Embedded well chip for three-dimensional cell culture

Abstract

This utility model relates to a porous chip for three-dimensional cell culture, belonging to the field of three-dimensional cell culture technology. It includes an outer aperture, an inner aperture, and a substrate. The inner aperture is embedded within the outer aperture, and the outer aperture, inner aperture, and substrate form an outer receiving cavity. The inner aperture and substrate form an inner receiving cavity, and the bottoms of the outer receiving cavity and the inner receiving cavity are connected. This utility model discloses a porous chip for three-dimensional cell culture, which creatively solves the systemic defects of traditional Transwell chambers.

Description

Technical Field

[0001] This invention relates to the field of three-dimensional cell culture technology, and in particular to a perforated chip for three-dimensional cell culture. Background Technology

[0002] Three-dimensional cell culture technology, as an important tool in modern biomedical research, is mainly used to simulate the in vivo microenvironment and plays a crucial role in drug screening, disease model construction, and tissue engineering. Currently, the mainstream three-dimensional cell culture devices mainly adopt the traditional Transwell chamber structure, which uses a porous membrane (typically with a pore size of 0.4-8 μm) to separate the upper and lower chambers for substance exchange. However, based on laboratory validation and clinical feedback, this structure has the following systemic drawbacks:

[0003] Functional deficiency: The porous membrane structure cannot simultaneously achieve stable gel support and cross-regional material exchange, resulting in difficulties in cell-gel co-culture and a 40%-60% reduction in macromolecular diffusion efficiency.

[0004] Sealing defects: The seal fails when used in ordinary petri dishes, with a liquid leakage rate of >30%, necessitating the use of special consumables. Utility Model Content

[0005] To overcome the shortcomings of existing technologies, this invention proposes a perforated chip for three-dimensional cell culture, which can creatively solve the problem of the systematic defects of traditional Transwell chambers.

[0006] To achieve this objective, the present invention adopts the following technical solution:

[0007] This utility model provides a perforated chip for three-dimensional cell culture, including an outer hole, an inner hole, and a substrate. The inner hole is embedded in the outer hole. The outer hole, the inner hole, and the substrate constitute an outer receiving cavity, and the inner hole and the substrate constitute an inner receiving cavity. The bottoms of the outer receiving cavity and the inner receiving cavity are connected.

[0008] This invention provides a perforated chip for three-dimensional cell culture, wherein the outer and inner holes are cylindrical and located on the same central axis.

[0009] This invention provides a perforated chip for three-dimensional cell culture, wherein a connecting rib is provided between the inner side of the outer hole and the outer side of the inner hole to connect the two.

[0010] This invention provides a perforated chip for three-dimensional cell culture, wherein the height of the connection between the bottom of the outer cavity and the inner cavity is 0.1-3mm.

[0011] This invention provides a perforated chip for three-dimensional cell culture, wherein the ratio of the diameter of the outer hole to the diameter of the inner hole is (3-5):1.

[0012] This invention provides a porous chip for three-dimensional cell culture, wherein the surface of the substrate is provided with a first layer to increase surface tension.

[0013] This invention provides a perforated chip for three-dimensional cell culture, wherein the outer and inner pores are provided with a second layer to increase surface tension.

[0014] This invention provides a perforated chip for three-dimensional cell culture, wherein the bottom of the substrate is provided with an optically transparent polymer or glass cover.

[0015] This invention provides a porous chip for three-dimensional cell culture, wherein the porous chip has a porous array structure.

[0016] The beneficial effects of this utility model are:

[0017] This invention proposes a porous chip for three-dimensional cell culture. In this embodiment, the bottoms of the outer and inner cavities of the porous chip are connected, forming a vertically nested and interconnected dual-cavity structure. This structural design has significant functional integration advantages: First, a fixed amount of gel can be injected into the bottom of the inner cavity according to experimental needs and stably supported, effectively reducing the amount of gel used and lowering experimental costs, while ensuring uniform gel thickness, which is beneficial for imaging quality control; second, the culture medium is contained in the outer cavity and connected to the bottom of the inner cavity to form a liquid gradient interface, thus achieving barrier-free material exchange without membrane separation. This invention effectively solves the systemic problems of functional deficiencies and sealing defects in the original Transwell chamber. Attached Figure Description

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

[0019] Figure 1 This is a three-dimensional view of a perforated chip for three-dimensional cell culture according to Example 1;

[0020] Figure 2 This is a top view of a perforated chip for three-dimensional cell culture according to Embodiment 1;

[0021] Figure 3 for Figure 2 A sectional view in the AA direction.

[0022] In the picture:

[0023] 1-Outer hole; 2-Inner hole; 3-Base; 4-Outer cavity; 5-Inner cavity; 6-Connecting rib; 7-Glass cover. Detailed Implementation

[0024] The technical solution of this utility model will be further described below with reference to the accompanying drawings and specific embodiments.

[0025] Example 1

[0026] like Figure 1-3 As shown, this embodiment provides a porous chip for three-dimensional cell culture, comprising an outer hole 1, an inner hole 2, and a substrate 3. The inner hole 2 is embedded within the outer hole 1. The outer hole 1, inner hole 2, and substrate 3 constitute an outer receiving cavity 4, and the inner hole 2 and substrate 3 constitute an inner receiving cavity 5. The bottoms of the outer receiving cavity 4 and the inner receiving cavity 5 are connected. This invention proposes a porous chip for three-dimensional cell culture, which, through innovative structural design, systematically solves several technical defects of traditional Transwell chambers in three-dimensional cell culture applications. In this embodiment, the bottoms of the outer receiving cavity and the inner receiving cavity are connected, forming a vertically nested and interconnected dual-cavity structure. This structural design has significant functional integration advantages: First, a quantitative amount of gel can be injected into the bottom of the inner hole according to experimental needs and stably supported, effectively reducing the amount of gel used, lowering experimental costs, and ensuring consistent gel thickness, which is beneficial for imaging quality control; second, the culture medium is contained in the outer hole and connected to the bottom of the inner hole to form a liquid gradient interface, thereby achieving barrier-free material exchange without membrane separation, overcoming the problem of low diffusion efficiency of large molecules due to the limited pore size of traditional membranes. The overall chip structure adopts an integrated design, avoiding the dependence on dedicated scaffolds and culture dishes required by traditional Transwells, thus improving operational convenience and reducing the risk of membrane damage. In summary, this invention creatively achieves cell-gel co-culture through a dual-well vertical nested structure and multi-dimensional structural optimization of the bottom liquid-passing interface, thereby breaking through the systemic bottlenecks of traditional Transwell chambers in terms of function, efficiency, and applicability.

[0027] Preferably, the outer hole 1 and the inner hole 2 are cylindrical, and the outer hole 1 and the inner hole 2 are located on the same central axis. In this preferred embodiment, the cylindrical structure has axisymmetric geometric characteristics, which can ensure that the gel matrix and culture medium are uniformly distributed in the vertical direction, avoiding dead corners and stagnation zones caused by asymmetrical structures, thereby improving the material exchange efficiency and the hydrodynamic performance of the culture medium; secondly, the coaxial design of the outer hole and the inner hole can achieve accurate part positioning, which is beneficial for mold casting and mass production.

[0028] Specifically, to achieve the goal of the inner hole 2 being embedded within the outer hole 1 and the bottoms of the outer receiving cavity 4 and the inner receiving cavity 5 being connected, a connecting rib plate 6 is provided between the inner side of the outer hole 1 and the outer side of the inner hole 2. In this embodiment, two symmetrically distributed connecting rib plates 6 are provided between the inner side of the outer hole 1 and the outer side of the inner hole 2. This symmetrically distributed connecting rib plate can enhance the mechanical strength of the structure, prevent the inner hole from slightly shifting or tilting relative to the outer hole due to improper operation or external force, and can also effectively limit the filling area of ​​the matrix gel and cell suspension, avoiding leakage of the gel between the pore walls.

[0029] Preferably, the height h at the bottom connection point between the outer cavity 4 and the inner cavity 5 is 0.1-3 mm. This range of connection height ensures open material exchange between the two chambers while maintaining a sufficient surface tension barrier to prevent cross-contamination during inversion, tilting, or shaking experiments. Secondly, this size serves as a microchannel to ensure restricted molecular diffusion paths, allowing researchers to further refine flux and gradient distribution by changing the matrix gel formulation, without significantly increasing manufacturing complexity due to excessive thinness. In this embodiment, 0.5 mm is used.

[0030] Preferably, the ratio of the diameter of the outer aperture 1 to the diameter of the inner aperture 2 is (3-5):1, where the diameter refers to the inner diameter. In this embodiment, the diameter of the outer aperture 1 is selected as 12 mm, and the diameter of the inner aperture 2 is selected as 3 mm. The 12 mm outer aperture diameter is compatible with commercially available microscope stages and most well plate slots, requiring no additional support modifications, while the 3 mm inner aperture size perfectly matches the optimal range for cell clumping and gel thickness control, improving the resolution of three-dimensional imaging.

[0031] Preferably, the surface of the substrate 3 is provided with a first surface tension-enhancing layer. Firstly, the surface-modified substrate layer significantly improves the stability of the gel and cell suspension on the surface, allowing them to quickly adhere to the substrate after injection without slippage or gel runoff, and maintaining colloidal stability during continuous tilting tests. Secondly, this tension-enhancing layer can be formed by spraying a fluorinated or silicon-containing surfactant polymer film onto the substrate surface after plasma treatment, combined with UV cross-linking curing, to create a durable and chemically sterile hydrophilic-hydrophobic coexistence interface. Similarly, the outer pore 1 and inner pore 2 are further provided with a second surface tension-enhancing layer. This second tension-enhancing layer can form a stable interfacial film at the interface between the pore wall and the culture medium or gel, preventing capillary loss of liquid along the wall surface, thereby effectively avoiding leakage when the carrier is tilted, shaken, or the culture medium is changed.

[0032] Preferably, the bottom of the substrate 3 is provided with an optically transparent polymer or glass cover plate. This preferred transparent cover plate balances high light transmittance and low refractive index difference, meeting the requirements of various high-resolution imaging techniques such as confocal microscopy and fluorescence microscopy, and effectively eliminating the scattering and light absorption effects of traditional Transwell porous membranes on imaging. In this embodiment, the bottom of the substrate 3 is directly replaced by a glass cover plate 7.

[0033] Preferably, the via chip is a multi-hole array structure. The multi-hole array design allows for the simultaneous placement of multiple via units (such as 6×4 or 8×8 arrays) on a single glass slide or a single substrate, significantly increasing experimental throughput and facilitating parallel comparisons under different drug concentration gradients or matrix ratios. Secondly, by designing multiple cavities on the same injection mold or microfluidic photomask and employing a unified molding or integrated packaging process, tight isolation and standardized dimensions between units are achieved, ensuring gradient consistency across different via locations and comparability of experimental results. In this embodiment, a 5×2 array is used.

[0034] This utility model has been described through preferred embodiments. Those skilled in the art will understand that various changes or equivalent substitutions can be made to these features and embodiments without departing from the spirit and scope of this utility model. This utility model is not limited to the specific embodiments disclosed herein; other embodiments falling within the scope of the claims of this application are all within the protection scope of this utility model.

Claims

1. A perforated chip for three-dimensional cell culture, characterized in that: It includes an outer hole (1), an inner hole (2) and a base (3), wherein the inner hole (2) is embedded within the outer hole (1); The outer hole (1), the inner hole (2) and the base (3) constitute an outer receiving cavity (4), and the inner hole (2) and the base (3) constitute a inner receiving cavity (5). The bottoms of the outer receiving cavity (4) and the inner receiving cavity (5) are connected.

2. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The outer hole (1) and the inner hole (2) are cylindrical, and the outer hole (1) and the inner hole (2) are located on the same central axis.

3. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: A connecting rib plate (6) is provided between the inner side of the outer hole (1) and the outer side of the inner hole (2) to connect the two.

4. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The height of the connection point between the bottom of the outer cavity (4) and the inner cavity (5) is 0.1-3mm.

5. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The ratio of the diameter of the outer hole (1) to the diameter of the inner hole (2) is (3-5):

1.

6. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The surface of the substrate (3) is provided with a first layer that increases surface tension.

7. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The outer hole (1) and the inner hole (2) are provided with a second surface tension-enhancing layer.

8. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The bottom of the substrate (3) is provided with an optically transparent polymer or glass cover.

9. The perforated chip for three-dimensional cell culture according to claim 1, characterized in that: The embedded chip has a multi-hole array structure.

Images