Vesicle fusion chip preparation device

Abstract

The utility model provides a vesicle fusion chip preparation device. The vesicle fusion chip preparation device comprises a vesicle generation device, a vesicle curing device and a vesicle receiving device, the vesicle generating device comprises a hydrogel solution pool 1, an injection device, a central pipeline 12 and a barometer 10, and the vesicle curing device comprises an atomization device, a biological ink pool module 13 and an ultraviolet curing device 15. The technical problems that in the prior art, the sizes of the vesicles cannot be uniform and controllable, the prepared vesicles are all large-size vesicles, and the vesicles can only be subjected to suspension culture in a culture medium are solved.

Description

Technical Field

[0001] The utility model belongs to the technical field of biotechnology, and particularly relates to a device for preparing a vesicle fusion chip. Background Art

[0002] Cell microencapsulation is to wrap target cells in one or several materials with good biocompatibility and semi-permeable membrane characteristics, enabling the target cells to achieve immune isolation and avoid the attack of macromolecular immune substances and immune cells, while allowing metabolites, small molecule nutrients and cell active substances to freely enter and exit the microcapsules. Compared with traditional cell culture, it has better advantages. Among them, the technology of encapsulating cells with vesicles can prepare organoids in a high-throughput one-step manner without a secondary process of washing, which is convenient for industrialization and standardization; the vesicles are uniform and controllable in size, and the influence of the size of cell spheres on viability and function can be studied, and it can also be used for the study of stem cell differentiation; however, the culture operation is difficult, and the vesicles are cultured in suspension in the culture medium, which is not conducive to changing the culture medium.

[0003] The droplet microfluidics technology based on the water-in-oil emulsion system can produce cell-loaded microspheres with uniform size, but the throughput is insufficient, and the process of demulsifying and washing the microspheres increases the complexity of the process. The gas jet microfluidics technology combined with rapid ion crosslinking can prepare organoid microcapsules in large quantities, but usually can only prepare large-sized microcapsules (larger than 500 μm) under the condition of low gas flow rate. As the gas flow rate increases, the morphology of the microcapsules is unstable, and it is difficult to prepare small-sized microcapsules with uniform size. More importantly, the organoids prepared by the above technologies can only grow in suspension in the culture medium, and none of them can reproduce a series of tumor physiological phenomena such as vascular infiltration and metastasis, and it is difficult to evaluate the role and effect of anti-angiogenic drugs in tumor treatment. Summary of the Invention

[0004] This application provides a device for preparing a vesicle fusion chip, which is used to solve the technical problems in the prior art that the size of vesicles cannot be uniformly controlled, the prepared vesicles are all large-sized vesicles, and the vesicles can only be cultured in suspension in the culture medium.

[0005] The present utility model requests protection for a preparation device for a vesicle fusion chip, which includes a vesicle generation device, a vesicle solidification device, and a vesicle receiving device; the vesicle generation device includes a hydrogel solution pool 1, an injection device, a central pipeline 12, and a gas device. The injection device is arranged below the hydrogel solution pool 1, the injection device is arranged above the central pipeline 12, the central pipeline 12 is arranged above the vesicle receiving device, and the gas device is arranged at the entrance of the central pipeline; the vesicle solidification device includes an atomization device, a bioink pool module 13, and an ultraviolet light curing device 15. The atomization device is arranged below the gas device and is connected to the central pipeline 12, the bioink pool 13 is arranged below the atomization device and is connected to the central pipeline 12, and the ultraviolet light curing device 15 is arranged below the vesicle receiving device.

[0006] Further, the gas device includes a gas pipeline 19, a barometer 10, and a gas controller 11. The gas pipeline 19 is connected to the central pipeline 12. The barometer 10 is used to detect the gas pressure. The gas controller 11 is used to adjust the gas flow rate.

[0007] Further, a primary digestion chamber 18 is connected to the hydrogel solution pool 1, and the separated tissue can be digested in the primary digestion chamber 18.

[0008] Further, the injection device includes an injection head size converter 3 and injection heads of different models.

[0009] Still further, the injection heads of different models include a 32G injection head 4, a 34G injection head 5, and a 28G injection head 6.

[0010] Further, a liquid controller 2 is arranged on the hydrogel solution pool 1.

[0011] Further, the atomization device includes an atomizer 7, an atomization spray pipe 8, and a valve 9. The atomized crosslinking agent solution forms an atomized spray through the atomizer, and the atomized spray enters the central pipeline 12 through the atomization spray pipe 8.

[0012] Further, the vesicle receiving device includes an X-Y axis moving device 17 and a receiving orifice plate 16. The receiving orifice plate 16 can move along the x and y axes, and the ultraviolet light curing device 15 is arranged below the receiving orifice plate 16.

[0013] Further, a waste discharge pipe 14 is arranged below the atomization spray pipe 8.

[0014] Beneficial effects

[0015] The fusion chip preparation technology of this application adopts the synchronous technology of air cutting water + positioning photocuring. By controlling the liquid flow diameter, the vesicle size can be accurately controlled to achieve regional vesicle quantitative curing. Since the liquid flow rate, the diameter of the needle, and the gas flow rate all affect the vesicle size, the thinner the inner diameter of the needle, the easier it is to produce smaller vesicles, and the faster the gas flow rate, the smaller the vesicles. The liquid flow rate of the hydrogel solution, the size of the injection head, and the gas flow rate in this application can all be adjusted. We can control the liquid flow rate of the hydrogel solution, the size of the injection head, and the gas flow rate according to the required vesicle size, so that the gas flow, the hydrogel solution flow rate, and the needle model are coordinated to produce small vesicles with uniform size. Compared with the prior art, which usually can only prepare large-sized microcapsules (greater than 500 μm) under the condition of low gas flow rate, the lower limit of the microcapsule size that can be achieved by the 28G injection head in this application is about 300 μm, the lower limit of the microcapsule size that can be achieved by the 32G injection head is about 150 μm, and the lower limit of the microcapsule size that can be achieved by the 34G injection head is about 80 μm.

[0016] In the prior art, the formed hydrogel droplets directly fall into the crosslinking pool. When the droplets collide with the liquid surface of the crosslinking pool, it will inevitably cause the splitting of some vesicles, thus affecting the uniformity of the microcapsules, and the influence is more significant when the solution flow rate is large. In this application, the crosslinking agent is atomized and then crosslinked with the vesicles to reduce the collision of the vesicles and gently crosslink the hydrogel droplets, thereby ensuring the size uniformity.

[0017] In the prior art, the microcapsules are in a suspended state during the subsequent microcapsule culture process after being crosslinked in the crosslinking pool, which is not convenient for changing the culture medium. During the process of changing the liquid, some floating microcapsules may be sucked away, resulting in loss. The inventor team found during the research process that after the vesicles are cured by the bioink and the ultraviolet light curing device, the microcapsules can be fixed in the hydrogel matrix, which is convenient for liquid changing operation. Description of the Drawings

[0018] Figure 1 Structural schematic diagram of the product of the present utility model

[0019] 1 - Hydrogel solution pool; 2 - Liquid controller; 3 - Injection head size converter; 4 - 32G injection head; 5 - 34G injection head; 6 - 28G injection head; 7 - Atomizer; 8 - Atomizing spray pipe; 9 - Valve; 10 - Barometer; 11 - Gas controller; 12 - Central pipeline; 13 - Bioink pool module; 14 - Waste discharge pipe; 15 - Ultraviolet light curing device; 16 - Receiving orifice plate; 17 - X - Y axis moving device; 18 - Primary digestion chamber; 19 - Gas pipeline Detailed Embodiments

[0020] The present application will be described in detail below through embodiments, but this does not mean any adverse limitations to the present application. The present application has been described in detail herein, and specific embodiments thereof have also been disclosed. It will be obvious to those skilled in the art that various changes and improvements can be made to the specific embodiments of the present application without departing from the spirit and scope of the present application.

[0021] Embodiment 1

[0022] A vesicle fusion chip preparation device includes a vesicle generation device, a vesicle solidification device, and a vesicle receiving device; the vesicle generation device includes a hydrogel solution pool 1, an injection device, a central pipeline 12, and a gas device. The injection device is arranged below the hydrogel solution pool 1, the injection device is connected to the central pipeline 12, the central pipeline 12 is arranged above the vesicle receiving device, and the gas device is arranged at the entrance of the central pipeline; the vesicle solidification device includes an atomization device, a bioink pool module 13, and an ultraviolet light curing device 15. The atomization device is arranged below the gas device and is connected to the central pipeline 12, the bioink pool 13 is arranged below the atomization device and is connected to the central pipeline 12, and the ultraviolet light curing device 15 is arranged above the vesicle receiving device. Cells and the hydrogel solution are mixed in the hydrogel solution pool 1, and the hydrogel solution mixed with cells flows into the central pipeline 12 through an injection head, and forms hydrogel droplets under the action of gas shearing at the entrance, and the hydrogel droplets fall from the central pipeline 12.

[0023] A primary digestion chamber 18 is connected to the hydrogel solution pool 1, and separated tissues can be digested in the primary digestion chamber 18. The primary cell suspension after digestion and the hydrogel solution meet and enter the hydrogel solution pool 1.

[0024] A liquid controller 2 is arranged on the hydrogel solution pool 1, and the flow rate of the hydrogel solution is controlled by the liquid controller 2.

[0025] The injection device includes an injection head size converter 3 and injection heads of different models. The injection heads of different models include a 32G injection head 4, a 34G injection head 5, and a 28G injection head 6, and the model of the injection head used can be adjusted by the injection head size converter 3.

[0026] The gas device includes a gas pipeline 19, a barometer 10, and a gas controller 11. The gas pipeline 19 is connected to the central pipeline 12. The barometer 10 is used to detect the gas pressure. The gas controller 11 is used to adjust the flow rate of the gas.

[0027] Since the flow rate of the liquid, the diameter of the needle, and the flow rate of the gas stream all affect the size of the vesicles, a thinner inner diameter of the needle is more likely to produce smaller vesicles, and the faster the gas stream speed, the smaller the vesicles. The flow rate of the hydrogel solution, the size of the injection head, and the flow rate of the gas in this application can all be adjusted. We can control the flow rate of the hydrogel solution, the size of the injection head, and the flow rate of the gas according to the size of the required vesicles, so that the gas stream, the flow rate of the hydrogel solution, and the needle model are coordinated to adjust to produce small vesicles with uniform size. The lower limit of the microcapsule size that can be achieved by the 28G injection head in this application is about 300μm, the lower limit of the microcapsule size that can be achieved by the 32G injection head is about 150μm, and the lower limit of the microcapsule size that can be achieved by the 34G injection head is about 80μm.

[0028] The atomization device includes an atomizer 7, an atomization spray pipe 8, and a valve 9. The atomized crosslinking agent solution forms an atomized spray through the atomizer, and the atomized spray enters the central pipe 12 through the atomization spray pipe 8. The surface of the hydrogel droplets crosslinks and solidifies to form microcapsules after contacting the crosslinking agent mist.

[0029] The vesicle receiving device includes an X-Y axis moving device 17 and a receiving orifice plate 16. The solidified hydrogel microcapsules continue to fall. The bioprinting ink pool module 13 pushes the crosslinkable hydrogel printing ink into the central pipe. The microcapsules are mixed with the bioprinting ink and fall into the receiving orifice plate below. The receiving orifice plate 16 can move along the x and y axes to achieve automated sample loading.

[0030] A UV curing device 15 is provided below the receiving orifice plate 16, which can cure the mixed bioprinting ink to fix the microcapsules in the photocrosslinked hydrogel.

[0031] A waste pipe 14 is provided below the atomization spray pipe 8. When the valve 9 is closed, the excess crosslinking agent spray is discharged through the waste pipe.

[0032] The method for preparing the vesicle fusion chip of the above preparation device includes the following preparation steps:

[0033] 1) Cells are mixed with the hydrogel solution in the hydrogel solution pool 1, and the hydrogel solution mixed with cells flows into the central pipe 12 through the injection head;

[0034] 2) The solution forms hydrogel droplets (i.e., vesicles) under the action of gas shearing at the inlet of the central pipe 12, and the hydrogel droplets fall from the central pipe 12;

[0035] 3) The atomized crosslinking agent solution forms an atomized spray through the atomizer 7. The valve 9 is opened, and the atomized spray enters the central pipe 12 through the atomization spray pipe 8. The surface of the hydrogel droplets crosslinks and solidifies to form microcapsules after contacting the crosslinking agent mist;

[0036] 4) The solidified hydrogel microcapsules continue to fall in the central pipe 12, and the bioprinting ink pool module 13 pushes the crosslinkable hydrogel printing ink into the central pipe. The microcapsules are mixed with the bioprinting ink and fall into the receiving orifice plate 16 below;

[0037] 5) The ultraviolet curing device 15 irradiates the receiving orifice plate 16 to crosslink and solidify the fused bioprinting ink, fixing the microcapsules in the photocrosslinked hydrogel.

[0038] Further, in the step 1), the separated tissue can be digested in the primary digestion chamber 18 first, and the digested primary cell suspension and the hydrogel solution meet and enter the hydrogel solution pool 1 for mixing.

[0039] Further, in the step 1), the flow rate of the hydrogel solution is controlled by the liquid controller 2, and the model of the injection head is adjusted by the injection head size converter 3.

[0040] Further, in the step 2), the flow rate of the gas is adjusted by the gas controller 11.

[0041] It should be understood that the present utility model is not limited to the specific embodiments of the present utility model described above, because changes can be made to the specific embodiments and still fall within the scope of the appended claims.

Claims

1. A vesicle fusion chip preparation device, characterized in that: It includes a vesicle generation device, a vesicle solidification device and a vesicle receiving device; the vesicle generation device includes a hydrogel solution pool (1), an injection device, a central pipeline (12) and a gas device. The injection device is arranged below the hydrogel solution pool (1), the injection device is arranged above the central pipeline (12), the central pipeline (12) is arranged above the vesicle receiving device, and the gas device is arranged at the entrance of the central pipeline; the vesicle solidification device includes an atomization device, a bio-ink pool module (13) and an ultraviolet light curing device (15). The atomization device is arranged below the gas device and connected to the central pipeline (12), the bio-ink pool module (13) is arranged below the atomization device and connected to the central pipeline (12), and the ultraviolet light curing device (15) is arranged above the vesicle receiving device.

2. The preparation device of a vesicle fusion chip according to claim 1, characterized in that: A primary digestion chamber (18) is connected to the hydrogel solution pool (1).

3. The preparation device of a vesicle fusion chip according to claim 1, wherein: The injection device includes an injection head size converter (3) and injection heads of different models.

4. The preparation device of a vesicle fusion chip according to claim 3, characterized in that: The injection heads of different models include a 32G injection head (4), a 34G injection head (5) and a 28G injection head (6).

5. The preparation device of a vesicle fusion chip according to claim 1, wherein: A liquid controller (2) is arranged on the hydrogel solution pool (1).

6. The preparation device of a vesicle fusion chip according to claim 1, characterized in that: The gas device includes a gas pipeline (19), a barometer (10) and a gas controller (11). The gas pipeline (19) is connected to the central pipeline (12), the barometer (10) is used to detect the gas pressure, and the gas controller (11) is used to adjust the gas flow rate.

7. The preparation device of a vesicle fusion chip according to claim 1, wherein: The atomization device includes an atomizer (7), an atomization spray pipe (8) and a valve (9). The atomized crosslinking agent solution forms an atomized spray through the atomizer, and the atomized spray enters the central pipeline (12) through the atomization spray pipe (8).

8. The preparation device of a vesicle fusion chip according to claim 1, characterized in that: The vesicle receiving device includes an X-Y axis moving device (17) and a receiving orifice plate (16). The receiving orifice plate (16) can move along the x and y axes, and the ultraviolet light curing device (15) is arranged above the receiving orifice plate (16).

9. The preparation device of a vesicle fusion chip according to claim 7, characterized in that: A waste discharge pipe (14) is arranged below the atomization spray pipe (8).

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