A microfluidic chip for centrifugal-driven droplet generation and spreading
By setting up a clock microchannel in the microfluidic chip to connect the aqueous phase and the oil phase chamber, and emulsifying it with light oil, the high cost and additional operation problems caused by the use of heavy oil in the prior art are solved, and the low-cost, dead volume generation is achieved.
Patent Information
- Application Number
- CN202211679852.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2022-12-26
- Publication Date
- 2025-06-17
- Estimated Expiration
- 2042-12-26
AI Technical Summary
Existing microfluidic chips that drive step emulsification to generate droplets require the use of denser heavy oil, which makes oil-phase active agents expensive and requires additional operation to ensure that the water phase has no residue, and at the same time, the production costs are high, limiting industrial mass production.
By setting up a clock microchannel in the microfluidic chip to connect the aqueous phase chamber at the proximal end and the oil phase chamber at the distal end, the emulsification is carried out using light oil with lower price and lower density to reduce costs, and the structural design ensures that the emulsification is free of dead volume.
The use of light oil for droplet generation is realized, which reduces costs and ensures that the emulsification is free of dead volume without additional operations, simplifies the operation process and reduces sample loss.
Smart Images
Figure CN116060145B_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the field of microfluidic technology applications, and more specifically, relates to a microfluidic chip for centrifugal-driven droplet generation and spreading. Background Art
[0002] Traditional microfluidic droplet generation methods induce the mutual shearing of the aqueous phase and the oil phase through a specific channel structure to generate droplets, which have the advantages of stable droplet generation, high reproducibility, and good uniformity. However, multiple pumps are required to provide power, which makes the droplet generation inseparable from pre-adjustment, resulting in sample loss. Different from these methods, the step emulsification method generates droplets due to the "Rayleigh-Taylor" instability when the aqueous phase enters the oil phase through a step. This method avoids the need for multiple power sources while retaining the aforementioned advantages. Compared with traditional pump-driven microfluidic chips, centrifugal-driven microfluidic chips use centrifugal force as the power and can quickly reach the specified rotation speed under the action of a centrifuge without additional adjustment, thus not causing sample loss.
[0003] Existing microfluidic chips for centrifugal-driven step emulsification to generate droplets require the aqueous phase to be emulsified in the proximal chamber, which is connected to the distal oil phase chamber through a "U"-shaped tube. The depth of the oil phase chamber is greater than that of the "U"-shaped tube, thus forming a step. Through this structure, the aqueous phase reaches the step through the "U"-shaped tube to generate droplets. When generating droplets in batches, in order to avoid the droplets colliding and squeezing around the nozzle, resulting in droplet fusion, this structure requires the use of heavier oil with a greater density. After the droplets are generated, they gather towards the proximal end away from the nozzle under the action of buoyancy. The existing method has the advantages of stable droplet generation and convenient imaging; however, it also has the disadvantages of expensive oil phase surfactants and the need for additional operations to ensure no residual aqueous phase.
[0004] In order to construct the step structure, existing methods use micro-milling technology to engrave polymer materials such as PMMA, or use soft lithography technology to engrave multiple layers of PDMS channels. These methods greatly increase the production cost of microfluidic chips and limit their industrial mass production. Summary of the Invention
[0005] Aiming at the defects and improvement requirements of the existing technology, the present invention provides a microfluidic chip for centrifugal-driven droplet generation and spreading, aiming to reduce the cost of droplet generation and ensure no dead volume in emulsification without additional operations.
[0006] To achieve the above object, according to one aspect of the present invention, there is provided a microfluidic chip for centrifugal-driven droplet generation and spreading, including:
[0007] A substrate;
[0008] An intermediate layer bonded to the substrate; one or more droplet generation units are provided on the intermediate layer, and the droplet generation unit includes: an oil phase pre-storage cavity, an aqueous phase pre-storage cavity, and a droplet generation cavity arranged in sequence from the inside out, a microchannel chip reserved area arranged between the aqueous phase pre-storage cavity and the droplet generation cavity, a waste liquid cavity, a first channel for connecting the oil phase pre-storage cavity and the droplet generation cavity, and a second channel for connecting the droplet generation cavity and the waste liquid cavity;
[0009] A microchannel chip placed in the microchannel chip reserved area, which is irreversibly bonded to the substrate, and a rectangular microchannel for connecting the aqueous phase pre-storage cavity and the droplet generation cavity is provided at the bottom; the depth of the rectangular microchannel is less than that of the droplet generation cavity;
[0010] An upper cover covering the intermediate layer and the microchannel chip; the upper cover includes: a sealing area connected to the microchannel chip reserved area, an aqueous phase sampling hole communicated with the aqueous phase pre-storage cavity, an oil phase sampling hole communicated with the oil phase pre-storage cavity, and an air hole communicated with the waste liquid cavity;
[0011] And a sealant filled in the gaps between the substrate, the intermediate layer, the microchannel chip and the upper cover;
[0012] The substrate, the intermediate layer and the upper cover are all disc-shaped, and positioning holes aligned with each other are provided at the center.
[0013] Further, the horizontal width of the proximal end of the aqueous phase pre-storage cavity is larger than that of the distal end, and the width of the distal end gradually decreases; the two sides of the distal end connected to the proximal end are two arcs of the same circle.
[0014] Further, the bottom of the oil phase pre-storage cavity is arc-shaped.
[0015] Further, the bottom of the droplet generation cavity is arc-shaped.
[0016] Further, two waste liquid cavities are provided, and two second channels are correspondingly provided;
[0017] The two waste liquid cavities are respectively located on both sides of the microfluidic chip reserved area, and the proximal end curved surfaces of the two waste liquid cavities are two arcs of the same circle.
[0018] In some alternative embodiments, the distal end curved surface of the droplet generation cavity is inclined from bottom to top towards the proximal end curved surface, and the droplet generation unit further includes: a droplet flattening cavity communicated with the droplet generation cavity and arranged above the droplet generation cavity.
[0019] In some alternative embodiments, the droplet generation unit further includes: a droplet flattening cavity arranged outside the droplet generation cavity and connected to the droplet generation cavity, and an active valve arranged between the liquid generation cavity and the liquid flattening cavity.
[0020] Further, the upper substrate is made of quartz glass, the upper microchannel chip is made of PDMS, and the upper intermediate layer is made of PMMA.
[0021] Further, the thickness of the droplet generation chamber is 2 to 3.5 times the diameter of the generated droplet.
[0022] According to another aspect of the present invention, there is provided another microfluidic chip for centrifugal-driven droplet generation and spreading, comprising: a centrifugal locator, and one or more cassette chips;
[0023] The centrifugal locator is provided with positioning holes and a card slot for loading the cassette chip;
[0024] The cassette chip includes:
[0025] A substrate;
[0026] An intermediate layer adhered to the substrate; the intermediate layer includes a droplet generation unit, and the droplet generation unit includes: an oil phase pre-storage chamber, an aqueous phase pre-storage chamber, and a droplet generation chamber arranged in sequence, a microchannel chip reserved area arranged between the aqueous phase pre-storage chamber and the droplet generation chamber, a waste liquid chamber, and a first channel for communicating the oil phase pre-storage chamber and the droplet generation chamber and a second channel for communicating the droplet generation chamber and the waste liquid chamber;
[0027] A microchannel chip placed in the microchannel chip reserved area, which is irreversibly bonded to the substrate, and a rectangular microchannel for communicating the aqueous phase pre-storage chamber and the droplet generation chamber is arranged at the bottom; the depth of the rectangular microchannel is less than that of the droplet generation chamber;
[0028] An upper cover covering the intermediate layer and the microchannel chip; the upper cover includes: a sealing area connected to the microchannel chip reserved area, an aqueous phase sampling hole communicating with the aqueous phase pre-storage chamber, an oil phase sampling hole communicating with the oil phase pre-storage chamber, and an air hole communicating with the waste liquid chamber;
[0029] And a sealant filled in the gaps between the substrate, the intermediate layer, the microchannel chip and the upper cover;
[0030] During emulsification, the oil phase pre-storage chamber is located at the proximal end, and the droplet generation chamber is located at the distal end.
[0031] Further, the horizontal width of the proximal end of the aqueous phase pre-storage chamber is larger than that of the distal end, and the width of the distal end gradually decreases; the two sides of the distal end connected to the proximal end are two arcs of the same circle.
[0032] Further, the bottom of the oil phase pre-storage chamber is arc-shaped.
[0033] Further, the bottom of the droplet generation chamber is arc-shaped.
[0034] Further, there are two waste liquid chambers, and two second channels are correspondingly arranged;
[0035] The two waste liquid chambers are respectively located on both sides of the reserved area of the microfluidic chip, and the curved surfaces of the proximal ends of the two waste liquid chambers are two arcs of the same circle.
[0036] In some alternative embodiments, the curved surface of the distal end of the droplet generation chamber inclines from bottom to top towards the curved surface of the proximal end, and the droplet generation unit further includes: a droplet flattening chamber that communicates with the droplet generation chamber and is disposed above the droplet generation chamber.
[0037] In some alternative embodiments, the droplet generation unit further includes: a droplet flattening chamber that is disposed outside the droplet generation chamber and connected to the droplet generation chamber, and an active valve that is disposed between the liquid generation chamber and the liquid flattening chamber.
[0038] Further, the upper substrate is made of quartz glass, the upper microchannel chip is made of PDMS, and the upper intermediate layer is made of PMMA.
[0039] Further, the thickness of the droplet generation chamber is 2 to 3.5 times the diameter of the generated droplets.
[0040] Generally speaking, through the above technical solutions conceived by the present invention, the following beneficial effects can be achieved:
[0041] (1) The microfluidic chip for centrifugal-driven droplet generation and flattening provided by the present invention (including a disc-type chip and a cartridge-type chip), wherein the aqueous phase pre-storage chamber located at the proximal end and the droplet generation chamber located at the distal end are connected through a rectangular microchannel at the bottom of the microchannel, and the depth of the rectangular microchannel is less than the depth of the droplet generation chamber, so that a step is formed by the substrate, the rectangular microchannel and the droplet generation chamber. Based on this step structure, the present invention can directly use light oil with a smaller density to complete emulsification. On the one hand, the industrial production of light oil and its surfactants is mature and the price is low, which can further reduce costs. On the other hand, when using light oil, no additional operation is required to ensure that there is no dead volume in emulsification, thereby minimizing manual operation and ensuring no loss of samples.
[0042] (2) In the preferred embodiment of the microfluidic chip for centrifugal-driven droplet generation and flattening provided by the present invention, the horizontal width of the proximal end of the aqueous phase pre-storage chamber is larger than that of the distal end, and the width of the distal end gradually decreases; the two sides of the distal end connected to the proximal end are two arcs of the same circle. Through this structural design of the aqueous phase pre-storage chamber, after adding the aqueous phase to the aqueous phase pre-storage chamber and then adding an appropriate amount of oil phase, it can be ensured that under the action of centrifugation, the oil phase liquid level is at the part with a wider horizontal position at the proximal end. After the aqueous phase reaches the step through the rectangular microchannel to generate droplets by emulsification and is completely emulsified, the oil phase liquid level is still at the part with a wider horizontal position at the proximal end, thereby minimizing the change in pressure at the step during the droplet generation process and ensuring the stability of droplet generation and the generation of the aqueous phase without dead volume.
[0043] (3) In the preferred embodiment of the centrifugal-driven droplet generation and spreading microfluidic chip provided by the present invention, the bottom of the oil-phase pre-storage chamber is arc-shaped, so that the oil phase in the oil-phase pre-storage chamber can enter the droplet generation chamber without dead volume under the action of centrifugation.
[0044] (4) In the preferred embodiment of the centrifugal-driven droplet generation and spreading microfluidic chip provided by the present invention, the bottom of the droplet generation chamber is arc-shaped, so that the droplets in the droplet generation chamber are uniformly stressed during centrifugation, which is beneficial to ensuring the stability of droplet generation.
[0045] (5) In the preferred embodiment of the centrifugal-driven droplet generation and spreading microfluidic chip provided by the present invention, the proximal curved surfaces of the two waste liquid chambers are two arcs of the same circle, so that the pressure in the waste liquid chamber can be kept constant, thereby ensuring the stability of droplet generation.
[0046] (6) In the preferred embodiment of the centrifugal-driven droplet generation and spreading microfluidic chip provided by the present invention, the distal curved surface of the droplet generation chamber inclines from bottom to top towards the proximal curved surface, and a droplet spreading chamber is separately provided. Based on this structure, after the droplet generation is completed and the droplets are accumulated at the bottom of the droplet generation chamber, the entire microfluidic chip is horizontally flipped by 180° and centrifuged, so that the droplets can enter the droplet spreading chamber along the inclined distal curved surface of the droplet generation chamber and be arranged in a single layer to achieve spreading; through the separately provided droplet spreading chamber, the present invention can achieve active spreading, and the whole process has fewer operation steps, is more controllable, and is more automated.
[0047] (7) In the preferred embodiment of the centrifugal-driven droplet generation and spreading microfluidic chip provided by the present invention, a droplet spreading chamber connected to the droplet generation chamber is further provided outside the droplet generation chamber, and an active valve is provided between the two. Based on this structure, after the droplet generation is completed and the droplets are accumulated at the bottom of the droplet generation chamber, by dissolving the solidified soluble material in the active valve and centrifuging, the droplets can enter the droplet spreading chamber and be arranged in a single layer to achieve spreading; through the separately provided droplet spreading chamber, the present invention can achieve active spreading, and the whole process has fewer operation steps, is more controllable, and is more automated.
[0048] (8) In the preferred embodiment of the centrifugal-driven droplet generation and spreading microfluidic chip provided by the present invention, the step is composed of a combination of multiple materials. Specifically, the substrate, rectangular microchannel and droplet generation chamber that make up the step are made of quartz glass, PDMS and PMMA respectively, which simplifies the manufacturing process, further improves the processing speed, and further reduces the production cost.
[0049] (9) In the preferred embodiment of the microfluidic chip for centrifugal-driven droplet generation and tiling provided by the present invention, the thickness of the droplet generation chamber is 2 to 3.5 times the diameter of the generated droplet. Through this design, after the droplet is generated, the microfluidic chip is placed horizontally for 5 - 15 minutes, and the proximal end of the droplet can be arranged in a single layer, and the distal end can be arranged in a double layer, greatly simplifying the tiling operation. BRIEF DESCRIPTION OF THE DRAWINGS
[0050] Figure 1 FIG. 6 is a schematic structural diagram of the microfluidic chip for centrifugal-driven droplet generation in Embodiment 1 of the present invention;
[0051] Figure 2 FIG. 7 is a front view of the microfluidic chip for centrifugal-driven droplet generation in Embodiment 1 of the present invention;
[0052] Figure 3 FIG. 8 is an exploded view of the microfluidic chip for centrifugal-driven droplet generation in Embodiment 1 of the present invention;
[0053] Figure 4 FIG. 9 is a top view of the microfluidic chip for centrifugal-driven droplet generation in Embodiment 1 of the present invention;
[0054] Figure 5 FIG. 10 is a schematic diagram of the step structure in Embodiment 1 of the present invention;
[0055] Figure 6 FIG. 11 is a CFD simulation diagram of the step emulsification process in Embodiment 1 of the present invention;
[0056] Figure 7 FIG. 12 is a droplet generation diagram and its diameter frequency histogram in Embodiment 1 of the present invention;
[0057] Figure 8 FIG. 13 is a schematic structural diagram of the cartridge chip for centrifugal-driven droplet generation and tiling in Embodiment 2 of the present invention;
[0058] Figure 9 FIG. 14 is a schematic structural diagram of the centrifugal locator loaded with the cartridge chip in Embodiment 2 of the present invention;
[0059] Figure 10 FIG. 15 is a schematic diagram of the droplet tiling principle of the cartridge chip for centrifugal-driven droplet generation and high-throughput tiling in Embodiment 2 of the present invention;
[0060] Figure 11 FIG. 16 is a schematic structural diagram of the microfluidic chip for centrifugal-driven droplet generation and high-throughput tiling in Embodiment 3 of the present invention;
[0061] Figure 12 FIG. 17 is a schematic structural diagram of the main body of the microfluidic chip for centrifugal-driven droplet generation and high-throughput tiling in Embodiment 3 of the present invention;
[0062] Figure 13This is a schematic structural diagram of the active valve-assisted high-throughput droplet tiling in Embodiment 4 of the present invention.
[0063] In all the drawings, the same reference numerals are used to represent the same elements or structures, where:
[0064] 1 - Substrate; substrate positioning hole 11;
[0065] 2 - Intermediate layer 2; 211 - Oil phase pre-storage cavity, 212 - Aqueous phase pre-storage cavity, 213 - Droplet generation cavity, 214 - Waste liquid cavity, 215 - Droplet tiling cavity, 22 - Microchannel chip reserved area, 231 - First channel, 232 - Second channel, 24 - Intermediate layer positioning hole;
[0066] 3 - Microchannel chip; 31 - Cuboid microchannel;
[0067] 4 - Upper cover; 41 - Air hole, 421 - Oil phase sampling hole, 422 - Aqueous phase sampling hole, 43 - Sealing area, 44 - Upper cover positioning hole;
[0068] 5 - Sealing glue;
[0069] 6 - Active valve. Detailed implementation manners
[0070] In order to make the objectives, technical solutions and advantages of the present invention clearer and more understandable, the present invention will be further described in detail below with reference to the drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention. In addition, the technical features involved in the various embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.
[0071] In the present invention, the terms "first", "second", etc. (if any) in the present invention and the drawings are used to distinguish similar objects and do not necessarily need to describe a specific order or sequence.
[0072] Regarding the existing microfluidic chip for centrifugal-driven step emulsification to generate droplets, by setting the depth of the "U"-shaped tube connecting the proximal aqueous phase chamber and the distal oil phase chamber to be less than the step formed by the oil phase chamber, this connection structure requires the use of heavier oil with a greater density. Therefore, there are technical problems such as expensive oil phase surfactants and the need for additional operations to ensure no residue in the aqueous phase. The present invention provides a microfluidic chip for centrifugal-driven droplet generation and tiling. The overall idea is as follows: Improve the droplet generation structure and directly connect the proximal aqueous phase chamber and the distal oil phase chamber using a vertical channel, so that emulsification can be completed using lighter oil with a lower price and a smaller density, achieving the purpose of reducing costs and ensuring no dead volume in emulsification without additional operations.
[0073] The following are the embodiments.
[0074] Example 1:
[0075] A microfluidic chip for centrifugal-driven droplet generation and spreading, the structure of which is as Figures 1 to 4 shown, including: a substrate 1, an intermediate layer 2, a microchannel chip 3, an upper cover 4 and a sealant 5.
[0076] As Figure 1 and Figure 3 shown, the substrate 1 is disc-shaped, and a substrate positioning hole 11 is provided at its center; optionally, in this embodiment, the material of the substrate 1 is quartz glass and it is a hydrophilic substrate; it should be noted that the hydrophilic or hydrophobic state of the substrate 1 is related to the size of the generated droplets, and in actual applications, it can be set accordingly according to the droplet size. In some other embodiments of the present invention, the substrate 1 can also be in a hydrophobic state.
[0077] As Figure 1 、 Figure 3 and Figure 4 shown, in this embodiment, the intermediate layer 2 is disc-shaped, a middle layer positioning hole 24 is provided at its center, and 4 droplet generation units are symmetrically arranged around the disc center thereon. The droplet generation unit includes: an oil phase pre-storage cavity 211, an aqueous phase pre-storage cavity 212, and a droplet generation cavity 213 arranged in sequence from inside to outside, a microchannel chip reserved area 22 provided between the aqueous phase pre-storage cavity 212 and the droplet generation cavity 213, a waste liquid cavity 214, a first channel 231 for connecting the oil phase pre-storage cavity 211 and the droplet generation cavity 213, and a second channel 232 for connecting the droplet generation cavity 213 and the waste liquid cavity 214. Optionally, in this embodiment, the intermediate layer 2 is formed by bonding PMMA plates after laser engraving, and the thickness of the droplet generation cavity 213 is 0.5 mm.
[0078] The microchannel chip 3 is placed in the microchannel chip reserved area 22; optionally, in this embodiment, the microchannel chip 3 is a rectangular PDMS chip (10 mm in length, 5 mm in width, and about 2 mm in height), which is processed and formed by traditional soft lithography technology and then cut by a die cutter. Ten through rectangular microchannels 31 (5 mm in length, 50 μm in width, and 20 μm in height) are recessed at the bottom thereof, with a spacing of 300 μm; after the microchannel chip 3 is placed in the microchannel chip reserved area 22, it is bonded to the substrate 1 by oxygen plasma. As Figure 5 shown, since the depth of the rectangular microchannels 31 is less than the depth of the droplet generation cavity 213, a stepped structure is formed by the substrate 1, the rectangular microchannels 31 and the droplet generation cavity 213, and since the three parts forming the step are composed of a variety of materials with mature processing technologies, the manufacturing process is further simplified and the production cost is further reduced.
[0079] The upper cover 4 covers the intermediate layer 2 and the microchannel chip 3, as Figure 3As shown in the figure, the upper cover 4 includes: a sealing area 43 connected to the reserved area 22 of the microchannel chip, an aqueous-phase sampling hole 422 communicating with the aqueous-phase pre-storage cavity, an oil-phase sampling hole 421 communicating with the oil-phase pre-storage cavity 211, and an air hole 41 communicating with the waste liquid cavity 214; the upper cover 4 is also in a disc shape, and an upper cover positioning hole 44 is provided at its center. The substrate positioning hole 11, the intermediate layer positioning hole 24, and the upper cover positioning hole 44 coincide, and their centers are the centers of the centrifugation of the chip.
[0080] The sealant 5 is coated on the gaps between the reserved area 22 of the microchannel chip, the sealing area 43, the microchannel chip 3, and the upper cover 4; optionally, in this embodiment, the sealant 5 is a high-viscosity silicone sealant, which is left standing for one hour at room temperature and cured by combining with water molecules in the air, making the rectangular microchannel 31 the only channel connecting the aqueous-phase pre-storage cavity 212 and the droplet generation cavity 213, and isolating the air around the opening of the rectangular microchannel 31. As Figure 3 and Figure 4 shown in the figure, in order to facilitate the coating of the sealant 5, the shape of the reserved area 22 of the microchannel chip matches the shape of the microchannel chip 3 to ensure the accurate positioning and fixing of the microchannel chip 3, and after the microchannel chip 3 is placed, there is still a certain space left in the reserved area 22 of the microchannel chip; the position of the sealing area 43 on the upper cover 4 corresponds to the reserved area 22 of the microchannel chip one by one and has the same shape.
[0081] The process of droplet generation and spreading using the centrifugal-driven droplet generation and spreading microfluidic chip provided in this embodiment is as follows:
[0082] First, 200 μl of isopropyl palmitate (containing 7% w / w EM180) is added to the oil-phase pre-storage cavity 211 through the oil-phase sampling hole 421 and centrifuged at a speed of 3000 RPM. The oil phase can enter and fill the droplet generation cavity 213 through the first channel 231. The air originally in the droplet generation cavity 213 enters the waste liquid cavity 214 along the second channel 232 connected in the middle of the cavity and is discharged from the chip through the air hole 41; it is easy to understand that in order to ensure that the oil phase fills the droplet generation cavity 213, the volume of the oil-phase pre-storage cavity 211 is slightly larger than the volume of the droplet generation cavity 213. After stopping centrifugation, under capillary action, the oil phase wets the rectangular microchannel 31; in order to facilitate the wetting of the rectangular microchannel 31 by the oil phase and make the droplet generation more stable, in this embodiment, the rectangular microchannel 31 has been pre-treated with hydrophobic treatment.
[0083] Then, 10 μl of red pigment (aqueous phase) and 200 μl of oil phase are successively added to the aqueous-phase pre-storage cavity 212 through the aqueous-phase sampling hole 422, and centrifuged at 3000 RPM for 10 minutes. All the aqueous phase generates droplets through the steps. The CFD simulation time series during the generation process is as Figure 6As shown, where the first row is the top view of the droplet generation process and the second row is the side view of the droplet generation process; the generated droplets accumulate at the bottom of the droplet generation chamber 213, and the surfactant in the oil phase protects the droplets from fusing after collision and extrusion. Since the thickness of the droplet generation chamber 213 is greater than the diameter of the generated droplets, the droplets are arranged in multiple layers.
[0084] Next, tilt the chip by 10 - 20°, so that the bottom end of the droplet generation chamber 213 is in the upper position in space and the centrifugal center is in the lower position in space. After standing for 10 minutes, the droplets are arranged in a single layer. Obtain the stitched image of the two-dimensional droplet array through a confocal microscope, as Figure 7 shown in the left image. The results show that the droplet spreading function has a good effect and will not cause droplet fusion; further obtain the droplet diameter through the image processing toolbox in the MATLAB software and draw the diameter frequency histogram, as Figure 7 shown in the right image. The results show that the average value of the droplet diameter is about 142 μm, and the coefficient of variation is about 3.1%, with good uniformity.
[0085] In this embodiment, the droplet spreading is realized by tilting the chip. It should be noted that this is only an optional embodiment of the present invention and should not be construed as the only limitation of the present invention. In some other embodiments of the present invention, the process of droplet spreading can also be simplified by specially designing the size of the droplet generation chamber 213. Specifically, the thickness of the droplet generation chamber 213 can be set to 2 - 3.5 times the diameter of the generated droplets. Through this design, after the droplets are generated, place the microfluidic chip horizontally for 5 - 15 minutes, and the proximal ends of the droplets can be arranged in a single layer, and the distal ends can be arranged in a double layer, greatly simplifying the spreading operation.
[0086] Refer to Figure 3 and Figure 4 For further optimizing the effect of droplet generation, this embodiment further improves the chip structure. Specifically as follows:
[0087] Refer to Figure 3 and Figure 4 In this embodiment, the proximal end horizontal width of the aqueous phase pre-storage chamber 212 is larger than that of the distal end, and the width of the distal end gradually decreases; the two sides of the distal end connected to the proximal end are two arcs of the same circle; through this structural design, after adding the aqueous phase to the aqueous phase pre-storage chamber 212 and then adding an appropriate amount of oil phase, it can make the oil phase liquid level in the position with a wider horizontal width at the proximal end under the centrifugal action. After the aqueous phase reaches the step through the rectangular microchannel 31 to generate droplets and is completely emulsified, the oil phase liquid level is still in the position with a wider horizontal width at the proximal end, thereby minimizing the change in pressure at the step during the droplet generation process and ensuring the stability of droplet generation and the generation of the aqueous phase without dead volume.
[0088] Refer to Figure 3 andFigure 4 , in this embodiment, the bottom of the oil-phase pre-storage chamber 211 is arc-shaped, so that the oil phase in the oil-phase pre-storage chamber 211 can enter the droplet generation chamber 213 without dead volume under the action of centrifugation; the bottom of the droplet generation chamber 213 is also arc-shaped, so that the droplets in the droplet generation chamber 213 are uniformly stressed during centrifugation, which is beneficial to ensuring the stability of droplet generation.
[0089] Refer to Figure 3 and Figure 4 , in this embodiment, there are two waste liquid chambers 214, and correspondingly two second channels 232 are also provided, which are respectively used to connect the droplet generation chamber 213 and the two waste liquid chambers 214. And in order to facilitate the exclusion of gas during centrifugation, the second channels 232 are as close as possible to the microchannel chip 3; the two waste liquid chambers 214 are respectively located on both sides of the microchannel chip reserved area 22, and the curved surfaces of the proximal ends of the two waste liquid chambers 214 are two arcs of the same circle, so that the pressure in the waste liquid chamber 214 can be kept constant, thereby ensuring the stability of droplet generation.
[0090] Refer to Figure 3 and Figure 4 , in this embodiment, in order to accelerate the speed at which the oil phase in the oil-phase pre-storage chamber 211 enters the droplet generation chamber 213 under the action of centrifugation, a pair of first channels 231 are also provided, and are respectively arranged on both sides of the bottom end of the oil-phase pre-storage chamber 211.
[0091] In this embodiment, the multiple droplet generation units in the chip can respectively adopt the rectangular microchannels 31 of different sizes, and are centrifuged at the same rotation speed. Multiple droplet generation units in the same chip can respectively generate uniform droplets with different diameters. Similarly, it should be noted that in this embodiment, the number of droplet generation units, the sizes of each chamber, etc. can all be flexibly adjusted according to actual needs.
[0092] Embodiment 2:
[0093] A microfluidic chip for centrifugal-driven droplet generation and spreading, different from the disc-type chip in Embodiment 1, this embodiment specifically includes a centrifugal locator and one or more cartridge chips;
[0094] The centrifugal locator is provided with positioning holes and a slot for loading the cartridge chip;
[0095] The structure of a single cartridge chip is as Figure 8 shown. Similar to Embodiment 1, in this embodiment, a single cartridge chip also includes a substrate, an intermediate layer, a microchannel chip, an upper cover and a sealant; among them, the intermediate layer only includes a single droplet generation unit, and the structure of this droplet generation unit is the same as that of the droplet generation unit in Embodiment 1 above. Specifically, it can refer to the description in Embodiment 1 above and will not be repeated here.
[0096] During centrifugation, Figure 8 the cassette chip shown needs to be loaded into a centrifugal locator for centrifugation. During emulsification, the oil phase pre-storage chamber is located at the proximal end, and the droplet generation chamber is located at the distal end. As Figure 9 shown, the centrifugal locator can load four cassette chips.
[0097] The process of droplet generation and spreading using the microfluidic chip for centrifugal-driven droplet generation and spreading provided in this embodiment is as follows:
[0098] As Figure 10 shown, after the centrifugal locator loads four cassette chips (labeled "1", "2", "3", and "4" respectively), samples are added for centrifugation to generate droplets according to the sample addition method in the above-mentioned Embodiment 1. The generated droplets will accumulate at the outermost end of the droplet generation chamber. Subsequently, each cassette chip is horizontally rotated 180° and re-loaded so that the end face with accumulated droplets is located at the innermost side. After centrifugation at a speed of 60 RPM for 10 minutes, the droplets in the four droplet generation units are all arranged in a single layer.
[0099] Embodiment 3:
[0100] A microfluidic chip for centrifugal-driven droplet generation and spreading. This embodiment is similar to the above-mentioned Embodiment 1, except that, as Figure 11 and Figure 12 shown, in this embodiment, the horizontal area of the droplet generation chamber 213 is reduced and the thickness is increased to 2 mm. The distal end curved surface inclines from bottom to top towards the proximal end curved surface. Moreover, the droplet generation unit further includes: a droplet spreading chamber 215 that communicates with the droplet generation chamber 213 and is disposed above the droplet generation chamber 215. The size of the droplet spreading chamber 215 is the same as the size of the droplet generation chamber in the above-mentioned Embodiment 1.
[0101] The process of droplet generation and spreading using the microfluidic chip for centrifugal-driven droplet generation and spreading provided in this embodiment is as follows:
[0102] Samples are added for centrifugation to generate droplets according to the sample addition method in the above-mentioned Embodiment 1. Subsequently, the chip is horizontally flipped 180°. After centrifugation at a speed of 60 RPM for 10 minutes, the droplets in the four droplet generation units are all arranged in a single layer.
[0103] By providing an additional droplet spreading chamber and making corresponding designs for the structure of the droplet generation chamber, this embodiment can achieve active droplet spreading, and the whole process has fewer operation steps, is more controllable, and is more automated.
[0104] For the cassette chip, an additional droplet spreading chamber can also be provided with reference to this embodiment.
[0105] Embodiment 4:
[0106] A microfluidic chip for centrifugal-driven droplet generation and spreading. This embodiment is similar to the above-mentioned Embodiment 1, except that, as Figure 13 shown, in this embodiment, the droplet generation unit further includes: a droplet spreading cavity 215 disposed outside the droplet generation cavity 213 and connected to the droplet generation cavity 213, and an active valve 6 disposed between the liquid generation cavity 213 and the liquid spreading cavity 215; the droplet generation cavity 213 and the droplet spreading cavity 215 are at the same horizontal plane. Paraffin is solidified in the active valve 6.
[0107] The process of droplet generation and spreading using the microfluidic chip for centrifugal-driven droplet generation and spreading provided by this embodiment is as follows:
[0108] At room temperature, samples are added and centrifuged to generate droplets by referring to the sample addition method of the above-mentioned Embodiment 1. After the droplets are generated, they are blocked by the paraffin solidified in the active valve. After the aqueous phase is completely emulsified, the chip is placed in an environment at 50 °C and heated for 5 minutes. The solid paraffin is completely dissolved in the oil phase. Subsequently, after the chip is centrifuged at a speed of 60 RPM for 10 minutes, the droplets in the four droplet generation units are all arranged in a single layer.
[0109] By providing an additional droplet spreading cavity outside the droplet generation cavity and setting an active valve to control the connection relationship between the droplet spreading cavity and the droplet generation cavity, this embodiment can achieve active droplet spreading, and the entire process has fewer operation steps, is more controllable, and is more automated.
[0110] For the cassette chip, an additional droplet spreading cavity can also be set with reference to this embodiment.
[0111] Those skilled in the art can easily understand that the above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, and improvements made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.
Claims
1. A microfluidic chip for centrifugal-driven droplet generation and spreading, characterized in that, Comprising: A substrate; An intermediate layer adhered to the substrate; One or more droplet generation units are provided on the intermediate layer. The droplet generation unit includes: an oil phase pre-storage cavity, an aqueous phase pre-storage cavity, and a droplet generation cavity arranged in sequence from the inside to the outside, a microchannel chip reserved area arranged between the aqueous phase pre-storage cavity and the droplet generation cavity, a waste liquid cavity, a first channel for communicating the oil phase pre-storage cavity and the droplet generation cavity, and a second channel for communicating the droplet generation cavity and the waste liquid cavity; A microchannel chip placed in the microchannel chip reserved area, which is irreversibly bonded to the substrate, and a rectangular microchannel for communicating the aqueous phase pre-storage cavity and the droplet generation cavity is arranged at the bottom; the depth of the rectangular microchannel is less than that of the droplet generation cavity; An upper cover covering the intermediate layer and the microchannel chip; the upper cover includes: a sealing area connected to the microchannel chip reserved area, an aqueous phase sample addition hole communicated with the aqueous phase pre-storage cavity, an oil phase sample addition hole communicated with the oil phase pre-storage cavity, and an air hole communicated with the waste liquid cavity; And a sealant filled in the gaps between the substrate, the intermediate layer, the microchannel chip and the upper cover; The substrate, the intermediate layer and the upper cover are all disc-shaped, and positioning holes aligned with each other are arranged at the center; The horizontal width of the proximal end of the aqueous phase pre-storage cavity is larger than that of the distal end, and the width of the distal end gradually decreases; the two sides of the distal end connected to the proximal end are two arcs of the same circle; The curved surface of the distal end of the droplet generation cavity inclines from bottom to top towards the curved surface of the proximal end, and the droplet generation unit further includes: a droplet spreading cavity communicated with the droplet generation cavity and arranged above the droplet generation cavity.
2. A microfluidic chip for centrifugal-driven droplet generation and spreading, characterized in that, Comprising: A centrifugal locator, and one or more cassette chips; Positioning holes and a card slot for loading the cassette chip are arranged on the centrifugal locator; The cassette chip includes: A substrate; An intermediate layer adhered to the substrate; the intermediate layer includes a droplet generation unit, and the droplet generation unit includes: an oil phase pre-storage cavity, an aqueous phase pre-storage cavity, and a droplet generation cavity arranged in sequence, a microchannel chip reserved area arranged between the aqueous phase pre-storage cavity and the droplet generation cavity, a waste liquid cavity, a first channel for communicating the oil phase pre-storage cavity and the droplet generation cavity, and a second channel for communicating the droplet generation cavity and the waste liquid cavity; A microchannel chip placed in the microchannel chip reserved area, which is irreversibly bonded to the substrate, and a rectangular microchannel for communicating the aqueous phase pre-storage cavity and the droplet generation cavity is arranged at the bottom; the depth of the rectangular microchannel is less than that of the droplet generation cavity; An upper cover covering the intermediate layer and the microchannel chip; the upper cover includes: a sealing area connected to the microchannel chip reserved area, an aqueous phase sample addition hole communicated with the aqueous phase pre-storage cavity, an oil phase sample addition hole communicated with the oil phase pre-storage cavity, and an air hole communicated with the waste liquid cavity; And a sealant filled in the gaps between the substrate, the intermediate layer, the microchannel chip and the upper cover; During emulsification, the oil phase pre-storage cavity is located at the proximal end, and the droplet generation cavity is located at the distal end; The proximal end of the aqueous phase pre-storage chamber has a larger horizontal width than the distal end, and the width of the distal end gradually decreases; the two sides of the distal end connected to the proximal end are two arcs of the same circle; The curved surface of the distal end of the droplet generation chamber inclines from bottom to top towards the curved surface of the proximal end, and the droplet generation unit further includes: a droplet spreading chamber communicated with the droplet generation chamber and arranged above the droplet generation chamber.
3. The microfluidic chip for centrifugal-driven droplet generation and spreading according to claim 1 or 2, characterized in that, The bottom of the oil phase pre-storage chamber is arc-shaped.
4. The microfluidic chip for centrifugal-driven droplet generation and spreading according to claim 1 or 2, characterized in that, The bottom of the droplet generation chamber is arc-shaped.
5. The microfluidic chip for centrifugal-driven droplet generation and spreading according to claim 1 or 2, characterized in that, There are two waste liquid chambers, and correspondingly two second channels are provided; The two waste liquid chambers are respectively located on both sides of the reserved area of the microchannel chip, and the curved surfaces of the proximal ends of the two waste liquid chambers are two arcs of the same circle.
6. The microfluidic chip for centrifugal-driven droplet generation and spreading according to claim 1 or 2, characterized in that, The droplet generation unit further includes: a droplet spreading chamber arranged outside the droplet generation chamber and connected to the droplet generation chamber, and an active valve arranged between the droplet generation chamber and the droplet spreading chamber.
7. The microfluidic chip for centrifugal-driven droplet generation and spreading according to claim 1 or 2, characterized in that, The substrate is made of quartz glass, the microchannel chip is made of PDMS, and the intermediate layer is made of PMMA.
8. The microfluidic chip for centrifugal-driven droplet generation and spreading according to claim 1 or 2, characterized in that, The thickness of the droplet generation chamber is 2 to 3.5 times the diameter of the generated droplet.
Citation Information
Patent Citations
Micro-fluidic chip for centrifugally driving droplet generation and tiling
CN219400208U