A collagen microsphere reaction device and an application method thereof
By designing a collagen microsphere reaction device, a coaxial through-hole array is used to achieve instant mixing and emulsification of collagen solution and cross-linking agent into spheres. This solves the problems of monodispersity and batch-to-batch variability in microsphere preparation in traditional methods, and realizes efficient and convenient collagen microsphere preparation.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- INST OF BIOLOGICAL & MEDICAL ENG GUANGDONG ACAD OF SCI
- Filing Date
- 2023-08-18
- Publication Date
- 2026-05-12
AI Technical Summary
Existing methods for preparing collagen microspheres suffer from low monodispersity, complex preparation processes, and difficulty in precisely controlling microsphere particle size. Furthermore, microfluidic methods for pipelines are subject to problems such as large assembly errors and complex operation.
A collagen microsphere reaction device is designed, including a first container, a second container, a third container, and a sealing cover. The collagen solution and cross-linking agent are mixed and emulsified into microspheres in real time through a coaxial array of through holes and a gas interface. The collagen microspheres are prepared in batches using a series-type focused through hole array.
This method enables convenient and efficient preparation of collagen microspheres, solving the problems of high technical threshold, expensive equipment, and complex procedures in traditional methods, and improving microsphere production efficiency and batch-to-batch consistency.
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Figure CN117138731B_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the technical field of microsphere synthesis, and particularly to a collagen microsphere reaction device and an application method thereof. BACKGROUND
[0002] The existing collagen microsphere preparation methods include solvent evaporation method, seed swelling method, polymerization method, sintering method, spray drying method, phase separation method and microfluidic method, etc. Among them, the microspheres produced based on the traditional microsphere preparation method have low monodispersity, the preparation process is complex, and the particle size of the microspheres is difficult to accurately control; the microfluidic method has a significant advantage in preparing monodisperse microspheres, and the commercially available microfluidic technology mainly includes injection chip microfluidic and pipeline microfluidic methods, wherein the injection chip microfluidic method has a high technical threshold, needs to rely on expensive equipment and molds, and the integration of the injection chip with other equipment is difficult, and the overall microsphere production efficiency is low; the pipeline microfluidic method is a low-cost solution, but there are problems such as large coaxiality assembly error between pipelines, complex multi-channel assembly operation, and large difference between batches of generated microspheres. SUMMARY
[0003] Therefore, the present application provides a collagen microsphere reaction device and an application method thereof, which can conveniently and efficiently realize the preparation of collagen microspheres.
[0004] In one aspect, the present application provides a collagen microsphere reaction device, which comprises a first container, a second container, a third container and a sealing cover plate; the top of the first container, the second container and the third container is open;
[0005] The bottom of the first container is provided with a first through hole array, and the first container is used for loading a continuous phase solution;
[0006] The bottom of the second container is provided with a second through hole array, and the second container is used for loading a crosslinking agent solution; the second container is nested and installed in the first container, a first annular cavity is arranged between the outer diameter of the second container and the inner diameter of the first container, and a first gap is arranged between the lower surface of the bottom of the second container and the upper surface of the bottom of the first container;
[0007] The bottom of the third container is provided with a third through hole array, and the third container is used for loading a dispersed phase solution; the third container is nested and installed in the second container, a second annular cavity is arranged between the outer diameter of the third container and the inner diameter of the second container, and a second gap is arranged between the lower surface of the bottom of the third container and the upper surface of the bottom of the second container;
[0008] The sealing cover covers the top of the first container, the second container, and the third container, and is used to isolate the first container, the first annular cavity, and the second annular cavity into independent spaces. The sealing cover is provided with a first air passage interface, a second air passage interface, and a third air passage interface in sequence above the first container, above the first annular cavity, and above the second annular cavity. The through holes in the first through hole array, the second through hole array, and the third through hole array are coaxially arranged. The first through hole array, the second through hole array, and the third through hole array form a series focusing through hole array based on the first gap and the second gap.
[0009] Optionally, the bottom of the first container is further provided with a first positioning blind hole array, and the bottom of the second container is further provided with a first positioning shaft array and a protruding first cylindrical array, and a second through hole array is disposed in the first cylindrical array; wherein, the first positioning blind hole array and the first positioning shaft array are aligned with the first through hole array and the second through hole array by an interference fit.
[0010] Optionally, the bottom of the second container is further provided with a second positioning blind hole array, and the bottom of the third container is further provided with a second positioning shaft array and a protruding second cylindrical array, and a third through hole array is disposed in the second cylindrical array; wherein, the second positioning blind hole array and the second positioning shaft array are aligned with the second through hole array and the third through hole array by an interference fit.
[0011] Optionally, the device further includes a drive unit, which is connected to the first air passage interface, the second air passage interface and the third air passage interface respectively, and is used to provide drive pressure to the first container, the second container and the third container respectively.
[0012] Optionally, the first through-hole array, the second through-hole array, and the third through-hole array include through-holes of different specifications to form focused through-hole arrays of different sizes.
[0013] On the other hand, embodiments of the present invention provide an application method for a collagen microsphere reaction device, applied to the aforementioned collagen microsphere reaction device; the application method includes:
[0014] A continuous phase solution is added to the first container through the first gas path interface, a crosslinking agent solution is added to the second container through the second gas path interface, and a discrete phase solution is added to the third container through the third gas path interface.
[0015] At a preset reaction temperature, a preset pneumatic pressure is input to the first, second, and third containers through the first, second, and third gas interfaces. This causes the discrete phase solution to mix with the crosslinking agent solution in the second gap through the third through-hole array and enter the second through-hole array. Consequently, the mixture of the continuous phase solution and the crosslinking agent solution and discrete phase solution in the second through-hole converges perpendicularly in the circumferential direction in the first gap between the second and first through-holes, separating the mixture into independent microspheres. These microspheres are then transported to the collection liquid through the first through-hole array to complete crosslinking and curing, resulting in collagen microspheres.
[0016] Optionally, the application method also includes:
[0017] The lyophilized collagen solids were added to deionized water or phosphate buffer solution and dissolved in a water bath to obtain a recombinant collagen solution as a discrete phase solution.
[0018] Optionally, the application method also includes:
[0019] Paraffin oil and surfactant were mixed and a continuous phase solution was prepared by shaking and magnetic stirring.
[0020] Optionally, the application method also includes:
[0021] 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride solid and N-hydroxysuccinimide solid were mixed and dissolved thoroughly in phosphate buffer solution to obtain a crosslinking agent solution.
[0022] Optionally, the device further includes a driving device, which is connected to the first air path interface, the second air path interface, and the third air path interface respectively, and inputs a preset pneumatic pressure to the first container, the second container, and the third container respectively through the first air path interface, the second air path interface, and the third air path interface, including:
[0023] Different preset pneumatic pressures are input to the first container, the second container, and the third container respectively through the first air circuit interface, the second air circuit interface, and the third air circuit interface connected by the drive device.
[0024] This invention provides a collagen microsphere reaction device, comprising: a first container, a second container, a third container, and a sealing cover; the tops of the first, second, and third containers are all open; the bottom of the first container is provided with a first through-hole array, and the first container is used to load a continuous phase solution; the bottom of the second container is provided with a second through-hole array, and the second container is used to load a crosslinking agent solution; the second container is nested within the first container, a first annular cavity is provided between the outer diameter of the second container and the inner diameter of the first container, and a first gap is provided between the lower surface of the bottom of the second container and the upper surface of the bottom of the first container; the bottom of the third container is provided with a third through-hole array, and the third container is used to load a discrete phase solution; the third container is nested within the second container. A second annular cavity exists between the outer diameter of the third container and the inner diameter of the second container, and a second gap is provided between the lower surface of the bottom of the third container and the upper surface of the bottom of the second container. A sealing cover covers the tops of the first, second, and third containers, isolating the first container, the first annular cavity, and the second annular cavity into independent spaces. A first gas path interface, a second gas path interface, and a third gas path interface are sequentially provided above the first container, above the first annular cavity, and above the second annular cavity, respectively. The through holes in the first, second, and third through-hole arrays are coaxially arranged. The first, second, and third through-hole arrays form a series-connected focused through-hole array based on the first and second gaps. This invention, based on structural components with through-hole arrays, forms a series-connected focused through-hole array, enabling automated batch preparation of collagen microspheres through instant mixing of collagen solution and crosslinking agent, emulsification into spheres, and crosslinking and curing. This solves the industry pain points of traditional collagen microsphere preparation methods, which involve high technical barriers, expensive equipment, and complex processes. This invention can conveniently and efficiently prepare collagen microspheres. Attached Figure Description
[0025] To more clearly illustrate the technical solutions in the embodiments of this application, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.
[0026] Figure 1 This is a schematic diagram of the appearance of a collagen microsphere reaction device provided in an embodiment of the present invention;
[0027] Figure 2 This is a schematic diagram of the structural composition of a collagen microsphere reaction device provided in an embodiment of the present invention;
[0028] Figure 3 This is a cross-sectional view of a collagen microsphere reaction device provided in an embodiment of the present invention;
[0029] Figure 4 A schematic diagram of a solution flow mixing reaction provided in an embodiment of the present invention;
[0030] Figure 5 This is a schematic diagram of the collagen microspheres prepared according to an embodiment of the present invention. Detailed Implementation
[0031] To make the objectives, technical solutions, and advantages of this application clearer, the following detailed description is provided in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the scope of this application.
[0032] First, it's important to note that the collagen industry is a crucial component of the currently booming biopharmaceutical and bio-economic sectors. The importance placed on biopharmaceuticals and the bio-industry is constantly increasing, and related plans are providing increasingly detailed descriptions of the development and application of protein preparation technologies. Currently, collagen products on the market are mainly divided into animal-derived collagen and recombinant humanized collagen.
[0033] Animal-derived collagen mainly refers to collagen extracted from animal tissues such as the hides and bones of livestock (pigs and cattle), freshwater fish skin and scales, and deep-sea fish skin. Among them, bovine collagen and porcine collagen have been used in food, cosmetics, medical devices, and other fields, and have a high degree of industrialization. However, the immunogenicity of animal-derived collagen and contamination by pathogens such as viruses are the main obstacles limiting its clinical application.
[0034] Recombinant humanized collagen is a functional fragment of human collagen and its analogues prepared using DNA recombination technology and bio-fermentation technology. Recombinant collagen effectively addresses the safety concerns associated with animal-derived collagen, while also improving collagen's hydrophilicity and immune rejection, and its market share is gradually increasing.
[0035] The main applications of recombinant humanized collagen are in the fields of biomedicine, beauty and skincare, and food. Products mainly include wound dressings, hemostatic materials, injectable fillers, drug carriers, collagen masks, collagen facial cleansers, and creams.
[0036] Porous microspheres prepared based on recombinant collagen maintain tissue compatibility and good hydrophilicity while also having a high specific surface area, low mass density, and strong cell adhesion, making them an ideal high-quality material for medical aesthetic filling, drug delivery, and large-scale 3D cell culture.
[0037] Currently, the application of collagen microspheres in medical aesthetic filling, drug delivery, and large-scale cell culture is still in its early stages. However, with further research on microsphere preparation technology, the application prospects of collagen will be even broader.
[0038] In view of the problems of traditional collagen microsphere preparation methods, on the one hand, embodiments of the present invention provide a collagen microsphere reaction device, including a first container, a second container, a third container and a sealing cover; the tops of the first container, the second container and the third container are all open;
[0039] The bottom of the first container is provided with a first through-hole array, and the first container is used to load a continuous phase solution;
[0040] The bottom of the second container is provided with a second through-hole array. The second container is used to load the crosslinking agent solution. The second container is nested in the first container. A first annular cavity is provided between the outer diameter of the second container and the inner diameter of the first container. A first gap is provided between the lower surface of the bottom of the second container and the upper surface of the bottom of the first container.
[0041] The bottom of the third container is provided with a third through-hole array. The third container is used to load the discrete phase solution. The third container is nested in the second container. There is a second annular cavity between the outer diameter of the third container and the inner diameter of the second container. There is a second gap between the lower surface of the bottom of the third container and the upper surface of the bottom of the second container.
[0042] The sealing cover covers the top of the first container, the second container, and the third container, and is used to isolate the first container, the first annular cavity, and the second annular cavity into independent spaces. The sealing cover is provided with a first air passage interface, a second air passage interface, and a third air passage interface in sequence above the first container, above the first annular cavity, and above the second annular cavity. The through holes in the first through hole array, the second through hole array, and the third through hole array are coaxially arranged. The first through hole array, the second through hole array, and the third through hole array form a series focusing through hole array based on the first gap and the second gap.
[0043] In some embodiments, the bottom of the first container is further provided with a first positioning blind hole array, and the bottom of the second container is further provided with a first positioning shaft array and a protruding first cylindrical array, and a second through hole array is disposed in the first cylindrical array; wherein, the first positioning blind hole array and the first positioning shaft array are aligned with the first through hole array and the second through hole array by an interference fit.
[0044] In some embodiments, the bottom of the second container is further provided with a second positioning blind hole array, and the bottom of the third container is further provided with a second positioning shaft array and a protruding second cylindrical array, and a third through hole array is disposed in the second cylindrical array; wherein, the second positioning blind hole array and the second positioning shaft array are aligned with the second through hole array and the third through hole array by an interference fit.
[0045] In some embodiments, the apparatus further includes a drive device connected to a first air passage interface, a second air passage interface, and a third air passage interface, respectively, and the drive device is used to provide drive pressure to the first container, the second container, and the third container, respectively.
[0046] In some embodiments, the first through-hole array, the second through-hole array, and the third through-hole array include through-holes of different specifications to form focused through-hole arrays of different sizes.
[0047] To explain the device of the present invention in detail, the structure of the device of the present invention will be described below with reference to some specific embodiments. It is easy to understand that the following is an explanation of the technical principle of the present invention and should not be regarded as a limitation of the present invention.
[0048] Reference Figure 1 For example, the assembled collagen microsphere reaction device, Figure 2 As shown, the collagen microsphere reaction device includes a first container 2-1, a second container 2-2, a third container 2-3, and a sealing cover 2-4.
[0049] Among them, such as Figure 3 As shown, the bottom of the first container is provided with a first through hole array 3-1 and a first positioning blind hole array 3-2; the bottom of the second container is provided with a second through hole array 3-3, a second positioning blind hole array 3-4, a first positioning shaft array 3-5 and a first cylindrical array 3-6; the bottom of the third container is provided with a third through hole array 3-7, a second positioning shaft array 3-8 and a second cylindrical array 3-9; the sealing cover is provided with an air passage interface 3-10, including a first air passage interface, a second air passage interface and a third air passage interface.
[0050] Among them, the positioning blind holes and positioning shaft array are used to align the through hole array. Through the interference fit between the holes and the shafts, the structural components are fixed together while all the upper and lower through holes are kept coaxial.
[0051] The first container is used to load the continuous phase solution, the second container is used to load the crosslinking agent solution, and the third container is used to load the discrete phase solution. The first through-hole array is used to focus the continuous and discrete phases; the second through-hole array is used to transport the mixed solution of the crosslinking agent solution and the discrete phase solution; and the third through-hole array is used to transport the discrete phase solution.
[0052] The second through-hole array is coaxial with the first cylindrical array. The bottom surface of the first cylindrical array and the bottom surface of the first container form a first gap. This first gap is used to transport and accelerate the continuous phase solution, allowing it to intersect perpendicularly with the crosslinking agent and discrete phase mixture in the second through-holes in the circumferential direction. Based on shear and viscous forces, the crosslinking agent and discrete phase mixture are separated into independent microspheres, which are then transported to the collection liquid through the first through-hole array to complete crosslinking and curing. For example... Figure 4 The diagram shows the flow mixing reaction of the discrete phase solution, the crosslinking agent solution, and the continuous phase solution, including the flow path 4-1 of the discrete phase solution, the flow path 4-2 of the crosslinking agent solution, the flow path 4-3 of the continuous phase solution, and the collagen microspheres emulsified into spheres 4-4.
[0053] The sealing cover is used to divide the first container, the second container, and the third container into three independent spaces. The first gas passage interface, the second gas passage interface, and the third gas passage interface are used to connect to an external drive device to provide a power source for the flow of the liquid phase.
[0054] It should be noted that the through-hole array and the cylindrical array are coaxial and concentric. The through-hole array is used to transport and focus the solution or mixed solution in the upper container; the cylindrical array provides sufficient space for the solution in the outer container to fill around each cylinder and be uniformly focused into the through-hole from the gap between the bottom surface and the cylinder.
[0055] Depending on the required size of the collagen microspheres, appropriate pore size and spacing can be selected. In some embodiments, the container spacing ranges from 1 to 300 μm, and the pore size ranges from 50 to 500 μm.
[0056] On the other hand, embodiments of the present invention provide a method for applying the collagen microsphere reaction device, which is applied to the aforementioned collagen microsphere reaction device. The method includes:
[0057] A continuous phase solution is added to the first container through the first gas path interface, a crosslinking agent solution is added to the second container through the second gas path interface, and a discrete phase solution is added to the third container through the third gas path interface.
[0058] At a preset reaction temperature, a preset pneumatic pressure is input to the first, second, and third containers through the first, second, and third gas interfaces. This causes the discrete phase solution to mix with the crosslinking agent solution in the second gap through the third through-hole array and enter the second through-hole array. Consequently, the mixture of the continuous phase solution and the crosslinking agent solution and discrete phase solution in the second through-hole converges perpendicularly in the circumferential direction in the first gap between the second and first through-holes, separating the mixture into independent microspheres. These microspheres are then transported to the collection liquid through the first through-hole array to complete crosslinking and curing, resulting in collagen microspheres.
[0059] In some embodiments, the application method may further include:
[0060] The lyophilized collagen solids were added to deionized water or phosphate buffer solution and dissolved in a water bath to obtain a recombinant collagen solution as a discrete phase solution.
[0061] In some embodiments, the application method may further include:
[0062] Paraffin oil and surfactant were mixed and a continuous phase solution was prepared by shaking and magnetic stirring.
[0063] In some embodiments, the application method may further include:
[0064] 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride solid and N-hydroxysuccinimide solid were mixed and dissolved thoroughly in phosphate buffer solution to obtain a crosslinking agent solution.
[0065] In some embodiments, the device further includes a driving device connected to a first air path interface, a second air path interface, and a third air path interface, respectively, and inputting a preset pneumatic pressure to the first container, the second container, and the third container through the first air path interface, the second air path interface, and the third air path interface, respectively, including:
[0066] Different preset pneumatic pressures are input to the first container, the second container, and the third container respectively through the first air circuit interface, the second air circuit interface, and the third air circuit interface connected by the drive device.
[0067] To explain in detail the principle of the technical solution of this invention, the overall process of this invention is described below with reference to some specific embodiments. It is easy to understand that the following is an explanation of the technical principle of this invention and should not be considered as a limitation thereof. The process can be as follows:
[0068] 1. Preparation of discrete phase solution: Weigh 0.5 g of recombinant type III collagen lyophilized solid, add 10 mL of deionized water or 1xPBS (phosphate buffer solution), and dissolve it completely in a water bath at 45-50℃ to obtain a 5% concentration of recombinant collagen solution.
[0069] 2. Preparation of continuous phase solution: Take 20 mL of paraffin oil and 4 mL of Span-80 surfactant, mix them, shake for 2 min, and then stir with a magnetic stirrer for 5 min to ensure thorough mixing;
[0070] 3. Preparation of cross-linking agent solution: Take 0.25 g of EDC (1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride) solid and 0.25 g of NHS (N-hydroxysuccinimide) solid, add them together to a 15 mL centrifuge tube, add 10 mL of 1xPBS solution to dissolve them completely, and the cross-linking agent is obtained;
[0071] 4. Microsphere reactor specifications: First through-hole array pore diameter 350μm, second through-hole array pore diameter 350μm, third through-hole array pore diameter 350μm, cylindrical array and container bottom surface gap 100μm;
[0072] 5. Microsphere reactor setup: Add the prepared continuous phase solution, crosslinking agent solution and discrete phase solution to the first, second and third containers respectively, cover with sealing plates, and connect the first, second and third gas connection ports to external power sources through gas pipes, and input pressures of 8 kPa, 2 kPa and 10 kPa respectively;
[0073] 6. Automated production of collagen microspheres: The microsphere reactor is placed above the surface of the collection liquid at -10℃. The flow-limiting valve is opened, and the pneumatic pressure drives the liquid phase in each container to flow. The microspheres are instantly mixed and emulsified into spheres in a series of through-hole arrays and then transported to the low-temperature collection liquid to complete cross-linking and curing.
[0074] 7. Microsphere morphology characterization: The collected collagen microspheres were solidified in the collection solution for 2 hours. A small number of microspheres were then observed under a microscope. Figure 5 The diameter measured is 210.5 ± 5 μm.
[0075] It should be noted that the pressure parameter is used to adjust the flow rate or velocity of each phase. Based on the flow rate ratio, the concentration of the crosslinking agent and the size of the microspheres can be adjusted. Temperature conditions primarily affect the curing speed of the collagen microspheres.
[0076] In summary, this invention, based on the interlocking of structural components with through-hole arrays to form a series-connected focused through-hole array, solves the problems of high integration difficulty and complex operation in pipeline microfluidic technology. The series-connected focused through-hole array enables instantaneous mixing and emulsification of collagen solution and cross-linking agent into spheres, solving the problem of premature solidification of solution caused by premixing of cross-linking agent, which leads to changes in solution fluidity. In addition, multiple containers use the same driving force to synchronously generate monodisperse collagen microspheres in the focused through-hole array, enabling standardized mass production of collagen microspheres and solving the difficulty of large batch-to-batch variability in traditional methods.
[0077] In some alternative embodiments, the functions / operations mentioned in the block diagrams may not occur in the order shown in the operation diagrams. For example, depending on the functions / operations involved, two consecutively shown blocks may actually be executed substantially simultaneously, or the blocks may sometimes be executed in reverse order. Furthermore, the embodiments presented and described in the flowcharts of this invention are provided by way of example to provide a more comprehensive understanding of the technology. The disclosed methods are not limited to the operations and logic flows presented herein. Alternative embodiments are contemplated in which the order of various operations is altered and sub-operations described as part of a larger operation are executed independently.
[0078] In the description of this specification, references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.
[0079] Although embodiments of the invention have been shown and described, those skilled in the art will understand that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the claims and their equivalents.
[0080] The above is a detailed description of the preferred embodiments of the present invention, but the present invention is not limited to the embodiments described. Those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are all included within the scope defined by the claims of this application.
Claims
1. A collagen microsphere reaction device, characterized in that, It includes a first container, a second container, a third container, and a sealing cover; the tops of the first container, the second container, and the third container are all open; The bottom of the first container is provided with a first through-hole array, and the first container is used to load a continuous phase solution; The bottom of the second container is provided with a second through-hole array. The second container is used to load a crosslinking agent solution. The second container is nested in the first container. A first annular cavity is provided between the outer diameter of the second container and the inner diameter of the first container. A first gap is provided between the lower surface of the bottom of the second container and the upper surface of the bottom of the first container. The bottom of the third container is provided with a third through-hole array. The third container is used to load a discrete phase solution. The third container is nested in the second container. There is a second annular cavity between the outer diameter of the third container and the inner diameter of the second container. There is a second gap between the lower surface of the bottom of the third container and the upper surface of the bottom of the second container. The sealing cover covers the top of the first container, the second container, and the third container, and is used to isolate the first container, the first annular cavity, and the second annular cavity into independent spaces. The sealing cover is provided with a first air passage interface, a second air passage interface, and a third air passage interface in sequence above the first container, the first annular cavity, and the second annular cavity. The through holes in the first through hole array, the second through hole array, and the third through hole array are coaxially arranged. The first through hole array, the second through hole array, and the third through hole array form a series focusing through hole array based on the first gap and the second gap.
2. The collagen microsphere reaction device according to claim 1, characterized in that, The bottom of the first container is also provided with a first positioning blind hole array, and the bottom of the second container is also provided with a first positioning shaft array and a protruding first cylindrical array, and the second through hole array is disposed in the first cylindrical array; wherein, the first positioning blind hole array and the first positioning shaft array are aligned with the first through hole array and the second through hole array by an interference fit.
3. The collagen microsphere reaction device according to claim 1, characterized in that, The bottom of the second container is also provided with a second positioning blind hole array, and the bottom of the third container is also provided with a second positioning shaft array and a protruding second cylindrical array. The third through hole array is disposed in the second cylindrical array; wherein, the second positioning blind hole array and the second positioning shaft array are aligned with the second through hole array and the third through hole array by an interference fit.
4. The collagen microsphere reaction device according to claim 1, characterized in that, The device further includes a driving device, which is connected to the first air passage interface, the second air passage interface and the third air passage interface respectively, and is used to provide driving pressure to the first container, the second container and the third container respectively.
5. The collagen microsphere reaction device according to claim 1, characterized in that, The first through-hole array, the second through-hole array, and the third through-hole array include through-holes of different specifications to form focused through-hole arrays of different sizes.
6. A method of applying a collagen microsphere reaction device, wherein the device is used in accordance with any one of claims 1 to 5, characterized in that, The application method includes: The continuous phase solution is added to the first container through the first gas path interface, the crosslinking agent solution is added to the second container through the second gas path interface, and the discrete phase solution is added to the third container through the third gas path interface; At a preset reaction temperature, a preset pneumatic pressure is input to the first container, the second container, and the third container through the first gas path interface, the second gas path interface, and the third gas path interface. This causes the discrete phase solution to mix with the crosslinking agent solution in the second gap through the third through-hole array and enter the second through-hole array. Consequently, the continuous phase solution and the mixture of the crosslinking agent solution and the discrete phase solution in the second through-hole converge perpendicularly in the circumferential direction in the first gap between the second through-hole and the first through-hole, separating the mixture into independent microspheres. These microspheres are then transported to the collection liquid through the first through-hole array to complete crosslinking and curing, resulting in collagen microspheres.
7. The application method of the collagen microsphere reaction device according to claim 6, characterized in that, The application method further includes: The lyophilized collagen solid is added to deionized water or phosphate buffer solution and dissolved in a water bath to obtain a recombinant collagen solution as the discrete phase solution.
8. The application method of the collagen microsphere reaction device according to claim 6, characterized in that, The application method also includes: The continuous phase solution is prepared by mixing paraffin oil with a surfactant and then mixing by shaking and magnetic stirring.
9. The application method of the collagen microsphere reaction device according to claim 6, characterized in that, The application method also includes: The crosslinking agent solution was obtained by mixing 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride solid and N-hydroxysuccinimide solid, and then dissolving them thoroughly in a phosphate buffer solution.
10. The application method of the collagen microsphere reaction device according to claim 6, characterized in that, The device further includes a driving device, which is connected to the first air passage interface, the second air passage interface, and the third air passage interface respectively. The step of inputting a preset pneumatic pressure to the first container, the second container, and the third container through the first air passage interface, the second air passage interface, and the third air passage interface respectively includes: The first air path interface, the second air path interface, and the third air path interface connected through the drive device respectively input different preset pneumatic pressures to the first container, the second container, and the third container.