, further increasing the cost and complexity during chip manufacturing or use, and even affecting the normal fluid operation of the microfluidic chip. Summary of the Invention

[0005] To solve the above problems, the object of the present invention is to provide a centrifugal microfluidic chip. It is composed of a three-way channel, a phase change material, and an energy storage device connected thereto, which can conveniently achieve the controllable batch release and precise quantitative release of the upstream liquid. Its structural composition is simple, which is convenient for reducing the production and manufacturing costs and simplifying the complexity of fluid controlled release.

[0006] The present invention adopts the following technical solutions:

[0007] A centrifugal microfluidic chip, comprising: a chip body; an upstream cavity located on the chip body; a downstream cavity located on the chip body, and the distal end side of the upstream cavity is communicated with the downstream cavity through a flow channel; a repeated switching valve disposed on the flow channel, with a phase change material inside, and by controlling heating and rotation speed, it can repeatedly block or connect the upstream cavity and the downstream cavity.

[0008] Further, the repeated switching valve includes: a three-way channel, the first channel of the three-way channel is communicated with the upstream cavity through a flow channel, the second channel of the three-way channel is communicated with the downstream cavity through a flow channel, and the distal end of the third channel of the three-way channel is communicated with the first channel and the second channel; an energy storage device, the energy storage device is arranged closer to the rotation center than the three-way channel, and the proximal end of the third channel is communicated with the side of the energy storage device away from the rotation center.

[0009] Further, the connection intersection of the first channel, the second channel and the third channel is located in the part of the three-way channel away from the rotation center.

[0010] Further, as the first channel and the second channel are away from the connection intersection, their inner diameters gradually increase.

[0011] Further, the first channel and the second channel are bent towards the rotation center with the connection intersection as the base point to form a bending area; when the repeated switching valve is closed, the position of the phase change material in the first channel and the second channel does not exceed the bending area; as the third channel is away from the connection intersection, its inner diameter gradually decreases.

[0012] Further, the energy storage device is of a cavity structure.

[0013] Further, the connections of the first channel, the second channel and the third channel are arc-connected; the minimum cross-sectional position of the first channel and the minimum cross-sectional position of the second channel form the smallest radius with the rotation center; the inner surface of the three-way channel in the area outside the connection of the two smallest radii is subjected to non-wetting treatment.

[0014] Furthermore, a buffer channel is provided between the proximal end of the third channel and the energy storage device.

[0015] Furthermore, the relationship between the minimum value of the minimum cross-sectional areas of the buffer channel and the third channel and the minimum cross-sectional areas of the first channel and the second channel satisfies the following formula:

[0016] ;

[0017] In the formula, S1 is the minimum cross-sectional area of the buffer channel, S2 is the minimum cross-sectional area of the third channel, S3 is the minimum cross-sectional area of the first channel, and S4 is the minimum cross-sectional area of the second channel.

[0018] Furthermore, when it is necessary to block the flow channel, heat the phase change material, control the rotation of the centrifugal microfluidic chip. At the first preset speed, the liquid phase change material moves from the proximal end to the distal end to seal the flow channel between the upstream cavity and the downstream cavity; then stop heating and wait for the liquid phase change material to solidify; when it is necessary to connect the upstream cavity and the downstream cavity, heat the solid phase change material. Under the action of negative pressure, the phase change material transformed from solid to liquid flows through the third channel towards the energy storage device. When the upstream cavity and the downstream cavity are connected, stop heating the phase change material and wait for the liquid phase change material to solidify; or, while heating the solid phase change material, control the rotation speed of the centrifugal microfluidic chip to ensure that the phase change material transformed from solid to liquid flows through the third channel towards the energy storage device at a certain flow rate. When the upstream cavity and the downstream cavity are connected, stop heating the phase change material and wait for the liquid phase change material to solidify.

[0019] The present invention provides a centrifugal microfluidic chip, which has the following beneficial technical effects compared with the prior art:

[0020] (1) First, by controlling two conditions of centrifugal force and heating, the repeated connection and closing of the repeated switching valve are realized. The sequential dropping of the reaction liquid ensures the normal progress of the reaction process. And by closing the repeated switching valve, it is ensured that the reaction in the downstream cavity is not affected by the liquid in the upstream cavity. The controllable batch release and accurate quantitative release of the upstream liquid make the detection result more accurate.

[0021] (2) Secondly, after the phase change material melts, the stable centrifugal force at a certain rotational speed and the constant gas pressure generated by the volume change are used to control the opening and closing of the valve, making the valve control method more efficient, stable and reliable; based on the balance relationship between the centrifugal force and the gas pressure, the repeated switching valve is repeatedly opened and closed, and then the batch release and precise quantitative release of the liquid reagent can be controlled, making the microfluidic chip design more functional and meeting various microfluidic control requirements; the repeated switching valve has a simple structure composition, an efficient and concise operation process, and extremely low requirements for operators and the hardware system, avoiding the complex components and operation processes of other active valve control methods. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following will briefly introduce the drawings required for the description of the embodiments. Obviously, the following drawings are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on these drawings.

[0023] Figure 1 Structural diagram of the centrifugal microfluidic chip provided in Embodiment 1 of the present invention;

[0024] Figure 2 Structural schematic diagram of the repeated switching valve in the open state provided in Embodiment 1 of the present invention;

[0025] Figure 3 Another structural schematic diagram of the repeated switching valve in the open state provided in Embodiment 1 of the present invention;

[0026] Figure 4 Structural schematic diagram of the repeated switching valve in the closed state provided in Embodiment 1 of the present invention;

[0027] Figure 5 Structural schematic diagram of the repeated switching valve in the closed state when the third channel has the same diameter provided in Embodiment 1 of the present invention;

[0028] Figure 6 Another structural schematic diagram of the repeated switching valve in the closed state provided in Embodiment 1 of the present invention;

[0029] Figure 7 Structural schematic diagram of the three-way structure provided in Embodiment 1 of the present invention;

[0030] Figure 8 Schematic diagram of the non-wetting treatment area of the three-way provided in Embodiment 1 of the present invention;

[0031] Figure 9 Structural schematic diagram of the repeated switching valve in the closed state with a buffer channel provided in Embodiment 1 of the present invention;

[0032] Figure 10 Structural diagram of a centrifugal microfluidic chip for in-situ synthesis of oligonucleotides provided in Embodiment 2 of the present invention.

[0033] Reference numerals: 1, chip body; 01, deprotecting agent storage chamber; 02, nucleotide monomer storage chamber; 020, adenine A storage chamber; 021, thymine T storage chamber; 022, cytosine C storage chamber; 023, guanine G storage chamber; 03, oxidant storage chamber; 04, capping agent storage chamber; 05, fluid channel; 06, oligonucleotide synthesis chamber; 07, waste liquid pool; 2, upstream cavity; 3, downstream cavity; 4, flow channel; 5, repeat switching valve; 50, phase change material; 51, three-way path; 510, first channel; 511, second channel; 512, third channel; 52, energy storage device; 53, buffer channel. Detailed implementation manners

[0034] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are some, but not all, of the embodiments of the present invention. Components of the embodiments of the present invention usually described and illustrated in the accompanying drawings here can be arranged and designed in various different configurations.

[0035] Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the claimed present invention, but merely represents selected embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts fall within the scope of protection of the present invention.

[0036] It should be noted that similar reference numerals and letters denote similar items in the following drawings. Therefore, once an item is defined in one drawing, it does not need to be further defined and explained in subsequent drawings.

[0037] In the description of the present invention, it should be noted that the orientation or positional relationship indicated by the terms "center", "upper", "lower", "left", "right", "vertical", "horizontal", "inner", "outer", etc. is based on the orientation or positional relationship shown in the accompanying drawings, or the orientation or positional relationship in which the product of the present invention is usually placed during use. It is only for the convenience of describing the present invention and simplifying the description, and does not indicate or imply that the device or element referred to must have a specific orientation, be constructed and operated in a specific orientation, and therefore cannot be construed as a limitation to the present invention. In addition, the terms "first", "second", "third", etc. are only used for descriptive distinction and cannot be construed as indicating or implying relative importance.

[0038] In the description of the present invention, it should also be noted that, unless otherwise clearly specified and defined, the terms "arranged", "installed", "connected" and "coupled" should be understood in a broad sense. For example, it may be a fixed connection, a detachable connection, or an integral connection; it may be a mechanical connection or an electrical connection; it may be a direct connection or an indirect connection through an intermediate medium, and it may be the communication inside two components. For those of ordinary skill in the art, the specific meanings of the above terms in the present invention can be understood according to specific situations.

[0039] Embodiment 1

[0040] An embodiment of the present invention discloses a centrifugal microfluidic chip, as Figure 1 and 2 shown, Figure 2 in which the black represents a solid-phase change material, including: a chip body 1; an upstream cavity 2, the upstream cavity 2 is located on the chip body 1; a downstream cavity 3, the downstream cavity 3 is located on the chip body 1, and the far-center end side of the upstream cavity 2 is communicated with the downstream cavity 3 through a flow channel 4; a repetitive switching valve 5, the repetitive switching valve 5 is arranged on the flow channel 4, and a phase change material 50 is arranged inside, and by controlling heating and rotation speed, the upstream cavity 2 and the downstream cavity 3 can be blocked or communicated repeatedly.

[0041] An installation hole is arranged on the chip body 1 for connecting with a device, and under the drive of the device, the chip body 1 is driven to rotate. The upstream cavity 2 and the downstream cavity 3 are in an upstream-downstream relationship, and the liquid in the upstream cavity can be controlled by the repetitive switching valve 5 to be dropped into the downstream cavity 3 multiple times to complete related reactions. The upstream cavity 2 and the downstream cavity 3 can be any step in the steps required for a biochemical reaction, and are used for the transfer or reaction of liquid. The upstream cavity 2 and the downstream cavity 3 can be provided with corresponding reagents or magnetic beads for accelerating the reaction inside according to specific reaction conditions, which will not be elaborated here.

[0042] The upstream cavity 2 and the downstream cavity 3 are in an upstream-downstream relationship in the process steps. The upstream cavity 2 and the downstream cavity can be located on the same radius or different radii centered on the rotation center O, as Figure 1 shown. In this embodiment, the distance between the downstream cavity 3 and the rotation center O is greater than the distance between the upstream cavity 2 and the rotation center O.

[0043] In the present invention, the "distal end" and "proximal end" are defined relative to the rotation center O. The phase change material 50 in the present invention can be wax. For example, the wax can be paraffin wax, microcrystalline wax, synthetic wax or natural wax. Alternatively, the phase change material can also be a gel or a thermoplastic resin. The gel can be polyacrylamide, polyacrylate, polymethacrylate, or polyvinylamine. The thermoplastic resin can be cyclic olefin copolymer (COC), polymethyl methacrylate (PMMA), polycarbonate (PC), polystyrene (PS), polyoxymethylene (POM), perfluoroalkoxy (PFA), polyvinyl chloride (PVC), polypropylene (PP), polyethylene terephthalate (PET), polyetheretherketone (PEEK), polyacrylate (PA), polysulfone (PSU) or polyvinylidene fluoride (PVDF), etc.

[0044] For the centrifugal microfluidic chip provided by the present invention, firstly, by controlling two conditions of centrifugal force and heating, the repeated connection and closing of the repeated switching valve 5 are realized, and the sequential dropping of the reaction solution ensures the normal progress of the reaction process. Moreover, by closing the repeated switching valve 5, it is ensured that the reaction in the downstream cavity 3 will not be affected by the liquid in the upstream cavity 2. The controllable batch release and precise quantitative release of the upstream liquid make the detection result more accurate. Secondly, when the phase change material melts, the stable centrifugal force at a certain rotational speed and the constant gas pressure generated by the volume change are used to control the opening and closing of the valve, making the valve control method more efficient, stable and reliable; based on the balance relationship between the centrifugal force and the gas pressure, the repeated switching valve 5 is switched on and off multiple times, and then the batch release and precise quantitative release of the liquid reagent can be controlled, making the microfluidic chip design more functional and meeting various microfluidic control requirements; the structure of the repeated switching valve is simple, the operation process is efficient and concise, and the requirements for the operator and the hardware system are extremely low, avoiding the complex components and operation processes of other active valve control methods.

[0045] It can be understood that the present application does not limit the heating method, as long as the heat source cooperates with the phase change material to realize the conversion of the phase change material into a liquid state under heating conditions.

[0046] Specifically, such as Figure 1 and 2As shown, the repeat switching valve 5 includes: a three-way passage 51. The first passage 510 of the three-way passage 51 is communicated with the upstream cavity 2 through a flow passage 4. The second passage 511 of the three-way passage 51 is communicated with the downstream cavity 3 through the flow passage 4. The distal end of the third passage 512 of the three-way passage 51 is communicated with the first passage 510 and the second passage 511. An energy storage device 52 is arranged closer to the rotation center O than the three-way passage 51. The proximal end of the third passage 512 is communicated with one side of the energy storage device 52 away from the rotation center. By arranging the energy storage device 52 closer to the proximal end compared with the third passage 512, it can meet the requirement of blocking or opening the connection between the upstream cavity 2 and the downstream cavity 3 while reducing the probability of bubbles appearing in the phase change material. Specifically, the energy storage device stores energy in the form of negative pressure, and the negative pressure acts on the surface of the liquid phase change material. The gas inside the liquid phase change material escapes from the phase change material under the negative pressure, preventing the gas fusion in the phase change material, so that the phase change material is divided into multiple segments by bubbles, seriously affecting the sealing effect.

[0047] As another embodiment of the present invention, as Figure 3 shown, the first passage 510 and the second passage 511 in the three-way passage 51 extend in a direction away from the rotation center O, and the blocking and connection of the upstream cavity 2 and the downstream cavity 3 can also be realized. It should be noted that in the embodiment of the present invention, the extension directions of the first passage 510 and the second passage 511 are not specifically limited, as long as the repeat switching valve 5 can realize the blocking and connection of the flow passage 4. Secondly, the first passage 510 and the second passage 511 can be symmetrically arranged or asymmetrically arranged, and symmetric arrangement is preferred.

[0048] As a preferred embodiment, as Figure 2 shown, the connection intersection S of the first passage 510, the second passage 511 and the third passage 512 is located in the part of the three-way passage away from the rotation center O. This structure can further improve the sealing effect of the phase change material on the passage. As Figure 4As shown, the light gray represents the liquid phase change material. When the phase change material is heated to melt and the chip body 1 rotates, the centrifugal force drives the liquid phase change material 50 to move away from the rotation center O. At this time, the liquid phase change material is deposited near the connection intersection S. Since the liquid phase change material flows away from the rotation center O, a negative pressure is generated in the energy storage device 52, and the distance between the liquid level in the third channel 512 and the rotation center is smaller than the distances between the liquid levels in the first channel 510 and the second channel 511 and the rotation center. The liquid phase change material deposited at the connection intersection S is driven by the centrifugal force to move the gas inside it towards the liquid level direction, that is, to move towards the rotation center direction in the channel, preventing the aggregation of gas inside the phase change material. Secondly, the centrifugal force has a compaction effect on the phase change material. The volume increased by the phase change material after the volume of the liquid phase change material solidifying and shrinking is offset by the volume removed by the centrifugal force, preventing the liquid phase change material from solidifying and shrinking and causing ineffective sealing of the first channel 510 and the second channel 511.

[0049] Specifically, as Figure 4 shown, the inner diameters of the first channel 510 and the second channel 511 gradually increase as they move away from the connection intersection S. As the liquid phase change material solidifies, a relatively large contraction length will be generated in the relatively long axial direction of the first channel 510 and the second channel 511, while the radial contraction is small. By setting the inner diameters of the first channel 510 and the second channel 511 to gradually increase as they move away from the connection intersection S, it is convenient for the phase change material with larger inner diameters at both ends after solidifying and shrinking to form tensile stress and be in close contact with the inner walls of the first channel 510 and the second channel 511, forming a good seal.

[0050] For the convenience of processing, in the embodiments of the present invention, the cross-sections of the flow channel 4, the first channel 510, the second channel 511, and the third channel 512 are preferably circular, and can also be other shapes.

[0051] The first channel 510 and the second channel 511 are bent towards the rotation center O with the connection intersection S as the base point, forming a bent area; when the repeated switching valve 5 is closed, the position of the phase change material 50 in the first channel 510 and the second channel 511 does not exceed the bent area; the inner diameter of the third channel 512 gradually decreases as it moves away from the connection intersection S. For the reasons mentioned above, when the first channel 510 and the second channel 511 are bent towards the rotation center O, as Figure 4As shown, during the solidification process, the gradually solidifying phase change material in the first channel 510 and the second channel 511 is compressed by the resultant force F towards the connection intersection S under the action of centrifugal force. The gradually solidifying phase change material with a variable diameter compacts the pipe walls of the first channel and the second channel under the action of pressure, improving the sealing effect. Secondly, when the third channel 512 is a constant-diameter channel, there is a possibility that, under the conditions of centrifugal force and solidification shrinkage of the liquid phase change material, there are still some partially solidified phase change materials on both sides of the first channel 510 and the second channel 511 that cannot compensate for the pores generated by the central solidification under the constraint of the inner wall of the variable-diameter channel, easily causing negative pressure cracks in the core. The generation of cracks is extremely likely to absorb local small bubbles, which then fuse and grow larger, causing the phase change material to be separated by the bubbles, resulting in a poor sealing effect and a small number of repeated uses, such as Figure 5 As shown, to solve the above problems, the present invention proposes that the inner diameter of the third channel 512 gradually decreases as it moves away from the connection intersection S. On the one hand, on the side closer to the rotation center O, the phase change material inside the third channel 512 is prone to solidify, and on the side farther from the rotation center O, the phase change material inside the third channel 512 takes a longer time to solidify. The liquid phase change material at the lower part of the third channel 512 compensates for the cracks generated during the solidification shrinkage of the liquid phase change material in the first channel 510 and the second channel 511, improving the sealing effect of the liquid phase change material on the three-way passage 51.

[0052] It can be understood that the first channel 510 and the second channel 511 can be bent upward in an arc, such as Figure 1 、 2 and shown in Figure 4, or can be bent upward in a straight line, such as Figure 6 shown.

[0053] To improve the compensation effect of the liquid phase change material in the third channel 512 on the first channel 510 and the second channel 511, as Figure 7 shown, the relationship between the cross-sectional area at the connection of the third channel 512 with the first channel 510 and the second channel 511 and the minimum cross-sectional areas of the first channel 510 and the second channel 511 satisfies the following formula:

[0054] ;

[0055] where S3 is the minimum cross-sectional area of the first channel, S4 is the minimum cross-sectional area of the second channel, S5 is the cross-sectional area at the connection of the third channel with the first channel and the second channel, or S5 is the maximum cross-sectional area of the third channel. The above size limitation ensures the compensation effect.

[0056] Energy storage device 52. The energy storage device 52 of the present application is a negative pressure energy storage. Specifically, under the action of centrifugal force, when the liquid phase change material flows from the proximal end to the distal end, the negative pressure in the energy storage device 52 increases. In the present invention, the energy storage device is a cavity structure sealed by a phase change material. To improve the energy storage effect, the cavity structure has a certain volume. It should be noted that the volume of the cavity structure will affect the selection of the rotation speed range and the rotation speed control accuracy; when the volume of the cavity structure is small, a larger rotation speed is required to increase the centrifugal force to drive the liquid phase change material to flow from the proximal end to the distal end. The rotation speed selection range is large, but the requirement for rotation speed control accuracy is high. A small change in the rotation speed will also affect the position and state of the liquid phase change material in the channel, and the energy consumption is large; the larger the volume of the cavity structure, the smaller the energy consumption, but the rotation speed selection range is small and the control accuracy requirement is relatively low, and the position change of the phase change material in the channel is large; by setting the cavity structure to have a certain volume, under the condition of moderate energy consumption, the liquid phase change material can be controlled in the channel at a lower rotation speed. On the other hand, when the chip body is in a stationary state, the reserved space in the energy storage device 52 can be relatively small. Even after the repeated switching valve 5 is repeated many times and part of the gas leaks into the cavity in the energy storage device 52, it can still be used for a long time. Through measurement, when the liquid phase change material fills one-third of the volume of the energy storage device, more than 1000 repetitions can be achieved and the blocking and connecting effects are good.

[0057] The above technical solution can meet the repeated blocking and connection between the upstream cavity 2 and the downstream cavity 3, but there is also a problem. For some special reactions, the reaction cavity needs to be composed of reaction substances inside and no impurities can be mixed in. For example, inert phase change materials should not be mixed into the reaction system. In order to ensure that the liquid phase change material does not mix into the downstream cavity during the use of the repeated switching valve 5, the present invention proposes a corresponding technical solution.

[0058] As Figure 8 shown, the connections of the first channel 510, the second channel 511 and the third channel 512 are arc-connected; the minimum cross-sectional area S3 position of the first channel 510 and the minimum cross-sectional area S4 position of the second channel 511 form the minimum radii R1 and R2 with the rotation center O; the inner surfaces of the three-way channel in the area outside the connection line of the two minimum radii R1 and R2 are treated with non-wetting, as shown by the slanted area in the figure. During the actual use of the product, the liquid phase change material is prone to adhesion at the connection point S. The phase change material adhered to the bottom of the three-way 51 enters the downstream cavity 3 under the flushing of the liquid discharged from the upstream cavity 2, thus polluting the reaction system in the downstream cavity 3. By treating the lower surface with non-wetting, the internal cohesive force of the liquid phase change material with a certain viscosity is greater than its cohesive force with the inner wall of the channel, which is beneficial to sucking the liquid phase change material into the energy storage device 52 and the third channel 512 through negative pressure.

[0059] For non-wetting treatment, a coating that is non-wetting with the phase change material can be applied to the inner wall of the channel, or micro-channels can be provided on the inner wall of the channel. Due to the certain surface tension of the liquid phase change material, the existence of the surface tension makes the liquid phase change material unable to wet the inside of the micro-channels, thereby reducing the bonding area with the inner wall of the channel and making the bonding force between the liquid phase change material and the inner wall of the channel smaller.

[0060] It should be noted that non-wetting treatment is not performed in the three-way channel within the region connecting the two minimum radii R1 and R2. The main reason is that if micro-channels are provided on the inner wall of the channel, the sealing effect of the phase change material on the three-way will be damaged.

[0061] As Figure 9 shown, a buffer channel 53 is provided between the proximal end of the third channel 512 and the energy storage device 52. The above setting can further reduce the residual probability of the liquid phase change material at the connection intersection S. Specifically, after the surface of the channel is subjected to non-wetting treatment, the internal bonding force of the liquid phase change material with a certain viscosity is greater than its bonding force with the inner wall of the channel. However, if the negative pressure in the energy storage device 52 is relatively large, the flow rate of the liquid phase change material is too fast, resulting in the internal bonding force of the local liquid phase change material being less than its bonding force with the inner wall of the channel, so that part of the liquid phase change material remains on the surface of the channel. By providing a buffer channel, the flow rate of the liquid phase change material can be further reduced, and the probability of the phase change material adhering to the inner wall of the channel can be reduced.

[0062] To achieve the buffering effect, the buffer channel can be set to be longer to increase the friction between the liquid phase change material and the channel, or the pipe diameter can be limited to increase the movement resistance of the buffer channel.

[0063] Specifically, the relationship between the minimum value of the minimum cross-sectional area of the buffer channel 53 and the third channel 512 and the minimum cross-sectional area S3 of the first channel 510 and the minimum cross-sectional area S4 of the second channel 511 satisfies the following formula:

[0064] ;

[0065] In the formula, S1 is the minimum cross-sectional area of the buffer channel, S2 is the minimum cross-sectional area of the third channel, S3 is the minimum cross-sectional area of the first channel, and S4 is the minimum cross-sectional area of the second channel. Through the above size limitation, when the chip body 1 is in a stationary state, the suction of the negative pressure on the liquid phase change material will not cause the flow rate of the liquid phase change material to be too fast, resulting in the residual phase change material at the connection intersection S.

[0066] The operating principle of the centrifugal microfluidic chip proposed by the present invention is as follows:

[0067] (1)When it is necessary to block the flow channel, heat the phase change material, control the rotation of the centrifugal microfluidic chip. At a preset speed, the liquid phase change material moves from the proximal end to the distal end, sealing the flow channel between the upstream cavity and the downstream cavity; then stop heating and wait for the liquid phase change material to solidify.

[0068] It should be noted that heating the phase change material and controlling the rotation of the centrifugal microfluidic chip include two control states: a. Heat the phase change material, and after the phase change material changes from solid state to liquid state, then control the rotation of the centrifugal microfluidic chip; b. Heat the phase change material while controlling the rotation of the centrifugal microfluidic chip. When the phase change material changes from solid state to liquid state, the liquid phase change material moves towards the distal end. Both of the above two situations are included in the technical solutions protected by the present invention.

[0069] Secondly, in the technical solutions protected by the present application, "then stop heating and wait for the liquid phase change material to solidify" also includes two scenarios: then stop heating, and after the liquid phase change material solidifies, it is possible to control the centrifugal microfluidic chip to stop rotating or keep rotating continuously.

[0070] (2)When it is necessary to connect the upstream cavity 2 and the downstream cavity 3, heat the solid phase change material. Under the action of negative pressure, the phase change material that changes from solid state to liquid state flows through the third channel towards the energy storage device. At this time, the chip body 1 does not need to rotate, and the liquid phase change material is sucked under the action of negative pressure, thereby connecting the upstream cavity and the downstream cavity. When the upstream cavity and the downstream cavity are connected, stop heating the phase change material. After the liquid phase change material solidifies, it is possible to control the centrifugal microfluidic chip to stop rotating or keep rotating continuously; or, control the rotation speed of the centrifugal microfluidic chip while heating the solid phase change material to ensure that the phase change material that changes from solid state to liquid state flows through the third channel towards the energy storage device at a certain flow rate. When the upstream cavity and the downstream cavity are connected, stop heating the phase change material. After the liquid phase change material solidifies, it is possible to control the centrifugal microfluidic chip to stop rotating or keep rotating continuously.

[0071] Embodiment 2

[0072] Such as Figure 10As shown in the figure, the embodiment of the present invention uses the repeated switching valve 5 of Embodiment 1. The centrifugal microfluidic chip described in this solution is used for in-situ synthesis of oligonucleotides, and mainly consists of raw material / reagent storage chambers, related fluid channels, the repeated switching valve 5, and a waste liquid pool located on the chip body 1. The rest of the supporting functional components and the specific synthesis mechanism are not shown and described in detail here, which conforms to the conventional understanding of professionals in this field. Among them, the deprotectant storage chamber 01, the nucleotide monomer storage chamber 02, the oxidant storage chamber 03, and the capping agent storage chamber 04 are all located at the position closest to the rotation center O. The oligonucleotide synthesis chamber 06 is connected to the deprotectant storage chamber 01, the nucleotide monomer storage chamber 02, the oxidant storage chamber 03, and the capping agent storage chamber 04 through several fluid channels 05 respectively. The nucleotide monomer storage chamber 02 is actually composed of 4 sub-storage chambers such as the adenine A storage chamber 020, the thymine T storage chamber 021, the cytosine C storage chamber 022, and the guanine G storage chamber 023. Each sub-storage chamber is connected to the oligonucleotide synthesis chamber 06 through its own separate fluid channel 05. The oligonucleotide synthesis chamber 06 is located farther from the rotation center O than each storage chamber, and the distal end of the oligonucleotide synthesis chamber 06 is connected to the waste liquid pool 07 at a farther end through the fluid channel 05. The repeated switching valve 5 is provided on all the fluid channels 05.

[0073] In the specific implementation manner involved in this proposal, the in-situ synthesis of oligonucleotides with a specific sequence is mainly realized by the phosphoramidite chemistry method. The synthesis process mainly includes four basic links: deprotection reaction, coupling of active nucleotide monomers, oxidation stabilization reaction, and capping and blocking reaction. By repeatedly cycling through these four basic links, the in-situ synthesis of oligonucleotides with a specific sequence on the microfluidic chip can be achieved. The specific chemical reaction process and mechanism are not described in detail here, and only the fluid release process of the microfluidic chip and the corresponding fluid regulation mechanism are introduced in detail.

[0074] The following are the specific operation steps to achieve the in-situ synthesis of oligonucleotides with a specific sequence by regulating the repeated opening and closing of the structure of the repeated switching valve 5:

[0075] (1) Fix a series of first protected nucleotide molecules in the oligonucleotide synthesis chamber 06 in advance, and add the corresponding liquid reagents to all the liquid reagent storage chambers. At this time, all the repeated switching valves 5 are in a completely closed state, that is, the upstream chamber and the downstream chamber are completely sealed and isolated through the closed repeated switching valve 5;

[0076] (2) Continuously and locally heat only the area of the repeat switching valve 5 between the deprotectant storage chamber 01 and the oligonucleotide synthesis chamber 06, while the centrifugal microfluidic chip remains stationary or rotates at a low speed. Stop heating after the melted phase change material enters the energy storage device 52. After the phase change material cools and solidifies, the opening of the repeat switching valve 5 is completed.

[0077] (3) Rotate the centrifugal microfluidic chip at a certain speed to apply a centrifugal force, so that part of the deprotectant in the deprotectant storage chamber 01 is released into the oligonucleotide synthesis chamber 06.

[0078] (4) After part of the deprotectant enters the oligonucleotide synthesis chamber 06, continuously and locally heat only the area of the repeat switching valve 5 between the deprotectant storage chamber 01 and the oligonucleotide synthesis chamber 06 again. While the centrifugal microfluidic chip is at a preset speed, after the phase change material in the energy storage device melts and enters the three-way passage under the action of centrifugal force, gradually stop heating while maintaining the rotation speed. After the phase change material cools and solidifies, the repeat switching valve 5 is closed again.

[0079] (5) Nucleotide molecule deprotection reaction occurs in the oligonucleotide synthesis chamber 06.

[0080] (6) Continuously and locally heat only the area of the repeat switching valve 5 between the oligonucleotide synthesis chamber 06 and the waste liquid pool 07, while the centrifugal microfluidic chip remains stationary or rotates at a low speed. Stop heating after the melted phase change material enters the energy storage device 52 of the repeat switching valve 5. After the phase change material cools and solidifies, the opening of the repeat switching valve 5 is completed.

[0081] (7) Rotate the centrifugal microfluidic chip at a certain speed to apply a centrifugal force, so that the waste liquid after the nucleotide molecule deprotection reaction in the oligonucleotide synthesis chamber 06 is completely released into the waste liquid pool 07.

[0082] (8) After the deprotection reaction waste liquid completely enters the waste liquid pool 07, continuously and locally heat only the area of the repeat switching valve 5 between the oligonucleotide synthesis chamber 06 and the waste liquid pool 07 again. While the centrifugal microfluidic chip rotates at a high speed, after the phase change material in the energy storage device melts and enters the three-way passage under the action of centrifugal force, gradually stop heating while maintaining the rotation speed. After the phase change material cools and solidifies, the repeat switching valve 5 is closed again.

[0083] (9) At this time, the first link - the deprotection reaction link - in the four basic links of in-situ oligonucleotide synthesis has been completed. At this time, all the repeat switching valves 5 are in a completely closed state, and a series of first nucleotide molecules pre-fixed in the oligonucleotide synthesis chamber 06 have been deprotected.

[0084] (10) Repeat the above steps (2)-(8) to selectively open and close the nucleotide monomer storage chamber 02 in a timely manner, completing the second of the four basic steps in in-situ oligonucleotide synthesis - the active nucleotide monomer coupling step. At this time, all the repeated switching valves 5 are in a fully closed state, and the first required nucleotide molecule has been coupled in the oligonucleotide synthesis chamber 06;

[0085] (11) Repeat the above steps (2)-(8) to selectively open and close the oxidant storage chamber 03 in a timely manner, completing the third of the four basic steps in in-situ oligonucleotide synthesis - the oxidation stabilization reaction step. At this time, all the repeated switching valves 5 are in a fully closed state, and the stabilization treatment after the coupling of the first required nucleotide molecule has been completed in the oligonucleotide synthesis chamber 06;

[0086] (12) Repeat the above steps (2)-(8) to selectively open and close the capping agent storage chamber 04 in a timely manner, completing the fourth of the four basic steps in in-situ oligonucleotide synthesis - the capping reaction step. At this time, all the repeated switching valves 5 are in a fully closed state, and the capping treatment of the first nucleotide that has not been coupled with the required nucleotide molecule has been completed in the oligonucleotide synthesis chamber 06 to prevent the extension of the sequence with deletion mutations;

[0087] (13) Repeat the above four basic steps in in-situ oligonucleotide synthesis. By selectively releasing different types of active nucleotide monomers in the nucleotide monomer storage chamber 02, the in-situ synthesis of oligonucleotides with specific sequences is finally completed on the centrifugal microfluidic chip.

[0088] The above are only examples of the present application and are not used to limit the protection scope of the present application. For those skilled in the art, various changes and modifications can be made to the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A centrifugal microfluidic chip, characterized in that, Comprising: Chip body; Upstream cavity, which is located on the chip body; Downstream cavity, which is located on the chip body, and one side of the telecentric end of the upstream cavity is communicated with the downstream cavity through a flow channel; Repeated switching valve, which is arranged on the flow channel and internally provided with a phase change material. By controlling heating and rotation speed, the upstream cavity and the downstream cavity can be repeatedly blocked or communicated; When it is necessary to block the flow channel, heat the phase change material, control the rotation of the centrifugal microfluidic chip. At the first preset speed, the liquid phase change material moves from the centripetal end to the telecentric end to seal the flow channel between the upstream cavity and the downstream cavity; then stop heating and wait for the liquid phase change material to solidify; When it is necessary to communicate the upstream cavity and the downstream cavity, heat the solid phase change material. Under the action of negative pressure, the phase change material that changes from solid to liquid flows through the third channel towards the energy storage device. When the upstream cavity and the downstream cavity are communicated, stop heating the phase change material and wait for the liquid phase change material to solidify; or, while heating the solid phase change material, control the rotation speed of the centrifugal microfluidic chip to ensure that the phase change material that changes from solid to liquid flows through the third channel towards the energy storage device at a certain flow rate. When the upstream cavity and the downstream cavity are communicated, stop heating the phase change material and wait for the liquid phase change material to solidify.

2. The centrifugal microfluidic chip according to claim 1, characterized in that, The repeated switching valve includes: Three-way passage, the first channel of the three-way passage is communicated with the upstream cavity through a flow channel, the second channel of the three-way passage is communicated with the downstream cavity through a flow channel, and the telecentric end of the third channel of the three-way passage is communicated with the first channel and the second channel; Energy storage device, which is arranged closer to the rotation center than the three-way passage, and the centripetal end of the third channel is communicated with one side of the energy storage device away from the rotation center.

3. The centrifugal microfluidic chip according to claim 2, wherein The connection intersection of the first channel, the second channel and the third channel is located in the part of the three-way passage away from the rotation center.

4. The centrifugal microfluidic chip according to claim 3, wherein The inner diameters of the first channel and the second channel gradually increase as they are away from the connection intersection.

5. The centrifugal microfluidic chip according to claim 4, wherein The first channel and the second channel are bent towards the rotation center with the connection intersection as the base point to form a bent area; When the repeated switching valve is closed, the position of the phase change material in the first channel and the second channel does not exceed the bent area; The third channel, as it is away from the connection intersection, Its inner diameter gradually decreases.

6. The centrifugal microfluidic chip according to claim 2, wherein The energy storage device is a cavity structure.

7. The centrifugal microfluidic chip according to claim 5, wherein The connections of the first channel, the second channel and the third channel are arc-connected; The minimum cross-sectional position of the first channel and the minimum cross-sectional position of the second channel form the minimum radius with the rotation center; The inner surface of the three-channel passage in the region outside the connection line of the two minimum radii is subjected to a non-wetting treatment.

8. The centrifugal microfluidic chip according to claim 5, wherein A buffer channel is provided between the proximal end of the third channel and the energy storage device.

9. The centrifugal microfluidic chip according to claim 8, wherein The relationship between the minimum value of the minimum cross-sectional areas of the buffer channel and the third channel and the minimum cross-sectional areas of the first channel and the second channel satisfies the following formula: 0.05(S3 + S4) ≤ min{S1, S2} ≤ 0.1(S3 + S4); In the formula, S1 is the minimum cross-sectional area of the buffer channel, S2 is the minimum cross-sectional area of the third channel, S3 is the minimum cross-sectional area of the first channel, and S4 is the minimum cross-sectional area of the second channel.

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