A method and apparatus for tiling droplets

Abstract

The application discloses a preparation method and device for tiling droplets, and relates to the technical field of biological microfluids, comprising the steps of mold preparation, upper cover preparation, droplet tiling, cleaning and reuse, etc. The method adopts a simple process of film limiting, potting glue forming a substrate, and secondary forming of a prepolymer, does not need precise photoetching equipment, and has the advantages of simple process, low cost, and strong replicability. The device realizes self-assembly tiling of droplets by means of micro-grooves, does not need external driving equipment, can effectively prevent droplet slipping and fusion, and stably obtains a uniform and ordered high-density droplet array. Different particle sizes of droplets can be adapted by adjusting the thickness of the film, and multiple biological detection scenes can be adapted. The device has a small and compact structure, is easy to integrate, can be repeatedly used after being cleaned and dried, effectively breaks through the bottleneck of traditional technology, and has prominent practical and industrial values.

Description

Technical Field

[0001] This invention relates to the field of biomicrofluidics, and in particular to a method and apparatus for fabricating flat droplets. Background Technology

[0002] Droplet microfluidics has become a key platform widely used in digital PCR, single-cell analysis, and other fields in recent years. One of its core steps is to achieve high-density, uniform, and stable planar arrangement of droplets for high-throughput fluorescence detection and quantitative analysis. Existing droplet spreading methods mainly rely on mechanisms such as passive capture by microcavity arrays, guidance by hydrophilic or hydrophobic surface patterns, or gravity sedimentation. While these methods are effective in specific scenarios, they generally suffer from poor spreading uniformity, easy droplet fusion and accumulation, reliance on external driving devices (such as centrifuges or pressure pumps), and low system integration. Especially in practical applications, how to realize a droplet spreading device that requires no complex peripherals, has a simple structure, and can be stably replicated remains a technical bottleneck that urgently needs to be solved.

[0003] To address the aforementioned issues, some studies have attempted to fabricate planar substrates with microstructures using soft lithography and polymer molding processes. For example, PDMS arrays can be molded using photoresist templates, utilizing capillary action to guide droplet positioning; or droplet self-assembly can be achieved by modifying surface properties to control hydrophilicity and hydrophobicity. However, these methods often involve complex processes, high mold costs, insufficient bonding stability between the PDMS structure and the rigid substrate, and difficulty in ensuring the consistency of microstructure size and morphology, thus limiting their scalability and reusability.

[0004] Therefore, there is an urgent need to develop a droplet spreading device that is simple in process, low in cost, stable in structure, reusable, and easy to integrate with detection modules, so as to achieve efficient, uniform, and reliable droplet array arrangement and further improve the practicality of droplet microfluidic systems. Summary of the Invention

[0005] To address the above technical problems, this invention provides a method and apparatus for fabricating flat droplets, achieving efficient, uniform, and reliable droplet array arrangement, and improving the practicality of droplet microfluidic systems.

[0006] To achieve the above objectives, the present invention provides the following solution: This invention provides a method for preparing a flattened liquid droplet, comprising the following steps: Mold preparation: A thin film is placed in a first container. The thickness of the film is the difference between the total internal height of the bottom and the diameter of the droplet. Potting compound is added to the first container and made to completely cover the film. The potting compound is left to cure. The cured potting compound is removed from the first container and separated from the film. A groove is formed at the position where the cured potting compound contacts the film. To prepare the top cover, the cured potting compound is placed in a second container with the groove facing upwards. A prepolymer is added to the second container and the prepolymer completely covers the groove. A clean cover plate is then placed on the prepolymer to form a quasi-mold. After the prepolymer in the quasi-mold has cured, the cover plate is removed from the second container along with the cured prepolymer to obtain the top cover. To create a flat droplet, drop the droplet into the bottom and cover it with the top cover to form a closed droplet flattening device with an internal microstructure. After cleaning and use, the top cover and the bottom should be cleaned and dried for later use.

[0007] Optionally, in the mold preparation step, a room-temperature curing potting compound may be used.

[0008] Optionally, in the mold preparation step, the film is a polydimethylsiloxane film.

[0009] Optionally, in the top cover preparation step, the prepolymer is polydimethylsiloxane.

[0010] Optionally, in the top cover preparation step, the prepolymer is cured in an oven at 70-100 degrees Celsius for 12-16 hours.

[0011] Optionally, in the cover preparation step, the cover plate is made of a glass slide.

[0012] Optionally, in the cover preparation step, the cover plate avoids introducing air bubbles when covering the prepolymer.

[0013] Optionally, during the cleaning step, a spray bottle containing HFE-7500 electronic fluorination solution is used to thoroughly rinse the lower surface of the top cover and the upper surface of the bottom that came into contact with the droplets; subsequently, an air gun is used to completely blow away any remaining HFE-7500 electronic fluorination solution to ensure that the surface is clean and dry for the next experiment.

[0014] Optionally, the first container and the second container are petri dishes.

[0015] The present invention also provides an apparatus for preparing a flat droplet, comprising an upper cover and a lower base, wherein the upper cover has a raised structure on the side facing the lower base, and the top of the lower base is open, and when the upper cover covers the lower base, the distance between the bottom of the raised structure and the inner bottom of the lower base is equal to the diameter of the droplet.

[0016] The present invention achieves the following technical effects compared to the prior art: The method and apparatus for fabricating droplets provided by this invention employ a simplified process involving thin-film positioning, curing of the encapsulated substrate to form a groove, and secondary molding of the prepolymer to prepare the top cover. This eliminates the need for high-precision photolithography equipment and customized precision molds, significantly simplifying the processing flow and reducing manufacturing costs. By relying on the microgroove structure and material capillary effect, droplet self-assembly and flattening are achieved, eliminating the need for external driving equipment such as centrifuges and pressure pumps. The closed microstructure effectively prevents droplet slippage, aggregation, and fusion, stably forming a high-density, uniform, ordered, and monodisperse droplet array, greatly improving the accuracy and repeatability of high-throughput fluorescence detection and quantitative analysis. By adjusting the film thickness, different droplet sizes can be accommodated, making it widely applicable in various scenarios such as digital analysis, single-cell analysis, and high-throughput biological detection, demonstrating strong adaptability and versatility.

[0017] Meanwhile, this device features a simple structure, is easy to disassemble and clean, and can be repeatedly dried and reused. The equipment is compact and highly integrated, directly adaptable to various microfluidic detection devices, and easily modularized. This invention boasts a simple process, low cost, and strong reproducibility, effectively overcoming the bottlenecks in the large-scale application of traditional technologies, significantly improving the practicality of droplet microfluidic systems, and possessing excellent industrialization and promotion value. Attached Figure Description

[0018] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0019] Figure 1 This is a schematic diagram of the mold structure in the device for spreading droplets according to the present invention; Figure 2 This is a schematic diagram of the structure of the upper cover in the device for spreading liquid droplets according to the present invention; Figure 3 This is a schematic diagram of the upper cover and lower base of the device for spreading liquid droplets according to the present invention; Figure 4 This is a cross-sectional view of the upper cover and lower base of the device for spreading droplets according to the present invention.

[0020] Explanation of reference numerals in the attached figures: 1. Mold; 2. Glass slide; 3. Prepolymer; 4. Top cover; 5. Bottom base; 11. Groove. Detailed Implementation

[0021] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0022] Example 1: like Figures 1 to 4 As shown, this embodiment provides a method for fabricating droplets, including the following steps: Mold preparation: A thin film is placed in a first container. The thickness of the film is the difference between the total internal height of the bottom 5 and the diameter of the droplet. Potting compound is added to the first container and the potting compound completely covers the film. The potting compound is left to cure. The cured potting compound is removed from the first container and separated from the film. The cured potting compound forms a groove 11 at the contact point with the film. To prepare the top cover, the cured potting compound is placed in the second container with the groove 11 facing upwards. Prepolymer 3 is added to the second container and the prepolymer 3 completely covers the groove 11. A clean cover plate is placed on the prepolymer 3 to form a quasi-mold 1. After the prepolymer 3 in the quasi-mold 1 is cured, the cover plate is removed from the second container through the cured prepolymer 3 to obtain the top cover 4. To create a flat droplet, drop the droplet into the lower bottom 5 and cover it with the upper cover 4. As the prepolymer 3 below the upper cover 4 extends downward, a closed droplet flattening device is formed between the lower surface of the prepolymer 3 and the inner bottom of the lower bottom 5, which can only accommodate a layer of liquid molecules. After cleaning and use, clean and dry the top cover 4 and bottom 5 before use.

[0023] When using the method and apparatus for fabricating droplets provided by this invention, a simple process is employed: thin film limiting, potting adhesive curing to form a grooved substrate, and prepolymer 3 secondary molding to prepare the top cover 4. This eliminates the need for high-precision photolithography equipment and customized precision molds 1, significantly simplifying the processing flow and reducing manufacturing costs. Droplet self-assembly and flattening are achieved through the microgroove structure 11 and the capillary effect of the material. External driving equipment such as centrifuges and pressure pumps are unnecessary; the closed microstructure effectively limits droplet positioning, effectively preventing droplet slippage, aggregation, and fusion. This results in a stable, high-density, uniform, ordered, and monodisperse droplet array, significantly improving the accuracy and repeatability of high-throughput fluorescence detection and quantitative analysis. By adjusting the film thickness, different droplet sizes can be accommodated, making it widely applicable in various scenarios such as digital analysis, single-cell analysis, and high-throughput biological detection, demonstrating strong adaptability and versatility.

[0024] Meanwhile, this device features a simple structure, is easy to disassemble and clean, and can be repeatedly dried and reused. The equipment is compact and highly integrated, directly adaptable to various microfluidic detection devices, and easily modularized. This invention boasts a simple process, low cost, and strong reproducibility, effectively overcoming the bottlenecks in the large-scale application of traditional technologies, significantly improving the practicality of droplet microfluidic systems, and possessing excellent industrialization and promotion value.

[0025] In the mold preparation process, the potting compound is a room temperature curing potting compound, such as a two-component epoxy resin-based adhesive, which does not require heating to cure. This can effectively avoid problems such as film deformation and molding accuracy deviation caused by high temperature conditions. After curing, the colloid has high hardness, dense structure, and excellent dimensional stability, which greatly improves the structural strength and molding consistency of the base groove 11.

[0026] In the mold preparation process, polydimethylsiloxane (PDMS) film is used. PDMS film is chosen because it has a smooth surface, good flexibility, and good release properties, which effectively reduces the adhesion between the potting compound and the film after curing, ensuring the integrity and precise dimensions of the groove 11. The film thickness can be selected according to the droplet diameter, allowing the mold to fabricate devices suitable for different liquids.

[0027] In the top cover preparation step, prepolymer 3 is polydimethylsiloxane. After curing, polydimethylsiloxane prepolymer 3 has good light transmittance, good biocompatibility, and moderate flexibility, making it suitable for the encapsulation and tiling of microfluidic droplets. It can effectively reduce droplet interface adsorption and breakage, and can be softened by heating, making it easy to separate from the glass slide 2.

[0028] In the top cover preparation step, prepolymer 3 is cured in an oven at 70-100 degrees Celsius for 12-16 hours. The petri dish, serving as the second molding container, ensures a smooth surface for prepolymer 3, effectively preventing warping and deformation. After curing, polydimethylsiloxane prepolymer 3 exhibits excellent light transmittance, good biocompatibility, and moderate flexibility, making it suitable for microfluidic droplet encapsulation and tiling scenarios, effectively reducing droplet interface adsorption and breakage. The glass slide 2 cover plate has high flatness and rigidity; pressure covering ensures uniform thickness of prepolymer 3, eliminating unevenness on the surface of the top cover 4. Strictly avoiding air bubble ingress ensures a smooth and defect-free internal microstructure for the top cover 4, preventing air bubble defects from interfering with droplet arrangement accuracy. The 70-100℃ long-term constant-temperature curing process enables full cross-linking and curing of polydimethylsiloxane prepolymer 3, resulting in a stable, non-deformable, and dimensionally consistent top cover 4 suitable for long-term repeated use.

[0029] In the top cover preparation step, a glass slide 2 is used as the cover plate. The glass slide 2 has high flatness and rigidity; pressure covering ensures uniform thickness of the prepolymer 3 and eliminates unevenness on the surface of the top cover 4. When the glass slide 2 covers the PDMS liquid surface, air bubbles must be avoided to ensure a smooth and defect-free internal microstructure of the top cover 4 and prevent air bubble defects from interfering with the accuracy of droplet arrangement. A long-term constant-temperature curing process at 70-100℃ is used to fully cure the polydimethylsiloxane prepolymer 3. After molding, the top cover 4 has a stable structure, is not easily deformed, has high dimensional consistency, and can be used repeatedly for a long time.

[0030] During the cleaning process, a spray bottle containing HFE-7500 electronic fluorination solution was used to thoroughly rinse the lower surface of the top cover 4 and the upper surface of the bottom 5 that came into contact with the liquid droplets. Subsequently, an air gun was used to completely blow away any remaining HFE-7500 electronic fluorination solution, ensuring that the surface was clean and dry for use in the next experiment.

[0031] HFE-7500 electronic fluorinated fluid has a strong dissolving ability for residual impurities in microfluidic droplets, evaporates quickly and leaves no residue, and can efficiently remove trace amounts of droplet residue and contaminants from the surface of the device. At the same time, it will not corrode the epoxy resin substrate and polydimethylsiloxane structure, thus preserving the microstructure morphology of the device. Combined with an air gun purging process, it can thoroughly remove cleaning agent residue, avoiding problems such as droplet adhesion, uneven distribution, and fusion failure caused by residual liquid. This ensures the consistency and repeatability of experiments for each reuse of the device, effectively extending the device's service life.

[0032] The first and second containers are petri dishes, especially disposable petri dishes. The petri dishes have wide openings and flat bottoms, which can ensure the flatness of the potting compound during molding, reduce the problem of substrate warping and deformation after molding, and are easy to obtain and inexpensive, making them suitable for mass production.

[0033] Example 2: This embodiment is a more detailed embodiment based on Embodiment 1, and specifically includes the following steps: Mold preparation: First, a PDMS film measuring 41.5×11.0×0.05 mm is tightly adhered to the center area of ​​a disposable culture dish. Then, epoxy resin AB glue is mixed at a weight ratio of 4:1 and slowly poured into the culture dish until the PDMS film is completely covered. This mold 1 is then allowed to cure at room temperature on a horizontal platform for 24 hours.

[0034] Top cover preparation: After the epoxy resin AB glue has fully cured, peel it off from the petri dish and make the side with the PDMS film groove 11 facing up as mold 1; slowly pour the pre-prepared and degassed PDMS prepolymer 3 (a mixture of matrix and curing agent at a weight ratio of 10:1) onto the surface of mold 1, ensuring complete coverage of the microstructure area, i.e., the groove 11 part; then cover the PDMS prepolymer 3 solution with a clean 75.5×25.5×1.1 mm glass slide 2, and carefully operate during the process to avoid introducing air bubbles; The mold 1 covered with the glass slide 2 was placed in an 80°C oven for curing for 13 hours to allow the PDMS prepolymer 3 to fully cure. After curing, the glass slide 2, together with the cured PDMS layer (at this time, the PDMS layer has replicated the microstructure of the mold 1 and becomes the upper cover 4 of the device), was carefully peeled off from the epoxy resin mold 1. To create a flat droplet, align the prepared PDMS top cover 4 with the 75.5×25.5×2 mm glass counting chamber (with an internal groove 11 measuring 70.5×20.5×0.1 mm) which serves as the base component, and assemble them by physical pressing to form a closed droplet flattening device with an internal microstructure. In use, drop the generated droplet into the center of the bottom 5 of the device, and slowly cover it with the top cover 4. The droplet will spontaneously flatten in the microgroove gaps inside the device, forming a stable single-layer arrangement, which can then be directly used for subsequent fluorescence microscopy observation and digital analysis. Clean by thoroughly rinsing the lower surface of the top cover 4 and the upper surface of the bottom 5, which came into contact with the liquid droplets, using a spray bottle filled with HFE-7500 electronic fluorination solution. Then, use an air gun to completely blow away any remaining HFE-7500 electronic fluorination solution, ensuring the surfaces are clean and dry for the next experiment.

[0035] Example 3: This embodiment is an apparatus prepared based on the method for preparing droplets in Embodiment 1. Specifically, it includes an upper cover 4 and a lower base 5. The upper cover 4 has a raised structure on the side facing the lower base 5. The top of the lower base 5 is open. When the upper cover 4 covers the lower base 5, the distance between the bottom of the raised structure and the inner bottom of the lower base 5 is equal to the diameter of the droplet.

[0036] More specifically, the bottom 5 can be made of glass, with an open top and a groove 11 of a certain depth in the middle to accommodate droplets.

[0037] When in use, drop the generated droplet into the center of the bottom 5, and slowly cover it with the top cover 4. The droplet will spontaneously spread out in the micro-groove gap between the bottom 5 and the top cover 4, forming a stable single-layer arrangement, which can then be directly used for subsequent fluorescence microscopy observation and digital analysis.

[0038] It should be noted that, for those skilled in the art, it is obvious that the present invention is not limited to the details of the above exemplary embodiments, and that the present invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered exemplary and non-limiting in all respects, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, all variations falling within the meaning and scope of equivalents of the claims are intended to be included within the present invention, and no reference numerals in the claims should be construed as limiting the scope of the claims.

[0039] This specification uses specific examples to illustrate the principles and implementation methods of the present invention. The descriptions of the above embodiments are only for the purpose of helping to understand the method and core ideas of the present invention. Furthermore, those skilled in the art will recognize that, based on the ideas of the present invention, there will be changes in the specific implementation methods and application scope. Therefore, the content of this specification should not be construed as a limitation of the present invention.

Claims

1. A method for preparing droplets for spreading out, characterized in that, Includes the following steps: Mold preparation: A film is placed in a first container, the thickness of which is the difference between the total internal height of the bottom and the diameter of the droplet. Potting compound is added to the first container and made to completely cover the film. The potting compound is then allowed to cure. The cured potting compound is removed from the first container and separated from the film, and a groove is formed at the position where the cured potting compound contacts the film; To prepare the top cover, the cured potting compound is placed in a second container with the groove facing upwards. A prepolymer is added to the second container and the prepolymer completely covers the groove. A clean cover plate is then placed on the prepolymer to form a quasi-mold. After the prepolymer in the quasi-mold has cured, the cover plate is removed from the second container along with the cured prepolymer to obtain the top cover. To create a flat droplet, drop the droplet into the bottom and cover it with the top cover to form a closed droplet flattening device with an internal microstructure. After cleaning and use, the top cover and the bottom should be cleaned and dried for later use.

2. The method for preparing droplets for planarization according to claim 1, characterized in that, In the mold preparation process, room temperature curing potting compound is used.

3. The method for preparing droplets according to claim 1, characterized in that, In the mold preparation step, the film is a polydimethylsiloxane film.

4. The method for preparing droplets according to claim 1, characterized in that, In the top cover preparation step, polydimethylsiloxane is used as the prepolymer.

5. The method for preparing droplets according to claim 1, characterized in that, In the top cover preparation step, the prepolymer is cured in an oven at 70-100 degrees Celsius for 12-16 hours.

6. The method for preparing droplets according to claim 1, characterized in that, In the preparation step of the top cover, the cover plate is made of a glass slide.

7. The method for preparing droplets according to claim 1, characterized in that, In the top cover preparation step, the cover plate should be placed on the prepolymer to avoid introducing air bubbles.

8. The method for preparing droplets according to claim 1, characterized in that, During the cleaning process, a spray bottle containing HFE-7500 electronic fluorination solution was used to thoroughly rinse the lower surface of the top cover and the upper surface of the bottom that came into contact with the liquid droplets. Subsequently, an air gun was used to completely blow away any remaining HFE-7500 electronic fluorination solution, ensuring that the surface was clean and dry for use in the next experiment.

9. The method for preparing droplets according to claim 1, characterized in that, The first container and the second container are petri dishes.

10. An apparatus for preparing droplets, characterized in that, It includes an upper cover and a lower bottom. The upper cover has a raised structure on the side facing the lower bottom. The top of the lower bottom is open. When the upper cover covers the lower bottom, the distance between the bottom of the raised structure and the inner bottom of the lower bottom is equal to the diameter of the droplet.

Images