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JP-7856749-B2 - Control device and method for producing microdroplets

JP7856749B2JP 7856749 B2JP7856749 B2JP 7856749B2JP-7856749-B2

Inventors

  • マー， リァン

Assignees

  • マキュラ バイオテクノロジー カンパニー リミテッド

Dates

Publication Date
20260511
Application Date
20220826
Priority Date
20210826

Claims (20)

  1. A control device for producing microdroplets, wherein the control device includes a means for producing microdroplets and a means for controlling fluid. The microdroplet production means includes an asymmetric vibrating microdroplet generation mechanism and a first dynamic positioning member, the asymmetric vibrating microdroplet generation mechanism includes a vibrating member and a sample dosing needle, and the first dynamic positioning member is fixedly connected to the asymmetric vibrating microdroplet generation mechanism. The first dynamic positioning member is configured to position the asymmetric vibrating microdroplet generation mechanism, The fluid control means includes a fluid drive device and a conduit, one end of the conduit is connected to the fluid drive device, and the other end of the conduit is connected to the asymmetric vibrating microdroplet generation mechanism, and the fluid drive device is configured to set the flow velocity in order to cause the sample addition needle to absorb and drain liquid through the conduit . The asymmetric vibrating microdroplet generation mechanism further includes a vibration mounting base, a connecting guide structure, and a drive controller, wherein the vibration mounting base has a connection port, a pipe fitting, and a sample dosing needle adapter, one end of the connection port is connected to the liquid supply conduit of the conduit via a pipe fitting, and the other end is connected to the sample dosing needle by the sample dosing needle adapter. The vibrating member includes a housing, a vibrator, and a vibrating output rod. The vibration output rod of the vibrating member is connected to the vibration mounting base via the connecting guide structure and configured to provide power to the vibration mounting base, and the drive controller is electrically connected to the vibrating member. The control device for producing microdroplets is characterized in that the asymmetric vibrating microdroplet generation mechanism drives the vibration mounting base via the drive controller to perform asymmetric reciprocating vibration or asymmetric reciprocating oscillation in a direction along the central axis of the vibration output rod, thereby causing a sample-adding needle filled with a first liquid to perform asymmetric reciprocating motion below the liquid surface of a second liquid, thereby generating microdroplets of the first liquid .
  2. The control device according to claim 1, characterized in that the asymmetric vibrating microdroplet generation mechanism generates one microdroplet within one motion period , and the one motion period is the path of the vibrating member that goes from a reflection point RP1, through an equilibrium point EP to a reflection point RP2, and then returns from the reflection point RP2, again going through the equilibrium point EP to the reflection point RP1.
  3. The control device according to claim 2, characterized in that the central axis of the sample dosing needle is perpendicular to the axis of the vibration output rod.
  4. The control device according to claim 3, characterized in that the sample-dosing needle generates one microdroplet within one period of asymmetric reciprocating vibration or asymmetric reciprocating oscillation of the vibration mounting base.
  5. The vibration frequency of the vibrating member is 10 to 1000 Hz . The control device according to claim 3, characterized in that the vibration amplitude of the vibrating member is 0.1 to 5 mm .
  6. The oscillation frequency of the aforementioned vibration mounting base is 10 to 1000 Hz . The control device according to claim 3, characterized in that the distance between the liquid discharge opening of the sample dosing needle and the axis of the vibration output rod is 10 to 100 mm , and the oscillation angle width of the vibration mounting base is 0.05 to 10 ° .
  7. The control device according to claim 3, characterized in that the vibration mounting base and the vibration output rod are connected by a coupling.
  8. The vibrating member further includes a position sensor, and the drive controller achieves closed-loop control of motion by collecting real-time position feedback signals from the position sensor. The position sensor is one of the following : a grid scale sensor, a capacitive position sensor, a resistive sensor, a current sensor, or a differential transformer sensor. The control device according to claim 3, characterized in that the closed-loop control of the asymmetric reciprocating motion is performed by the drive controller, which collects a real-time position feedback signal from the position sensor of the vibrating member using a position signal acquisition module, compares it in real time with the asymmetric reciprocating motion control program, and feeds it back to the position correction module to adjust the control parameters of the vibration drive circuit.
  9. The asymmetric vibrating microdroplet generation mechanism further includes a support and fixing base for fixing the vibrating member, The control device according to claim 3, wherein the asymmetric vibrating microdroplet generation mechanism further includes a pump tube clamp base for clamping the liquid supply conduit.
  10. The aforementioned connection ports are multiple, and the multiple connection ports are provided at equal intervals inside the vibration mounting base. The control device according to claim 3, characterized in that the number of connection ports is 1 to 96 .
  11. The control device according to claim 10, wherein the connection guide structure is a ball spline including a spline shaft and a spline sleeve, and both ends of the spline shaft are fixedly connected to the vibration output rod and the vibration mounting base, respectively .
  12. The control device according to claim 10, wherein the connecting guide structure includes a first bearing and a second bearing, one end of the vibration mounting base is connected to the vibration output rod through the first bearing, the other end of the vibration mounting base is connected to the second bearing, and the first bearing is a bearing having an axial locking edge.
  13. The control device according to claim 3, further comprising a sample dosing needle removal mechanism for automatically removing the sample dosing needle after microdroplets have been generated.
  14. The sample dosing needle has a conical tubular structure with open ends, one end being a liquid supply opening for tight insertion with the sample dosing needle adapter, and the other end being a liquid discharge opening for generating microdroplets, the inner diameter of the liquid discharge opening being 20 to 300 μm and the outer diameter being 150 to 600 μm. The control device according to claim 3, characterized in that the liquid storage volume range of the sample -adding needle is 5 to 500 μL .
  15. When the vibrating mounting base performs the asymmetric reciprocating motion, the motion of the liquid dispensing portion of the sample-adding needle has one equilibrium point and two reflection points at both ends of the equilibrium point, and the position-time curve of the motion is configured to be asymmetric on both sides of either reflection point. The control device according to claim 3, characterized in that the asymmetric waveform of the periodic motion of the liquid discharge part of the sample dosing needle is at least one asymmetric combination of a sine wave, sawtooth wave, trapezoidal wave, triangular wave, and square wave.
  16. The control device according to claim 3, characterized in that the vibrating member is configured as a mechanism that generates continuous or intermittent motion, and the vibrating member is one selected from an electromagnetic vibration device, a piezoelectric ceramic vibration device, an eccentric wheel vibration device, a servo motor, a voice coil motor, and a galvanometer motor.
  17. The first dynamic positioning member includes a positioning member lifting displacement mechanism for controlling the lifting and lowering of the asymmetric vibrating microdroplet generation mechanism. The control device according to claim 1, wherein the first dynamic positioning member further includes a liquid level detection mechanism for assisting the first dynamic positioning member in accurately positioning the opening of the sample dosing needle.
  18. The control device further includes a second dynamic positioning member, the second dynamic positioning member used to fix and move the first opening container and the second opening container. The control device according to claim 17 , wherein the drive controller provides power to the fluid drive device, the first dynamic positioning member and the second dynamic positioning member, the first open container contains a first liquid, the sample addition needle draws in the first liquid to generate microdroplets of the first liquid, the second open container contains a second liquid, the sample addition needle performs asymmetric reciprocating vibration or asymmetric oscillation below the liquid surface of the second liquid to discharge the first liquid from the sample addition needle to generate microdroplets of the first liquid .
  19. The first open container is a single liquid storage tank, a one-dimensional liquid storage tank array, or a two-dimensional liquid storage tank array, and the volume of each liquid storage tank is 10 to 1000 μL . The control device according to claim 18, wherein the second opening container is a two-dimensional flat-bottom sample cell array for stacking the generated microdroplets , and the second opening container contains 24, 32, 96, or 384 flat-bottom sample cells of equal volume.
  20. The control device according to claim 19 , characterized in that the fluid drive device is a pulsation-free drive pump , and there is one or more fluid drive devices.

Description

This application relates to the technical field of microdroplet production, and more specifically, to a control device for microdroplet production and a method for producing microdroplets using the control device. Microdroplets are widely applied in various fields, and microfluidic control technologies based on microdroplets are experiencing rapid development and application in areas such as digital PCR, single-cell culture, single-cell genome/transcriptome sequencing, single-cell functional sorting, high-throughput screening, and protein crystallization. Microdroplet formation involves creating emulsified microdroplets using two immiscible phases; the microdroplet phase is called the dispersed phase, and the phase surrounding the microdroplet is called the continuous phase. After microdroplets are formed, operations such as splitting, fusion, mixing, dilution, collection, and sorting can be performed. Therefore, controlling the shape, size, and monodispersity of the microdroplets is crucial. Conventional microdroplet generation techniques can be broadly categorized into three types. The first involves generating microdroplets using a microfluidic chip, based on the principle of interfacial instability during the convergence of a dispersed phase and a continuous phase in a microchannel. The inventors found that generating microdroplets in such a microfluidic chip requires satisfying specific conditions such as flow velocity, oil-water interfacial tension, channel arrangement, and channel surface modification, and that the range of microdroplet volume control is also constrained by these factors. Furthermore, after the microdroplets are generated within the microfluidic chip's channels, they must be transferred to a storage container through specific steps and equipment, making it difficult to customize the conditions for a single microdroplet, and resulting in inconvenient operations such as microdroplet positioning, extraction, and analysis. The second method involves using a special device to spray a small amount of liquid to form microdroplets. For example, special spraying or microdroplet excitation methods such as piezoelectric ceramics, thermally excited expansion, and high-pressure spraying are employed. The inventors found that precisely controlling the volume of microdroplets using these methods is difficult, and that the corresponding fluid control system is complex. The third method involves injecting a small amount of liquid into a continuous phase via a micropipe, vibrating the outlet of the micropipe up and down at the gas-liquid interface of the continuous phase, and utilizing the cutting action of the surface tension at the phase interface to generate microdroplets of uniform size (Micropipe-based droplet generation method, Du Wenbin et al., Chinese Patent No.: ZL201410655191.5). This method makes it possible to produce microdroplets of uniform size and controllable volume. The inventors found that because the microdroplet samples produced by this method need to be stored in a microsyringe connected to the micropipe via a conduit, it is necessary to change the micropipe, conduit, and microsyringe when producing microdroplets for multiple samples, increasing the complexity of the operation. Furthermore, they found that this fluid control system makes it difficult to produce microdroplets for minute volume samples. Furthermore, once droplets are generated, the micropipe needs to be precisely positioned on the surface of the second liquid. However, inherent positioning errors and dynamic changes in the liquid surface during the droplet generation process can cause the micropipe to shift or gradually deviate from its optimal positioning height, making it impossible to consistently and continuously generate uniform droplets. This is a schematic diagram of the structure of a control device for producing microdroplets provided in this application.This is a schematic diagram of the three-dimensional structure of a microdroplet generation mechanism that vibrates asymmetrically along the central axis direction of the vibration output rod provided in this application, and is an enlarged view of reference numeral 1 in Figure 1.This is a schematic cross-sectional view of a microdroplet generation mechanism that vibrates asymmetrically along the central axis direction of the vibration output rod provided in this application, and is an enlarged view of reference numeral 1 in Figure 1.This is a schematic diagram of the three-dimensional structure of the asymmetrically oscillating microdroplet generation mechanism provided by the present invention, and is an enlarged view of reference numeral 1 in Figure 1.This is a front view of the asymmetrically oscillating microdroplet generation mechanism provided by the present invention, and is an enlarged view of reference numeral 1 in Figure 1.This is a schematic cross-sectional view of the asymmetrically oscillating microdroplet generation mechanism provided by the