Microfluidic chip

Abstract

The application discloses a microfluidic chip, which comprises a substrate, a cover plate and a groove structure. The substrate is provided with a hydrogel channel and a culture fluid channel which are in communication with each other. The cover plate is arranged on the substrate and covers the hydrogel channel and the culture fluid channel. The communication part of the hydrogel channel and the culture fluid channel is divided by the groove structure. The groove can prevent the hydrogel in the hydrogel channel from flowing into the culture fluid channel, so that the hydrogel is limited in the hydrogel channel and the hydrogel is prevented from entering the culture fluid channel. The hydrogel channel is provided with the hydrogel, and the groove can form a relatively stable gel interface at the junction of the hydrogel channel and the culture fluid channel, so as to help the cells in the culture fluid channel to adhere to the gel interface and exchange nutrients through the hydrogel. A growth environment close to a biological body is constructed, so as to help to ensure the authenticity and effectiveness of the data research of the cell in vitro culture.

Description

[0001] This application claims priority to the Chinese patent application No. 2022109082800, filed on July 29, 2022, and entitled "Microfluidic chip", the content of which is incorporated herein by reference in its entirety. TECHNICAL FIELD

[0002] The present application relates to the field of biomedical technology, in particular to a microfluidic chip. BACKGROUND

[0003] Organ chip technology is a technology of performing in vitro three-dimensional cell culture in a chip. By constructing microchannels, microreaction chambers and other functional components, the components in the chip, such as cells, fluids, gases and extracellular microenvironment, are precisely controlled, so as to generate human microtissues and microorgans with biological functions. Organ chip technology has the advantages of integration, low consumption, high throughput, high simulation degree and fast analysis, and has broad application prospects in new drug research and development, disease modeling, personalized medicine and space medicine.

[0004] In the human body, nutrients and gases are transported to various parts of the body through blood vessels, and exchange of nutrients, gases and metabolites is carried out through the interface between organs and blood vessels. In order to simulate this structure in the human body, many organ chip technology researches use porous membranes to construct such an interface, and vascular endothelial cells and cells corresponding to organs are cultured on both sides of the porous membrane to simulate the cell arrangement and interface function of the organ and blood vessel interface. The addition of the porous membrane on the one hand puts high requirements on the properties and biocompatibility of the porous membrane itself, and the presence of the porous membrane also affects the free interaction and migration of cells between the channels, and on the other hand, the combination of the porous membrane in the chip is relatively complex. SUMMARY

[0005] The present application provides a microfluidic chip, which can enhance the restriction effect of the fluid in the microfluidic chip to improve the yield of the microfluidic chip.

[0006] The present application provides a microfluidic chip, comprising:

[0007] a substrate having a hydrogel channel and a culture fluid channel in communication with each other;

[0008] a cover plate covering the substrate and covering the hydrogel channel and the culture fluid channel;

[0009] a groove structure, the communication between the hydrogel channel and the culture fluid channel is divided by the groove structure, the groove structure has one or more grooves, and the groove is recessed at least with respect to the channel wall of the hydrogel channel;

[0010] wherein the groove is capable of blocking the flow of the hydrogel into the culture fluid channel after the hydrogel is injected into the hydrogel channel.

[0011] In some embodiments, the groove is recessed with respect to both the hydrogel channel and the channel wall of the culture fluid channel.

[0012] In some embodiments, the groove structure comprises a first groove structure disposed on the substrate and a second groove structure disposed on the cover plate, the first groove structure comprises one or more first grooves, the second groove structure comprises one or more second grooves, and the groove comprises the first groove and the second groove.

[0013] In some embodiments, the first groove and the second groove are arranged in alignment in the stacking direction of the cover plate and the substrate, or the first groove and the second groove are arranged in misalignment in the stacking direction of the cover plate and the substrate.

[0014] In some embodiments, the flow direction of the hydrogel in the hydrogel channel towards the culture fluid channel is a cross-flow direction of the hydrogel, and when each groove structure comprises a plurality of grooves, the plurality of grooves are arranged in intervals along the cross-flow direction, and adjacent two grooves in the same groove structure have a protrusion therebetween.

[0015] In some embodiments, the extension direction of the hydrogel channel itself is a longitudinal flow direction of the hydrogel, and the groove extends along the longitudinal flow direction or a plurality of grooves are arranged in intervals along the longitudinal flow direction.

[0016] In some embodiments, the groove is a rectangular slot, one slot wall surface of one groove is in contact with one channel wall of the hydrogel channel, and the contacting slot wall surface is arranged perpendicularly to the channel wall.

[0017] In some embodiments, the number of the hydrogel channel is one, the number of the culture fluid channel is two, the hydrogel channel is located between the two culture fluid channels, and the groove structure is arranged between each culture fluid channel and the hydrogel channel.

[0018] In some embodiments, the hydrogel channel comprises a substance exchange section and a hydrogel inlet section and a hydrogel outlet section connected to both ends of the substance exchange section, respectively, and the substance exchange section is in communication with the culture fluid channel.

[0019] The groove structure comprises two first groove structures arranged at intervals on the substrate and two second groove structures arranged at intervals on the cover plate, the two first groove structures and the two second groove structures define the substance exchange section, and the first groove structure comprises one or more first grooves, and the second groove structure comprises one or more second grooves.

[0020] In some embodiments, the substrate has a plurality of liquid inlet flow channels, a plurality of liquid inlet communication holes, a plurality of liquid outlet flow channels, and a plurality of liquid outlet communication holes, the flow cross section of the liquid inlet communication hole is smaller than the flow cross section of the liquid inlet flow channel, and the flow cross section of the liquid outlet communication hole is smaller than the flow cross section of the liquid outlet flow channel.

[0021] One end of each of the hydrogel channels communicates with a liquid inlet flow channel through a liquid inlet communication hole, and the other end of each of the hydrogel channels communicates with a liquid outlet flow channel through a liquid outlet communication hole.

[0022] One end of each of the culture fluid channels communicates with a liquid inlet flow channel through a liquid inlet communication hole, and the other end of each of the culture fluid channels communicates with a liquid outlet flow channel through a liquid outlet communication hole.

[0023] Based on the microfluidic chip provided in the embodiments of the present application, the communication position of the hydrogel channel and the culture fluid channel is divided by a groove structure, the groove can block the flow of the hydrogel into the culture fluid channel, thereby limiting the hydrogel in the hydrogel channel and preventing the hydrogel from flowing into the culture fluid channel; further, the hydrogel flows into the hydrogel channel, and the arrangement of the groove can also form a relatively stable gel interface at the junction position of the hydrogel channel and the culture fluid channel, thereby helping the cells in the culture fluid channel to adhere to the gel interface and enabling the exchange of nutrients through the hydrogel; so as to construct a growth environment close to the biological body, thereby facilitating the authenticity and effectiveness of the data research of the in vitro cell culture. BRIEF DESCRIPTION OF DRAWINGS

[0024] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed in the embodiments or the prior art description will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative labor.

[0025] Figure 1 The structure diagram of the microfluidic chip in one embodiment of the present application;

[0026] Figure 2A schematic diagram of a structure of a substrate in an embodiment of the present application;

[0027] Figure 3 A schematic diagram of a cross-section of a microfluidic chip in an embodiment of the present application;

[0028] Figure 4 Another schematic diagram of a cross-section of a microfluidic chip in an embodiment of the present application;

[0029] Figure 5 A schematic diagram of a cross-section of a microfluidic chip (with multiple grooves) in an embodiment of the present application;

[0030] Figure 6 A schematic diagram of a structure of a hydrogel acting between a cover plate and a substrate in an embodiment of the present application;

[0031] Figure 7 Another schematic diagram of a structure of a hydrogel acting between a cover plate and a substrate in an embodiment of the present application;

[0032] Figure 8 Still another schematic diagram of a structure of a hydrogel acting between a cover plate and a substrate in an embodiment of the present application;

[0033] Figure 9 A schematic diagram of a structure of a cover plate in an embodiment of the present application;

[0034] Figure 10 A flow chart of a method of using a microfluidic chip for a cell aggregate chip in an embodiment of the present application;

[0035] Figure 11 A flow chart of a method of using a microfluidic chip for a kidney cell chip in an embodiment of the present application.

[0036] Reference signs:

[0037] 10, substrate; 11, hydrogel channel; 111, substance exchange section; 1111, first substance exchange section; 1112, second substance exchange section; 112, hydrogel inlet section; 113, hydrogel outlet section; 12, culture fluid channel; 121, culture fluid inlet section; 122, culture fluid outlet section; 123, culture fluid channel section; 13, bulge;

[0038] 20, cover plate; 21, liquid inlet flow channel; 22, liquid inlet communication hole; 23, liquid outlet flow channel; 24, liquid outlet communication hole;

[0039] 31, first groove structure; 311, first groove; 32, second groove structure; 321, second groove;

[0040] H, horizontal flow direction; Z, vertical flow direction; D, stacking direction. DETAILED DESCRIPTION

[0041] In order to make the purposes, technical solutions and advantages of the present application clearer, the present application 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 intended to explain the present application and not to limit the present application.

[0042] In the human body, nutrients and gases are transported to various parts of the body through blood vessels, and nutrients, gases and metabolites are exchanged through the interface between organs and blood vessels. In order to simulate this structure in the human body, a porous membrane is used to construct such an interface in many organ chip technology researches, and blood vessel endothelial cells and cells corresponding to organs are cultured on both sides of the porous membrane to simulate the cell arrangement and interface function of the interface between the organs and the blood vessels. The addition of the porous membrane on one hand puts forward higher requirements for the properties and biocompatibility of the porous membrane itself, and the presence of the porous membrane also affects the free interaction and migration of cells between the channels, and on the other hand, the combination of the porous membrane in the chip is relatively complex.

[0043] In order to solve the above problems, please see Figures 1 to 3 The present application provides a microfluidic chip, which comprises a substrate 10, a cover plate 20 and a groove structure.

[0044] The substrate 10 has a hydrogel channel 11 and a culture fluid channel 12, which can be used to transport at least one of the extracellular microenvironment components such as cells, fluids, gases and drugs.

[0045] The culture fluid channel 12 can be one or two. When the culture fluid channel 12 is one, the culture fluid channel 12 is located on one side of the hydrogel channel 11, and the culture fluid channel 12 is in communication with the hydrogel channel 11. The hydrogel channel 11 is filled with hydrogel, and the culture fluid channel 12 is filled with culture fluid, such as culture medium.

[0046] It should be noted that the addition of hydrogel and culture fluid can be pump-free gravity-driven, or devices such as external peristaltic pumps and syringe pumps can be used to drive the flow of hydrogel and culture medium, so that the hydrogel and culture medium can provide the fluid shear force environment required for cell growth.

[0047] The cover plate 20 is located on the side of the substrate 10 close to the hydrogel channel 11 and the culture fluid channel 12, and covers the hydrogel channel 11 and the culture fluid channel 12, so that the hydrogel channel 11 and the culture fluid channel 12 form a channel shape with the middle closed and the two ends open, so that the hydrogel can flow in the hydrogel channel 11, and the culture medium can flow in the culture fluid channel 12.

[0048] The recess structure has a restriction effect on fluid, and the hydrogel is injected into the hydrogel channel 11, and the recess structure can block the hydrogel from flowing into the culture fluid channel 12, thereby limiting the hydrogel in the hydrogel channel 11; further, when the hydrogel solidifies in the hydrogel channel 11, the hydrogel forms a gel interface at the junction of the recess structure and the hydrogel channel 11, which provides a relatively suitable base hardness for cells to adhere to the gel interface. The cells grow in the hydrogel, and the nutrients in the culture medium can exchange substances with the hydrogel through the gel interface to meet the growth needs of the cells located in the hydrogel.

[0049] Please refer to Figure 2 When the culture fluid channel 12 is two, the two culture fluid channels 12 are located on opposite sides of the hydrogel channel 11. The recess structure is located between the hydrogel channel 11 and the culture fluid channel 12, and the hydrogel channel 11 is filled with hydrogel to form a three-layer structure of culture medium, hydrogel, and culture medium.

[0050] The recess structure has a restriction effect on fluid, and the hydrogel is injected into the hydrogel channel 11, and the recess structure can block the hydrogel from flowing into the culture fluid channel 12, thereby limiting the hydrogel in the hydrogel channel 11; further, when the hydrogel solidifies in the hydrogel channel 11, the hydrogel forms a gel interface at the junction of the recess structure and the hydrogel channel 11, which provides a relatively suitable base hardness for cells to adhere to the gel interface. The cells grow in the culture medium, and the cells can adhere to the gel interface, and the gel interface provides the base hardness required for cell growth, so as to facilitate uniform adhesion and distribution of cells on the gel interface; at the same time, the hydrogel channel 11 is located between the two culture fluid channels 12, different cells can be cultured in the two culture fluid channels 12, and the culture medium in the two culture fluid channels 12 can exchange substances through the hydrogel in the hydrogel channel 11 to meet the nutritional needs of the cells in the two different culture media.

[0051] Taking a kidney biological model as an example, one of the culture fluid channels 12 is filled with renal epithelial cells to make the renal epithelial cells adhere to the gel interface; and then the culture fluid channel 12 is filled with renal epithelial cell culture medium to provide nutrients required for the growth of renal epithelial cells. The other culture fluid channel 12 is filled with vascular endothelial cells to make the vascular endothelial cells adhere to the gel interface; and then the culture fluid channel 12 is filled with vascular endothelial cell culture medium to provide nutrients required for the growth of vascular endothelial cells. The renal epithelial cell culture medium and the vascular endothelial cell culture medium can exchange substances through the hydrogel, so that the renal epithelial cells can absorb the nutrients in the vascular endothelial cell culture medium.

[0052] Further, the gel interface can simulate the real interface in the organism, and the environment for cell growth is closer to the growth environment in the organism, so as to ensure the authenticity and effectiveness of the data research on the cell culture in vitro.

[0053] In some embodiments, referring to Figure 3 , the groove structure has a groove, the groove is recessed relative to the channel wall of the hydrogel channel 11, the groove is located between the hydrogel channel 11 and the culture fluid channel 12, the groove has a restriction on the fluid, the hydrogel is injected into the hydrogel channel 11, and the groove can block the hydrogel from flowing into the culture fluid channel 12; meanwhile, the hydrogel can form a gel interface at the junction of the groove and the hydrogel channel 11, so as to facilitate the uniform adhesion and distribution of cells on the gel interface, and to provide the cells with the base hardness and growth microenvironment required for cell growth.

[0054] The number of grooves can be one, the groove has a restriction on the fluid, and the hydrogel can be limited in the hydrogel channel 11; referring to Figure 5 , when the number of grooves is multiple, the grooves can have multiple blocking effects, and increasing the number of grooves can improve the stability of the hydrogel in the hydrogel channel 11.

[0055] Further, the groove can also be recessed relative to the channel walls of both the hydrogel channel 11 and the culture fluid channel 12, so that the groove is located between the hydrogel channel 11 and the culture fluid channel 12.

[0056] Further, referring to Figure 3 , the groove structure can include a first groove structure 31 arranged on the substrate 10 and a second groove structure 32 arranged on the cover plate 20; the first groove structure 31 includes a first groove 311, the second groove structure 32 includes a second groove 321, the groove includes the first groove 311 and the second groove 321, the groove has a restriction on the fluid, and the groove can block the hydrogel from flowing into the culture fluid channel 12, so that the groove has a better limiting effect.

[0057] In some embodiments, referring to Figure 5 , the first groove structure 31 can include multiple first grooves 311, the second groove structure 32 can also include multiple second grooves 321, and the multiple first grooves 311 and the multiple second grooves 321 can be arranged in sequence in a direction perpendicular to the extension direction of the hydrogel channel 11, the multiple first grooves 311 and the multiple second grooves 321 can have multiple blocking effects, and appropriately increasing the number of grooves can improve the stability of the hydrogel in the hydrogel channel 11.

[0058] Further, referring to Figure 3 in combination with Figure 1, the first groove 311 and the second groove 321 can be arranged opposite to each other in the stacking direction D of the cover plate 20 and the base plate 10. Since the grooves have a limiting effect on the fluid, the flow of the hydrogel at the opposite positions of the cover plate 20 and the base plate 10 can be limited, and the hydrogel can form a nearly straight gel interface at the intersection of the first groove 311 and the second groove 321 in the hydrogel channel 11, so as to facilitate the uniform adhesion and distribution of cells on the gel wall. The hydrogel interface simulates the real interface in the organism, so that the cell growth environment is closer to the growth environment in the organism, thereby facilitating the authenticity and effectiveness of the cell culture data research.

[0059] In some embodiments, referring to Figure 4 In combination Figure 1 , the first groove 311 and the second groove 321 can also be arranged in a staggered manner in the stacking direction D of the cover plate 20 and the base plate 10. By using the limiting effect of the grooves on the hydrogel, a gel interface can be formed at the intersection of the first groove 311 and the second groove 321 in the hydrogel channel 11, thereby preventing the hydrogel from flowing from the hydrogel channel 11 to the culture fluid channel 12. Since the cells adhere to the gel interface, the limiting effect of the grooves can make the hydrogel form a relatively stable gel interface, thereby providing a relatively stable base hardness for cell adhesion, facilitating cell growth. Moreover, the arrangement of the gel interface also facilitates precise control of the pressure in the hydrogel channel 11 and the culture fluid channel 12, thereby providing a growth environment for cells that is closer to the growth environment in the organism, thereby facilitating the authenticity and effectiveness of the cell culture data research.

[0060] Referring to Figure 5 , further, the flow direction of the hydrogel in the hydrogel channel 11 to the culture fluid channel 12 is the cross-flow direction H of the hydrogel. When each groove structure includes a plurality of grooves, the plurality of grooves are arranged in the cross-flow direction H, and adjacent two grooves in the same groove structure have protrusions, and the adjacent protrusions are grooves. The grooves have a limiting effect on the fluid, thereby preventing the hydrogel from flowing to the culture fluid channel 12, and further ensuring the stability of the hydrogel in the hydrogel channel 11.

[0061] Further, the extension direction of the hydrogel channel 11 itself is the longitudinal flow direction Z of the hydrogel, and the grooves extend in the longitudinal flow direction or a plurality of grooves are arranged in the longitudinal flow direction. The extension direction of the plurality of grooves is consistent with the extension direction of the hydrogel channel 11, and the grooves have a limiting effect on the fluid. The plurality of grooves can limit the hydrogel in the longitudinal flow direction, and also improve the stability of the hydrogel in the hydrogel channel 11, so as to form a relatively stable gel interface, facilitate cell adhesion to the gel interface, and also facilitate the construction of a growth environment that is closer to the real growth environment in the organism.

[0062] Further, the groove is a rectangular groove, one groove wall of one groove is in contact with one channel wall of the hydrogel channel 11, and the contact groove wall is arranged perpendicularly to the channel wall, so that the contact angle is formed at the connection position of the groove and the hydrogel channel 11, and then the gel interface is formed at the junction position of the groove and the hydrogel channel 11, and the channel wall of the hydrogel channel 11 is arranged perpendicularly to the groove wall, so as to increase the restriction effect of the groove on the hydrogel in the hydrogel channel 11.

[0063] When the hydrogel is injected into the hydrogel channel 11 and spreads, please refer to Figure 6 , satisfies Young's equation: γ sg- γ sl = γ lg cos θ, where γ sg is the solid-gas interfacial tension, γ sl is the solid-liquid interfacial tension, γ lg is the liquid-gas interfacial tension, and θ is the contact angle of the liquid on the solid surface.

[0064] When the hydrogel is in contact with the surface of the cover plate 20, it satisfies: γ s1g- γ s1l = γ lg cos θ1, where γ s1g is the solid-gas interfacial tension of the surface of the cover plate 20, γ s1l is the solid-liquid interfacial tension of the surface of the cover plate 20, γ lg is the liquid-gas interfacial tension, and θ1 is the contact angle of the liquid on the surface of the cover plate 20. When the hydrogel is in contact with the surface of the substrate 10, it satisfies: γ s2g- γ s2l = γ lg cos θ2, where γ s2g is the solid-gas interfacial tension of the surface of the substrate 10, γ s2l is the solid-liquid interfacial tension of the surface of the substrate 10, γ lg is the liquid-gas interfacial tension, and θ2 is the contact angle of the liquid on the surface of the substrate 10.

[0065] According to the Laplace pressure difference: In the formula, R1 is the principal curvature radius of the liquid side curved surface in the plane perpendicular to the flat plate, and R2 is the principal curvature radius of the liquid side curved surface in the plane parallel to the flat plate. For the microfluidic chip structure, R2 = 0, R1 is related to the channel height (the distance between the opposite surfaces of the cover plate 20 and the substrate 10) h, and according to the geometric relationship, h = R1 cos θ1 + R1 cos θ2, where θ1 is the contact angle of the hydrogel with the surface of the cover plate 20, and θ2 is the contact angle of the hydrogel on the surface of the substrate 10. Bringing it into the above pressure difference formula can obtain: When the hydrogel reaches the edge, the characteristic angle θ edge , In the formula: is a geometric boundary angle. θ1or θ2in the formula is replaced by θ edge . Figure 6 In the formula

[0066] Figure 7 In the formula γ lg＝ 72 mN / m, h = 200 μm, θ1= 80°, θ edge = 170°, only when the groove is arranged on the substrate, the pressure difference is calculated as ΔP = -291 Pa, and the relative air pressure P1 in the hydrogel is 291 Pa.

[0067] Figure 8 In the formula γ lg＝ 72 mN / m, h = 200 μm, θ edge1 = 170°, θ edge2 = 170°, when the grooves are arranged on the substrate and the cover plate, the pressure difference is calculated as ΔP = -709 Pa. At this time, the relative air pressure P2 in the hydrogel is 709 Pa.

[0068] From the above calculation, it can be known that P2 > P1, when the grooves are arranged on the substrate 10 and the cover plate 20, the hydrogel can withstand a larger air pressure in the interior, and it can be known that the limiting effect when the grooves are arranged on both the substrate 10 and the cover plate 20 is better than the limiting effect when the grooves are arranged only on the substrate 10.

[0069] Therefore, when a pressure of 500 Pa is applied in the interior of the hydrogel, the grooves arranged on the substrate 10 and the cover plate 20 can withstand a larger pressure, and thus when 500 Pa is applied in the interior of the hydrogel, the hydrogel can still be limited in the hydrogel channel 11.

[0070] In some embodiments, when the groove arranged between the hydrogel channel 11 and the culture fluid channel 12 is one, the distance between the opposite surfaces of the cover plate 20 and the substrate 10 is greater than or equal to 0 μm and less than or equal to 500 μm, the width of the groove is greater than or equal to 100 μm and less than or equal to 400 μm, and the depth of the groove is greater than or equal to 50 μm and less than or equal to 400 μm, under the condition that the grooves are arranged on the substrate 10 and the cover plate 20, the grooves have a better blocking effect on the hydrogel located in the hydrogel channel 11.

[0071] When the grooves between the hydrogel channel 11 and the culture fluid channel 12 are multiple, the distance between the opposite surfaces of the cover plate 20 and the substrate 10 is greater than or equal to 0 μm and less than or equal to 500 μm, the width of the grooves is greater than or equal to 100 μm and less than or equal to 400 μm, the depth of the grooves is greater than or equal to 50 μm and less than or equal to 400 μm, and the number of the grooves is not less than two, so that the grooves have a relatively stable and effective liquid blocking effect on the hydrogel in the hydrogel channel 11.

[0072] Further, referring to Figure 2 , the hydrogel channel 11 comprises a substance exchange section 111 and a hydrogel inlet section 112 and a hydrogel outlet section 113 connected to both ends of the substance exchange section 111, and the substance exchange section 111 is in communication with the culture fluid channel 12.

[0073] The substance exchange section 111 can be provided as a straight channel, and the hydrogel inlet section 112 and the hydrogel outlet section 113 can also be provided as straight channels and in communication with the substance exchange section 111 to form the hydrogel channel 11 from the hydrogel inlet section 112 to the substance exchange section 111 and then to the hydrogel outlet section 113, so as to facilitate the flow of the hydrogel from the hydrogel inlet section 112 to the substance exchange section 111 for the cells in the culture fluid channel 12 on one side or both sides of the substance exchange section 111 to adhere.

[0074] Meanwhile, the central axes of the hydrogel inlet section 112, the hydrogel outlet section 113 and the substance exchange section 111 can be provided on the same straight line to set the hydrogel inlet section 112 and the hydrogel outlet section 113 at a more appropriate position to facilitate the addition of the hydrogel and the collection of the excess hydrogel. In some embodiments, the hydrogel inlet section 112 and the hydrogel outlet section 113 can also be provided with an included angle with the substance exchange section 111, which needs to be set according to the actual shape and size of the substrate 10 and the cover plate 20, and the present application does not make any limitation. It should be noted that the hydrogel inlet section 112, the hydrogel outlet section 113 and the substance exchange section 111 are provided as straight channels, which can reduce the flow resistance of the hydrogel in the hydrogel channel 11 to facilitate the flow of the hydrogel.

[0075] In some embodiments, the hydrogel inlet section 112, the hydrogel outlet section 113 and the substance exchange section 111 can also have other shapes, which can be set according to actual needs, and the present application does not make any limitation.

[0076] In some embodiments, referring to Figures 1 to 3The groove structure comprises two first groove structures 31 arranged at intervals on the substrate 10 and two second groove structures 32 arranged at intervals on the cover plate 20, the two first groove structures 31 and the two second groove structures 32 define a substance exchange section 111, and the first groove structure 31 comprises one or more first grooves 311, and the second groove structure 32 comprises one or more second grooves 321, the groove comprises the first groove 311 and the second groove 321.

[0077] Please refer to Figure 2 The substance exchange section 111 comprises a first substance exchange section (not shown in the figure) and a second substance exchange section (not shown in the figure), the first substance exchange section is close to the substrate 10, and the second substance exchange section is close to the cover plate 20. The two first grooves 311 arranged at intervals on the substrate 10 define the first substance exchange section, the groove has a limiting effect on the hydrogel, so that the groove can limit the hydrogel in the first substance exchange section; the two second grooves 321 arranged at intervals on the cover plate 20 define the second substance exchange section, the groove has a limiting effect on the hydrogel, so that the groove can limit the hydrogel in the second substance exchange section; the two first grooves 311 and the two second grooves 321 jointly limit the hydrogel in the hydrogel channel 11, preventing the hydrogel from entering the culture fluid channel 12. Further, due to the joint limitation of the first groove 311 and the second groove 321, a relatively stable gel interface is formed at the junction of the groove and the hydrogel channel 11, which helps the cells in the culture fluid channel 12 to exchange substances with the nutrients in the hydrogel; at the same time, the relatively stable gel interface facilitates the adhesion of cells, so as to be closer to the growth environment of the organism, thereby helping to ensure the authenticity and effectiveness of the data research of the in-vitro cell culture.

[0078] In some embodiments, the first groove structure 31 can comprise a plurality of first grooves 311, the second groove structure 32 can comprise a plurality of second grooves 321, the first groove 311 and the second groove 321 cooperate to form a groove, and the plurality of grooves are arranged in the cross-flow direction H of the hydrogel. The plurality of grooves can play a multiple blocking role, and appropriately increasing the number of grooves can prevent the hydrogel from entering the culture fluid channel 12 from the hydrogel channel 11, thereby improving the stability of the hydrogel in the hydrogel channel 11.

[0079] Please refer to Figure 2, the culture fluid channel 12 may also include a culture fluid channel segment 123, a culture fluid inlet segment 121 and a culture fluid outlet segment 122 located at both ends of the culture fluid channel segment 123, so as to form a culture fluid channel 12 from the culture fluid inlet segment 121 to the culture fluid channel segment 123 to the culture fluid outlet segment 122, facilitating the addition of the culture medium from the culture fluid inlet segment 121 into the culture fluid channel 12 to provide nutrients for cell growth.

[0080] The culture fluid channel 12 is located on one side of the hydrogel channel 11. The culture fluid inlet segment 121 and the culture fluid outlet segment 122 can be set as straight channels, and there is an included angle between them and the culture fluid channel segment 123, so as to set the liquid addition port and the liquid outlet of the culture medium at appropriate positions on the substrate 10 and the cover plate 20, facilitating the operator to add the culture medium.

[0081] Please refer to Figure 2 , the positions of the culture fluid channel 12 and the hydrogel channel 11 can be set in a shape like the Chinese character "chuan". The culture fluid channel 12 composed of the culture fluid inlet segment 121, the culture fluid channel segment 123 and the culture fluid outlet segment 122 is set to be concave, so as to set the liquid addition port of the culture fluid inlet segment 121 and the liquid outlet of the culture fluid outlet segment 122 at appropriate positions, facilitating the addition of the culture medium and the collection of the excess culture medium.

[0082] Furthermore, please refer to Figure 9 Refer to Figure 1 , the substrate 10 has a plurality of liquid inlet channels 21, a plurality of liquid inlet communication holes 22, a plurality of liquid outlet channels 23 and a plurality of liquid outlet communication holes 24. The flow cross-section of the liquid inlet communication hole is smaller than that of the liquid inlet channel 21, and the flow cross-section of the liquid outlet communication hole 24 is smaller than that of the liquid outlet channel 23.

[0083] One end of each hydrogel channel 11 is connected to a liquid inlet channel 2 through a liquid inlet communication hole 22, and the other end of each hydrogel channel 11 is connected to a liquid outlet channel 23 through a liquid outlet communication hole 24;

[0084] The hydrogel channel 11 comprises a hydrogel inlet section 112, a hydrogel outlet section 113 and a substance exchange section 111. The liquid inlet channel 21 corresponds to the liquid inlet of the hydrogel inlet section 112. The liquid inlet channel 21 is provided with a liquid inlet communication hole 22 at a side close to the hydrogel inlet section 112. The flow cross section of the liquid inlet communication hole 22 is smaller than that of the liquid inlet channel 21. The liquid inlet channel 21 with a large cross section is convenient for the outlet end of the pipette to enter, which helps the hydrogel in the pipette to enter the hydrogel inlet section 112 through the liquid inlet communication hole 22. Further, the outlet end of the pipette is placed in the liquid inlet channel 21, which can prevent the hydrogel in the pipette from being contaminated. The hydrogel inlet section 112 is provided with an enlarged portion 13 at a position close to the liquid inlet communication hole 22, so that the hydrogel in the liquid inlet communication hole 22 can enter the hydrogel inlet section 112 completely.

[0085] The liquid outlet channel 23 corresponds to the hydrogel outlet section 113. The liquid outlet channel 23 is provided with a liquid outlet communication hole 24 at a side close to the hydrogel outlet section 113. The flow cross section of the liquid outlet communication hole 24 is smaller than that of the liquid outlet channel 23. The liquid outlet communication hole 24 with a small cross section is convenient for connecting the liquid outlet channel 23 and the hydrogel outlet section 113, so that the excess hydrogel in the hydrogel outlet section 113 can be discharged to the liquid outlet channel 23. Meanwhile, the liquid outlet channel 23 with a large cross section can store more hydrogel. When the amount of the hydrogel in the liquid outlet channel 23 reaches a certain amount, the operator can remove it, so as to improve the removal efficiency of the excess hydrogel.

[0086] The hydrogel outlet section 113 is provided with an enlarged portion 13 at a position close to the liquid outlet communication hole 24, so that the excess hydrogel can be gathered in the enlarged portion 13, and then the excess hydrogel can enter the liquid outlet channel 23 through the liquid outlet communication hole 24.

[0087] Each culture fluid channel 12 is communicated with a liquid inlet channel 21 through a liquid inlet communication hole 22 at one end, and communicated with a liquid outlet channel 23 through a liquid outlet communication hole 24 at the other end.

[0088] The culture fluid channel 12 comprises a culture fluid channel segment 123 and a culture fluid inlet segment 121 and a culture fluid outlet segment 122 located at both ends of the culture fluid channel segment 123; the liquid inlet flow channel 21 corresponds to the liquid inlet of the culture fluid inlet segment 121, and the liquid inlet flow channel 21 is provided with a liquid inlet communication hole 22 on the side close to the culture fluid inlet segment 121; the flow cross section of the liquid inlet communication hole 22 is smaller than that of the liquid inlet flow channel 21, so as to add the culture medium in the liquid inlet flow channel 21 to the culture fluid channel 12 through the liquid inlet communication hole 22, while the liquid inlet flow channel 21 with a larger cross section facilitates the liquid outlet end of the pipette to enter, thereby helping the culture medium in the pipette to enter the culture fluid channel 12 through the liquid inlet communication hole 22, and further, placing the liquid outlet end of the pipette in the liquid inlet flow channel 21 can also prevent the culture medium in the pipette from being contaminated.

[0089] Meanwhile, the culture fluid inlet segment 121 is provided with an enlarged portion 13 at the position close to the liquid inlet communication hole 22, so that the culture medium in the liquid inlet communication hole 22 can all enter the culture fluid inlet segment 121.

[0090] The liquid outlet flow channel 23 corresponds to the culture fluid outlet segment 122, and the liquid outlet flow channel 23 is provided with a liquid outlet communication hole 24 on the side close to the culture fluid outlet segment 122; the flow cross section of the liquid outlet communication hole 24 is smaller than that of the liquid outlet flow channel 23, and the liquid outlet communication hole 24 with a smaller cross section facilitates the connection of the liquid outlet flow channel 23 and the culture fluid outlet segment 122, thereby allowing the excess culture medium in the culture fluid outlet segment 122 to be discharged to the liquid outlet flow channel 23; meanwhile, the liquid outlet flow channel 23 with a larger cross section can store more excess culture medium, and the operator can remove the excess culture medium when the amount of the excess culture medium in the liquid outlet flow channel 23 reaches a certain amount, so as to improve the removal efficiency of the excess culture medium.

[0091] Meanwhile, the culture fluid outlet segment 122 is provided with an enlarged portion 13 at the position close to the liquid outlet communication hole 24, so as to converge the excess culture medium to the enlarged portion 13, and then pass the excess culture medium to the liquid outlet flow channel 23 through the liquid outlet communication hole 24.

[0092] Please refer to Figure 10 When the number of the culture fluid channels 12 is one, the use method of the microfluidic chip for the cell aggregate chip comprises the following steps:

[0093] S11, assembling the cover plate 20 and the base plate 10 to form a microfluidic chip.

[0094] S12, sterilizing the microfluidic chip.

[0095] S13, add the mixed cell aggregate hydrogel into the hydrogel channel 11, and the groove can limit the mixed cell aggregate hydrogel in the hydrogel channel 11, and the excess hydrogel flows out from the outflow channel 23 through the outflow communication hole 24.

[0096] S14, after the mixed cell aggregate hydrogel is solidified, a mixed cell aggregate hydrogel strip is formed.

[0097] S15, add the cell aggregate culture medium from the inflow channel 21, and the excess culture medium flows out from the outflow channel 23 through the outflow communication hole 24.

[0098] S16, after the cell aggregate culture is completed, subsequent detection is performed.

[0099] Please refer to Figure 11 When the number of the culture fluid channels 12 is two, the use method of the microfluidic chip in the embodiment for the kidney cell chip comprises:

[0100] S21, assemble the cover plate 20 and the base plate 10 to form the microfluidic chip.

[0101] S22, sterilize the microfluidic chip.

[0102] S23, add the Matrigel / collagen / fibrin mixed hydrogel into the hydrogel channel 11, and the groove can limit the Matrigel / collagen / fibrin mixed hydrogel in the hydrogel channel 11, and the excess Matrigel / collagen / fibrin mixed hydrogel flows out from the outflow channel 23 through the outflow communication hole 24.

[0103] S24, after the Matrigel / collagen / fibrin mixed hydrogel is solidified, a Matrigel / collagen / fibrin mixed hydrogel strip is formed.

[0104] S25, add the renal epithelial cells into the culture fluid channel 12 from the inflow channel 21, so that the renal epithelial cells adhere to the gel interface, add the renal epithelial cell culture medium from the inflow channel 21, and the excess culture medium flows out from the outflow channel 23 through the outflow communication hole 24.

[0105] S26, add the vascular endothelial cells into the culture fluid channel 12 from the inflow channel 21, so that the vascular endothelial cells adhere to the gel interface, add the vascular endothelial cell culture medium from the inflow channel 21, and the excess culture medium flows out from the outflow channel 23 through the outflow communication hole 24.

[0106] S27、After the cell culture is successful, the kidney-endothelial organ model is constructed, which is used for subsequent tests.

[0107] The same or similar reference numerals in the drawings of the embodiments correspond to the same or similar components; in the description of the present application, it is understood that if the orientations or positional relationships indicated by the terms "upper", "lower", "left", "right" and the like are based on the orientations or positional relationships shown in the drawings, they are only for the convenience of describing the present application and simplifying the description, and do not indicate or imply that the devices or elements referred to must have a particular orientation, be constructed and operated in a particular orientation, therefore the terms describing the positional relationships in the drawings are only used for exemplary illustration, and cannot be understood as a limitation on the present patent, and for those of ordinary skill in the art, the specific meanings of the above terms can be understood according to the specific circumstances.

[0108] The above is only a preferred embodiment of the present application, and is not used to limit the present application, any modification, equivalent replacement and improvement made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A microfluidic chip, characterized by, The microfluidic chip comprises: a substrate, having a hydrogel channel and a culture fluid channel which are in communication with each other; a cover plate, covering the substrate and covering the hydrogel channel and the culture fluid channel; a groove structure, delimiting the communication between the hydrogel channel and the culture fluid channel, the groove structure having a plurality of grooves, the grooves being recessed at least with respect to the channel wall of the hydrogel channel, and the grooves being recessed with respect to the channel walls of the hydrogel channel and the culture fluid channel at the same time; wherein the flow direction of the hydrogel in the hydrogel channel to the culture fluid channel is the cross-flow direction of the hydrogel, when each groove structure comprises a plurality of grooves, the plurality of grooves are arranged in intervals along the cross-flow direction, and there is a protrusion between any two adjacent grooves in the same groove structure, and the grooves can block the flow of the hydrogel into the culture fluid channel after the hydrogel is injected into the hydrogel channel.

2. The microfluidic chip of claim 1, wherein, The groove structure comprises a first groove structure arranged on the substrate and a second groove structure arranged on the cover plate, the first groove structure comprises a plurality of first grooves, and the second groove structure comprises a plurality of second grooves, and the grooves comprise the first grooves and the second grooves.

3. The microfluidic chip of claim 2, wherein, The first grooves and the second grooves are arranged in a direction perpendicular to the stacking direction of the cover plate and the substrate, or the first grooves and the second grooves are arranged in a staggered manner in the stacking direction of the cover plate and the substrate.

4. The microfluidic chip of claim 1, wherein, The extension direction of the hydrogel channel itself is the longitudinal flow direction of the hydrogel, and the grooves extend along the longitudinal flow direction or a plurality of grooves are arranged in intervals along the longitudinal flow direction.

5. The microfluidic chip of claim 1, wherein, The grooves are rectangular grooves, one groove wall of one groove is in contact with one channel wall of the hydrogel channel, and the groove wall and the channel wall are arranged perpendicularly.

6. The microfluidic chip of claim 1, wherein, The number of the hydrogel channels is one, the number of the culture fluid channels is two, and the hydrogel channel is located between the two culture fluid channels, and the groove structure is arranged between each culture fluid channel and the hydrogel channel.

7. The microfluidic chip according to claim 6, wherein the hydrogel channel comprises a substance exchange section and a hydrogel inlet section and a hydrogel outlet section connected to both ends of the substance exchange section respectively, and the substance exchange section is in communication with the culture fluid channel; the groove structure comprises two first groove structures arranged in intervals on the substrate and two second groove structures arranged in intervals on the cover plate, the two first groove structures and the two second groove structures define the substance exchange section, and the first groove structure comprises a plurality of first grooves, and the second groove structure comprises a plurality of second grooves, and the grooves comprise the first grooves and the second grooves.

8. The microfluidic chip according to any one of claims 1-7, wherein, The cover plate has a plurality of liquid inlet flow channels, a plurality of liquid inlet communication holes, a plurality of liquid outlet flow channels, and a plurality of liquid outlet communication holes, the flow cross section of the liquid inlet communication hole is smaller than the flow cross section of the liquid inlet flow channel, and the flow cross section of the liquid outlet communication hole is smaller than the flow cross section of the liquid outlet flow channel. One end of each of the hydrogel channels is communicated with the liquid inlet channel through a liquid inlet communication hole, and the other end of each of the hydrogel channels is communicated with the liquid outlet channel through a liquid outlet communication hole; One end of each of the culture fluid channels is communicated with the liquid inlet channel through a liquid inlet communication hole, and the other end of each of the culture fluid channels is communicated with the liquid outlet channel through a liquid outlet communication hole.

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