Microfluidic organ chip and system, application of microfluidic organ chip and system, and multi-organ co-culture method

Abstract

The invention relates to the technical field of biological tissue engineering, and discloses a microfluidic organ chip which comprises a chip body provided with a plurality of culture modules, and each culture module comprises a culture area comprising a first culture area and a second culture area which are communicated with each other, two first liquid storage tanks communicated with the first culture area respectively, and two second liquid storage tanks communicated with the second culture area respectively, the two second liquid storage tanks are respectively communicated with the second culture area, and a porous membrane is arranged between the first culture area and the second culture area to realize substance exchange; the culture area of each culture module, the two first liquid storage tanks and the two second liquid storage tanks are arranged corresponding to array hole positions of the hole plate. Each culture module is arranged corresponding to the hole site of the hole plate array, can be seamlessly connected with high-throughput detection equipment, and can be directly applied to scenes such as large-scale drug screening and drug effect evaluation. Gravity-driven or pump-driven perfusion culture can be realized, and single-organ co-culture and multi-organ co-culture can also be realized. The invention further discloses a microfluidic organ chip system, application thereof and a multi-organ co-culture method.

Description

Technical Field

[0001] This application relates to the field of biological tissue engineering technology, such as a microfluidic organ-on-a-chip and system, its application, and a method for multi-organ co-culture. Background Technology

[0002] Organ-on-a-chip (OoC) is a microfluidic cell culture device that integrates micro- and nanofabrication, cell biology, and biomaterials technologies to simulate the key structures and physiological functions of human organs on a micrometer-scale chip. This technology replicates complex physiological responses at the organ level by culturing living cells in a continuously perfused, multi-chamber structure, precisely controlling the mechanical properties of the fluid, the biochemical microenvironment, and intercellular interactions. Compared to traditional research models, organ-on-a-chip addresses a core pain point that has long plagued biomedical research: two-dimensional cell culture cannot simulate the three-dimensional dynamic characteristics of tissue interfaces (such as the blood vessel-tissue barrier), while animal models suffer from inherent drawbacks such as significant interspecies differences, high costs, low throughput, and ethical controversies.

[0003] Current organ-on-a-chip designs can be broadly categorized into three types: complex organ-on-a-chip systems for multi-organ co-culture, biomimetic microenvironment-regulated chips, and high-throughput drug screening chips. Each type has its own advantages and limitations. For example, complex organ-on-a-chip systems for multi-organ co-culture typically rely on complex flow control systems, and their equipment and flow path interfaces are usually highly customized products, resulting in high procurement and operating costs, as well as high costs related to cross-platform compatibility. Biomimetic microenvironment-regulated chips require the use of elastic materials such as PDMS to achieve specific mechanical forces. These materials have the problem of lipophilic drug adsorption, which may affect the accuracy of drug evaluation. In high-throughput drug screening chips, existing designs rely on matrix protein gels to form channels, but the success rate of this process is low, leading to limitations in actual test throughput. Furthermore, to achieve high throughput, these systems utilize liquid level differences and gravity to drive fluid flow.

[0004] In the process of implementing the embodiments of this disclosure, it has been found that at least the following problems exist in the related technologies: existing microfluidic organ-on-a-chip cannot be seamlessly connected to high-throughput detection equipment, and cannot simultaneously accommodate gravity-driven perfusion culture and fluid pump perfusion culture. Summary of the Invention

[0005] To provide a basic understanding of some aspects of the disclosed embodiments, a brief summary is given below. This summary is not intended as a general commentary, nor is it intended to identify key / important components or describe the scope of protection of these embodiments, but rather as a prelude to the detailed description that follows.

[0006] This disclosure provides a microfluidic organ-on-a-chip and system, its application, and a method for co-culturing multiple organs, to solve the problems that existing microfluidic organ-on-a-chip cannot seamlessly connect to high-throughput detection equipment and cannot simultaneously accommodate gravity-driven perfusion culture and fluid pump perfusion culture.

[0007] In some embodiments, the microfluidic organ-on-a-chip includes: a chip body having multiple culture modules disposed thereon; wherein each culture module includes: a culture region including a first culture region and a second culture region connected together, wherein a porous membrane is disposed between the first culture region and the second culture region to facilitate material exchange; two first liquid reservoirs respectively connected to the first culture region, and the connection positions are located on opposite sides of the first culture region; and two second liquid reservoirs respectively connected to the second culture region, and the connection positions are located on opposite sides of the second culture region; wherein the culture region, the two first liquid reservoirs and the two second liquid reservoirs of each culture module correspond to the array pore positions of the well plate.

[0008] In some embodiments, the microfluidic organ-on-a-chip includes: a support frame having a plurality of support positions thereon; a plurality of chip units, the chip units being movably disposed on the support positions; the chip unit includes a unit body and a plurality of culture modules disposed on the unit body, the culture modules being the culture modules in the aforementioned co-culture chip.

[0009] In some embodiments, the microfluidic organ-on-a-chip system includes any of the aforementioned microfluidic organ-on-a-chip and a plurality of connecting connectors. Each connecting connector includes a first connecting part and a second connecting part that are connected. The first connecting part is used to seal and dock with a first or second liquid reservoir, and the second connecting part is used to connect with an external component to realize pump-driven perfusion culture in the co-culture zone.

[0010] In some embodiments, any of the aforementioned microfluidic organ-on-a-chip or the aforementioned microfluidic organ-on-a-chip system is used for the in vitro construction of single-organ models, single-organ co-culture models, and multi-organ co-culture models.

[0011] In some embodiments, the method for multi-organ co-culture using the aforementioned microfluidic organs includes: culturing organ models with different culture cycles on different chip units; wherein, the organ models with different culture cycles are controlled to reach the cell state required for co-culture when they are cultured to the same time point; when the organ modules on different chip units reach the cell state required for co-culture, the chip units are placed on a support frame, and then the different chip units are connected to perform multi-organ co-culture.

[0012] The microfluidic organ-on-a-chip and system, their applications, and the method for multi-organ co-culture provided in this disclosure can achieve the following technical effects: In the microfluidic organ-on-a-chip of this disclosure, the co-culture area, two first reservoirs, and two second reservoirs of each culture module correspond to the array well arrangement of a well plate (e.g., a 96-well plate or a 384-well plate). This allows for seamless integration with mainstream fluorescence imaging systems, automated liquid handling platforms, and other high-throughput detection equipment, significantly improving experimental throughput and detection efficiency. It can be directly applied to large-scale drug screening and efficacy evaluation scenarios, effectively reducing the time and cost of batch experiments. The reservoir design allows for gravity-driven perfusion culture; alternatively, microfluidic components can be externally connected to the reservoirs to achieve pump-driven perfusion culture, thus combining both gravity-driven and pump-driven perfusion cultures. When a fluid pump drives perfusion culture, multiple culture modules of the same organ can be connected for single-organ co-culture, or multiple culture modules of different organs can be connected for multi-organ co-culture. Furthermore, by connecting different numbers of culture modules of different organs, for example, by determining the number of culture modules of different organs according to the physiological proportions of different organs, the physiological proportion scaling and material balance between different organs can be maintained, thereby enabling the construction of a more biomimetic multi-organ co-culture model.

[0013] In another microfluidic organ-on-a-chip provided in this embodiment, the chip units are independent and detachable structures that can be freely combined and flexibly arranged in the support positions of the support frame according to experimental needs (supporting independent experiments with a single chip unit or parallel experiments with multiple chip units). In addition, different organ models can be seeded and constructed independently on each chip unit first. When co-culture is required, the chip unit representing the desired organ can be placed in the support frame.

[0014] The above general description and the description below are exemplary and illustrative only and are not intended to limit this application. Attached Figure Description

[0015] One or more embodiments are illustrated by way of example with reference to the accompanying drawings. These illustrations and drawings do not constitute a limitation on the embodiments. Elements having the same reference numerals in the drawings are shown as similar elements. The drawings are not to be scaled. And wherein: Figure 1 This is a schematic diagram of the structure of a microfluidic organ-on-a-chip provided in an embodiment of this disclosure; Figure 2 This is a schematic diagram of the structure of a microfluidic organ-on-a-chip provided in an embodiment of this disclosure; Figure 3 This is a schematic diagram of a traditional perforated plate. Figure 4 This is a schematic diagram of the structure of a cultivation module provided in an embodiment of this disclosure; Figure 5 This is a schematic diagram of another cultivation module provided in an embodiment of this disclosure; Figure 6 This is a schematic diagram of another cultivation module provided in an embodiment of this disclosure; Figure 7 This is a schematic diagram of another cultivation module provided in an embodiment of this disclosure; Figures 8a to 8e This is a schematic diagram of the structure of the second channel layer, porous membrane layer, first channel layer and liquid storage layer of a microfluidic organ chip provided in this embodiment of the present disclosure; Figure 8f This is a cross-sectional view of a culture module of a microfluidic organ-on-a-chip provided in an embodiment of this disclosure; Figure 9 This is a three-dimensional structural schematic diagram of a microfluidic organ-on-a-chip provided in an embodiment of this disclosure; Figure 10a This is a schematic diagram of the structure of a connecting joint provided in an embodiment of this disclosure; Figure 10b yes Figure 10a A schematic cross-sectional view of the connecting joint shown. Figure 11 yes Figure 10a The diagram shows a cross-sectional view of the connecting joint when an external Luer connector is connected to it. Figure 12 This is a schematic diagram of another connecting connector provided in an embodiment of this disclosure; Figure 13 yes Figure 12 A schematic cross-sectional view of the connecting joint shown. Figure 14a This is a schematic diagram of the structure of a connecting cover provided in an embodiment of this disclosure; Figure 14b yes Figure 14a Schematic diagram of the cross-sectional structure along the middle AA direction; Figure 15a This is a schematic diagram of the structure of a connecting cover provided in an embodiment of this disclosure; Figure 15b This is a schematic diagram of the structure of a connecting cover provided in an embodiment of this disclosure; Figure 15c This is a schematic diagram of the structure of a connecting cover provided in an embodiment of this disclosure; Figure 16 This is a schematic diagram of the assembly structure of a microfluidic organ-on-a-chip system provided in an embodiment of this disclosure; Figure 17 yes Figure 16 Schematic diagram of the cross-sectional structure in the middle BB direction; Figure 18 This is a schematic diagram of another microfluidic organ-on-a-chip provided in this embodiment of the present disclosure; Figure 19aThis is a schematic diagram of another microfluidic organ-on-a-chip provided in this embodiment of the present disclosure; Figure 19b yes Figure 19a Schematic diagram of the cross-sectional structure along the CC direction; Figure 20 This is a schematic diagram of the structure of a chip unit provided in an embodiment of this disclosure; Figure 21 This is a schematic diagram of a load-bearing frame provided in an embodiment of this disclosure; Figure 22 This is a schematic diagram of another load-bearing frame provided in an embodiment of this disclosure; Figure 23 This is a schematic diagram of another load-bearing frame provided in an embodiment of this disclosure; Figure 24 This is a schematic diagram of the structure of a connecting cover provided in an embodiment of this disclosure; Figure 25 This is a flowchart of a method for multi-organ co-culture using a second type of microfluidic organ-on-a-chip, provided in an embodiment of this disclosure.

[0016] Figure label: 100. Chip body; 101. First channel layer; 102. Porous membrane layer; 103. Second channel layer; 104. Liquid storage layer; 1041. Horizontal frame; 1042. Vertical frame; 105. Base plate; 10. Culture module; 11. Culture area; 111. First culture area; 112. Second culture area; 12. First liquid storage tank; 121. First liquid storage tank I; 122. First liquid storage tank II; 13. Second liquid storage tank; 131. Second liquid storage tank I; 132. Second liquid storage tank II; 141. First connecting channel; 1410. First through hole; 142. Second connecting channel; 1420, Second through hole; 200, Connecting connector; 210, First connecting part; 211, First channel hole; 220, Second connecting part; 230, Luer connector; 300, Cover plate; 310, Plate surface; 311, Frame; 400, Support frame; 401, Support position; 402, Operation clearance position; 410, Frame body; 411, Edge boss; 420, Divider; 500, Chip unit; 510, Unit body; 501, First channel unit layer; 502, Porous membrane unit layer; 503, Second channel unit layer; 504, Liquid storage unit layer; 505, Base plate unit layer. Detailed Implementation

[0017] To provide a more detailed understanding of the features and technical content of the embodiments of this disclosure, the implementation of the embodiments of this disclosure will be described in detail below with reference to the accompanying drawings. The accompanying drawings are for illustrative purposes only and are not intended to limit the embodiments of this disclosure. In the following technical description, for ease of explanation, several details are used to provide a full understanding of the disclosed embodiments. However, one or more embodiments may still be implemented without these details. In other cases, well-known structures and devices may be simplified in their depiction to simplify the drawings.

[0018] The terms "first," "second," etc., used in the specification, claims, and accompanying drawings of this disclosure are used to distinguish similar objects and are not necessarily used to describe a specific order or sequence. It should be understood that such data can be interchanged where appropriate for the embodiments of this disclosure described herein. Furthermore, the terms "comprising" and "having," and any variations thereof, are intended to cover non-exclusive inclusion.

[0019] In this disclosure, the terms "upper," "lower," "inner," "middle," "outer," "front," and "rear," etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. These terms are primarily for better description of the embodiments of this disclosure and their implementations, and are not intended to limit the indicated devices, elements, or components to having a specific orientation, or to require them to be constructed and operated in a specific orientation. Furthermore, some of the aforementioned terms may be used to indicate other meanings besides orientation or positional relationship; for example, the term "upper" may in some cases indicate a dependency or connection relationship. Those skilled in the art can understand the specific meaning of these terms in the embodiments of this disclosure according to the specific circumstances.

[0020] Furthermore, the terms "set up," "connect," and "fix" should be interpreted broadly. For example, "connection" can be a fixed connection, a detachable connection, or an integral structure; it can be a mechanical connection or an electrical connection; it can be a direct connection or an indirect connection through an intermediate medium, or it can be an internal connection between two devices, components, or parts. Those skilled in the art can understand the specific meaning of the above terms in the embodiments of this disclosure according to the specific circumstances.

[0021] Unless otherwise stated, the term "multiple" means two or more.

[0022] It should be noted that, unless otherwise specified, the embodiments and features described in the present disclosure can be combined with each other.

[0023] Combination Figures 1 to 9As shown, this embodiment of the disclosure provides a microfluidic organ-on-a-chip, including a chip body 100 on which multiple culture modules 10 are disposed. Each culture module 10 includes a culture region 11, two first liquid reservoirs (first liquid reservoir I 121 and first liquid reservoir II 122), and two second liquid reservoirs (second liquid reservoir I 131 and second liquid reservoir II 132). The culture region 11 includes a first culture region 111 and a second culture region 112 that are connected, with a porous membrane between the first culture region 111 and the second culture region 112 to facilitate material exchange. The two first liquid reservoirs are respectively connected to the first culture region 111, and the connection points are located on opposite sides of the first culture region 111. The two second liquid reservoirs are respectively connected to the second culture region 112, and the connection points are located on opposite sides of the second culture region 112. The co-culture region 11, the two first liquid reservoirs, and the two second liquid reservoirs of each culture module 10 are arranged corresponding to the array pore positions of a well plate.

[0024] In the microfluidic organ-on-a-chip (referred to as the first type of microfluidic organ-on-a-chip) of this disclosure, the co-culture area 11, two first reservoirs 12, and two second reservoirs 13 of each culture module 10 correspond to the array well arrangement of a well plate (e.g., a 96-well plate or a 384-well plate). This allows for seamless integration with mainstream fluorescence imaging systems, automated liquid handling platforms, and other high-throughput detection equipment, significantly improving experimental throughput and detection efficiency. It can be directly applied to large-scale drug screening, efficacy evaluation, and other scenarios, effectively reducing the time and cost of batch experiments. The reservoir design allows for gravity-driven perfusion culture; alternatively, microfluidic components can be externally connected to the reservoirs to achieve pump-driven perfusion culture, thus combining both gravity-driven and pump-driven perfusion cultures. In fluid pump-driven perfusion culture, multiple culture modules of the same organ can be connected for single-organ co-culture, or multiple culture modules of different organs can be connected for multi-organ co-culture. Furthermore, by connecting different numbers of culture modules of different organs—for example, determining the number of culture modules based on the physiological proportions of different organs—the physiological scaling and material balance between different organs can be maintained, thus enabling the construction of more biomimetic multi-organ co-culture models. For instance, based on the in vivo liver-kidney volume ratio, connecting 3 liver units (3 culture modules) with 1 kidney unit (1 culture module) can construct a more biomimetic liver-kidney co-culture model.

[0025] Furthermore, the culture module 10 of the microfluidic organ-on-a-chip in this embodiment is laid out with a reference plate. Therefore, compared with existing long-channel microfluidic chips, the culture area 11 of the culture module 10 in this embodiment has a smaller volume and a lower cell seeding amount. For scarce primary cells, organoids and other precious experimental materials, this can minimize sample consumption and improve resource utilization.

[0026] It is understood that the microfluidic organ-on-a-chip of this disclosure is designed to resemble the well plate design of existing conventional well plates. Multiple culture modules 10 are arrayed on the chip body to adapt to high-throughput testing equipment, and the external dimensions of the chip body are consistent with the external dimensions of the reference well plate. The well plate can be a conventional 96-well plate or a 384-well plate, determined according to actual needs.

[0027] In some embodiments, the size of the chip body 100 is consistent with the size of existing conventional via plates. For example... Figure 2 and Figure 3 As shown, the length L1 and width L2 of the chip body 100 are the same as the length L1′ and width L2′ of a conventional well plate, respectively. Optionally, the minimum distance D between two adjacent culture modules 10 is the same as the minimum distance D′ between adjacent culture wells in a conventional well plate.

[0028] Optionally, the chip body 100 is primarily made of low-molecular-weight adsorption polypropylene (PS) and PET materials to reduce drug adsorption. Of course, it is not limited to these materials; other usable materials can also be used in the microfluidic organ-on-a-chip of this disclosure.

[0029] In some embodiments, combined with Figure 2 and Figure 3 As shown, the culture area 11, two first reservoirs 12, and two second reservoirs 13 of each culture module 10 are arranged in a 3×3 array of well positions 10′ corresponding to the well plate. The culture area 11 corresponds to well position (2, 2), and the two first reservoirs 12 and two second reservoirs 13 correspond to well positions (1, 1), (1, 3), (3, 1), and (3, 3). In this embodiment, the number of culture modules 10 is determined based on the number and array of culture wells of the benchmark well plate. For example, a commercially available standard 384-well plate (such as...) Figure 3 As shown), it is a 16×24 well plate with 16 rows and 24 columns. Therefore, in this embodiment, the multiple culture modules 10 on the microfluidic organ chip are designed as a 5×8 array, with a total of 40 culture modules 10 (as shown). Figure 2 (As shown).

[0030] It is understandable that "correspondence" means that there is overlap in position, but the shapes and sizes of the two corresponding structures are not exactly the same. The correspondence method is determined based on parameters such as the shape and size of the culture zone 11 and the storage tank.

[0031] In this embodiment, the shape, area, and other parameters of the culture area 11 are not limited and can be determined according to actual needs to achieve different fluid flow rates, shear forces, or exposure areas.

[0032] In some embodiments, combined with Figure 2 , Figures 4 to 6As shown, the culture region 11 includes a channel-type culture region 11. In this embodiment, the length of the channel-type culture region 11 is greater than the side length of the culture well at its location, while the width is less than or equal to the side length of the culture well at its location. Therefore, depending on the width, the channel-type culture region 11 can completely or partially cover the culture well at its location. It is understood that compared to traditional long-channel microfluidic chips, the length of the channel-type culture region in this embodiment is significantly shorter. While minimizing sample consumption and improving resource utilization, it can also reduce the layout area of ​​the culture module 10, adapting to the integrated layout of the culture module. This avoids the problem of limited integration quantity caused by excessive space occupation in long channels, enabling high-density arrangement of multiple culture modules and constructing a high-throughput microfluidic organ-on-a-chip. Furthermore, it completely avoids the interference of regional effects in long channels: In traditional long-channel microfluidic systems, the acceleration effect of fluid at the inlet, the steady-state flow in the middle, and the deceleration effect at the outlet create a significant velocity gradient, resulting in uneven distribution of fluid shear force in different regions of the channel—the shear force is higher at the inlet and outlet, while the shear force in the middle region is relatively stable. This difference causes regional differences in cell adhesion, proliferation, and differentiation (i.e., "regional effect"), which seriously affects the uniformity and reproducibility of experimental results. In contrast, the short-channel design shortens the fluid flow path, making the velocity distribution within the channel more uniform and the shear force difference smaller, ensuring the consistency of the cell culture environment throughout the channel, effectively eliminating the interference of regional effects on experimental results, and improving the reliability and comparability of data.

[0033] Optionally, the length-to-width ratio of the channel-type culture zone 11 is 3–6:1 to achieve better perfusion culture results. For example, 3:1, 3.5:1, 4:1, 5:1, 5.5:1, 6:1, etc.

[0034] In an optional example, the length of the channel-type culture zone 11 l The ratio of width to width w is 3.5 to 5:1.

[0035] Optionally, the length of the channel-type culture zone 11 l The length is 5–10 mm. Optionally, the length of the channel-type culture zone 11 is... l The length is 6–10 mm. Optionally, the length of the channel-type culture zone 11 is... l The diameter is 6.5–9.5 mm.

[0036] Optionally, the width w of the channel-type culture zone 11 is 1.5–3 mm. Optionally, the width w of the channel-type culture zone 11 is 1.5–2 mm.

[0037] In other embodiments, the culture region 11 includes a pore-shaped culture region 11. A pore-shaped culture region 11 refers to a culture region 11 whose cross-sectional shape has the same or similar dimensions in all directions, such as a circular or regular polygonal cross-sectional shape. Figure 7As shown, the cross-sectional shape of the culture zone 11 is a regular hexagon. The pore-shaped culture zone 11 can completely cover the culture wells at its location.

[0038] Optionally, the distance between the opposite two edges of the well-shaped culture area 11 is in the range of 3 to 5 mm. Optionally, the distance between the opposite two edges of the well-shaped culture area 11 is in the range of 3.5 to 4.5 mm.

[0039] It is understood that the first culture area 111 and the second culture area 112 are connected to form the culture area 11. Therefore, when describing parameters such as shape and size, the culture area 11, the first culture area 111 and the second culture area 112 can be substituted for each other. For example, the channel-type culture area 11 can be replaced to obtain the channel-type first culture area 111 and the channel-type second culture area, and so on.

[0040] In some embodiments, the depths of the first culture zone 111 and the second culture zone 112 are each 200–800 μm, determined according to the actual situation. For example, 200 μm, 250 μm, 300 μm, 350 μm, 400 μm, 450 μm, 500 μm, 550 μm, 600 μm, 650 μm, 700 μm, 800 μm, etc.

[0041] Optionally, the depths of the first culture zone 111 and the second culture zone 112 are each 300–600 μm. Optionally, the depths are each 500 μm.

[0042] The liquid storage tank (the collective name for the first liquid storage tank 12 and the second liquid storage tank 13) has a liquid storage function, and its shape and size are designed to meet the requirements of gravity-driven irrigation, without being specifically limited here.

[0043] In some embodiments, the depths of the first liquid storage tank 12 and the second liquid storage tank 13 are each 5 to 15 mm, determined according to the actual situation. For example, 5 mm, 6 mm, 7 mm, 8 mm, 9 mm, 10 mm, 11 mm, 12 mm, 13 mm, 14 mm, 15 mm, etc.

[0044] In some embodiments, a plurality of culture modules 10 are arranged in an array on the chip body, and the center distance d of the culture areas 11 of any two adjacent culture modules 10 is equal. Figure 2 and Figure 3 As shown, the center distance d is equal to the center distance d′ of the two adjacent 3×3 array aperture positions 10′ on a conventional standard aperture plate (2,2).

[0045] In some embodiments, the communication positions between the two first liquid storage tanks 12 and the first culture zone 111 are located in a first direction of the first culture zone 111 (e.g., ...). Figure 4 The two second liquid storage tanks 13 and the second culture zone 112 are located at opposite ends on the first direction of the second culture zone 112 (in the direction indicated by the dashed line p).

[0046] Optionally, when the culture area 11 is a channel-type culture area, the first direction is the length direction of the channel-type culture area.

[0047] Optionally, when the culture area 11 is a pore-shaped culture area, the first direction is the length direction or the width direction of the chip body.

[0048] In some embodiments, combined with Figure 4 As shown, the first storage tank 12 and the first culture zone 111 are connected by a first connecting channel 141. The angle α1 between the first connecting channel 141 and the first culture zone 111 in a first direction is greater than 90° and less than 180°. Optionally, the angle α1 is greater than or equal to 100° and less than or equal to 170°. Optionally, the angle α1 is greater than or equal to 110° and less than or equal to 160°. By setting the angle α1, disturbance is avoided when the fluid enters the culture zone 11 from the first storage tank 12, so as to form a stable fluid shear force in the culture zone 11.

[0049] In some embodiments, the second reservoir 13 and the second culture zone 112 are connected via a second connecting channel 142, and the angle α2 between the second connecting channel 142 and the second culture zone 112 in a first direction is greater than 90° and less than 180°. Optionally, the angle α2 is greater than or equal to 100° and less than or equal to 170°. Optionally, the angle α2 is greater than or equal to 110° and less than or equal to 160°. By setting the angle α2, disturbances are avoided when the fluid enters the culture zone 11 from the second reservoir 13, so as to form a stable fluid shear force in the culture zone 11.

[0050] Optionally, the included angle α1 and included angle α2 are equal. This results in the formation of the same and stable fluid shear force in the first culture zone 111 and the second culture zone 112.

[0051] In this embodiment of the present disclosure, the co-cultivation area 11, the two first liquid storage tanks 12 and the two second liquid storage tanks 13 of each cultivation module are arranged in accordance with the array of holes of the orifice plate, and the two first liquid storage tanks 12 and the two second liquid storage tanks 13 are all connected to the cultivation area 11. Therefore, the cultivation area 11 is located in the middle of the cultivation module 10, and the first liquid storage tanks 12 and the second liquid storage tanks 13 are arranged around the cultivation area 11.

[0052] In some embodiments, the line connecting the two first liquid storage tanks 12 intersects the line connecting the two second liquid storage tanks 13. In this embodiment, the first liquid storage tanks 12 and the second liquid storage tanks 13 are alternately arranged around the culture zone 11.

[0053] Optionally, two first liquid storage tanks 12 are arranged on both sides of the first culture zone 111 along a first direction, and two second liquid storage tanks 13 are arranged on both sides of the second culture zone 112 along a first direction. That is, one first liquid storage tank (e.g., first liquid storage tank I 121) and one second liquid storage tank (e.g., second liquid storage tank I 131) are arranged on one side of the culture zone 11 along a first direction, and another first liquid storage tank (e.g., first liquid storage tank II 122) and another second liquid storage tank (e.g., second liquid storage tank II 132) are arranged on the other side of the culture zone 11 along a first direction.

[0054] In an optional example, such as Figures 4 to 7 As shown, the first liquid storage tank I121 and the second liquid storage tank I131 are located on one side of the first direction of the culture zone 11, and the first liquid storage tank II122 and the second liquid storage tank II132 are located on the other side of the first direction of the culture zone 11.

[0055] In an optional example, the culture zone 11, two first liquid storage tanks 12, and two second liquid storage tanks 13 of each culture module are arranged in a 3×3 array of well positions 10′ corresponding to the well plate. The culture zone 11 corresponds to well position (2, 2), the first liquid storage tank I 121 and the first liquid storage tank II 122 correspond to well positions (1, 1) and (3, 3), and the second liquid storage tank I 131 and the second liquid storage tank II 132 correspond to well positions (1, 3) and (3, 1). The specific well positions of each liquid storage well are determined according to the first direction of the culture zone 11.

[0056] Optionally, combined Figure 1 , Figure 2 and Figure 3 As shown, taking the first direction of the culture zone 11 as the horizontal direction as an example, on a 3×3 array of holes 10′, the culture zone 11 is set with hole (2,2), the first liquid storage tank I 121 is set with hole (1,1), the first liquid storage tank II 122 is set with hole (1,3), the second liquid storage tank I 131 is set with hole (3,1), and the second liquid storage tank II 132 is set with hole (3,3).

[0057] The distance d1 between the first liquid storage tank 12 and the second liquid storage tank 13 located on the same side is not limited, but the distance d1 can be minimized as much as possible to increase the area of ​​the liquid storage tank and improve the liquid storage capacity.

[0058] Optionally, the distance d1 between the first liquid storage tank 12 and the second liquid storage tank 13 arranged on the same side along the first direction of the culture area 11 is 0.5-1 mm. This maximizes the area of ​​the liquid storage tanks while avoiding mutual interference between the first liquid storage tank 12 and the second liquid storage tank 13. Optionally, the distance d1 is 0.6-1 mm. Optionally, the distance d1 is 0.7-0.9 mm. In this embodiment, it can be understood that while each liquid storage tank is arranged corresponding to its respective hole position, the first liquid storage tank 12 and the second liquid storage tank 13 located on the same side also move closer to the center, occupying part of the adjacent hole positions.

[0059] Optionally, when the first liquid storage tank I121 is set with corresponding hole position (1,1) and the second liquid storage tank I131 is set with corresponding hole position (1,3), the first liquid storage tank I121 and the second liquid storage tank I131 extend towards each other and each occupy a portion of the adjacent hole position (1,2); similarly, the first liquid storage tank II122 and the second liquid storage tank II132 extend towards each other and each occupy a portion of the adjacent hole position (3,2).

[0060] Optionally, the distance d1 between the first liquid storage tank I121 and the second liquid storage tank I131 is 0.5 to 1 mm, and / or the distance d1 between the first liquid storage tank II122 and the second liquid storage tank II132 is 0.5 to 1 mm.

[0061] In this embodiment, the cross-sectional shape of the first liquid storage tank 12 and the second liquid storage tank 13 is not limited, as long as their cross-sectional area is maximized and they do not affect each other.

[0062] Optionally, combined Figure 4 , Figure 5 and Figure 7 As shown, the cross-sectional shape of the first liquid storage tank 12 and the second liquid storage tank 13 is polygonal.

[0063] Optionally, combined Figure 6 As shown, the cross-sectional shape of the first liquid storage tank 12 and the second liquid storage tank 13 is circular.

[0064] In this embodiment of the present disclosure, the vertical projections of the first liquid storage tank 12 and the second liquid storage tank 13 fall outside the culture area 11. This facilitates observation of the culture area 11 from above.

[0065] In some embodiments, an observation window is provided above the culture area 11. It is understood that the area of ​​the chip body 11 (or the liquid storage layer 104) above the culture area 11 is hollowed out to directly expose the culture area 11, so as to facilitate observation of the culture area 11.

[0066] Optionally, an observation window may be provided above at least the culture region 11 of the chip body 100. For example, an observation window may be provided on the reservoir layer 104 in the region corresponding to the culture region 11.

[0067] Optionally, in the first direction of the culture area 11, observation windows opened in multiple culture modules located in the same column or row are connected. This facilitates shaping and simplifies manufacturing.

[0068] In some embodiments, combined with Figures 8a to 8f As shown, the chip body 100 includes a second channel layer 103, a porous membrane layer 102, a first channel layer 101, and a liquid storage layer 104 stacked sequentially. The second channel layer 103 has a second culture area 112 and a communication channel (e.g., a second communication channel 142) for communicating with the second liquid storage tank 13. Figure 8a As shown; Figure 8c As shown, the first channel layer 101 is provided with a first culture zone 111 and a connecting channel (e.g., a first connecting channel 141) for communicating with the first storage tank 12, and a second through hole 1420 connecting the second storage tank 13 and the second culture zone 112; Figure 8d As shown, a first liquid storage tank 12 and a second liquid storage tank 13 are provided on the liquid storage layer 104.

[0069] In this embodiment, the second culture region 112 and the connecting channel on the second channel layer 103 can be either a through-structure or a non-through-structure. When it is a through-structure, a base plate 105 can be added to the bottom of the chip body 100 during use.

[0070] Optionally, the first culture area 111 and the connecting channel provided on the first channel layer 101 can be a through structure extending through the thickness direction of the first channel layer 101, or a non-through structure. In this embodiment, it can be understood that when it is a through structure, the structural schematic diagrams of both surfaces of the first channel layer 101 can be as shown below. Figure 8c As shown.

[0071] Optionally, when the first culture zone 111 and the connecting channel are non-penetrating structures, the openings of the first culture zone 111 and the connecting channel are located on the surface in contact with the porous membrane layer 102, so as to achieve communication between the first culture zone 111 and the second culture zone 112 (specifically, through the porous membrane). In this embodiment, a first through hole 1410 connecting the first liquid storage tank 12 and the first culture zone 111 is also formed on the first channel layer 101. At this time, Figure 8c The diagram shown is a schematic diagram of the structure of the first channel layer 101 on the side of the porous membrane layer 102. A schematic diagram of the structure of the first channel layer 101 on the side of the liquid storage layer 104 in this embodiment is shown below. Figure 8e As shown.

[0072] in, Figure 8f This is a cross-sectional view of a culture module, taken as an example, along the length of the channel in a channel-type culture module. Combination Figure 9 and Figure 17 As shown, a horizontal frame portion 1041 and a vertical frame portion 1042 connected to the outer edge of the horizontal frame portion 1041 are formed extending outward from the peripheral edge of the liquid storage layer 104. This makes the outer contour of the chip body 100 the same as the outer contour of a conventional perforated plate, so as to be able to seamlessly connect to high-throughput detection equipment such as mainstream fluorescence imaging systems and automated liquid handling platforms.

[0073] The microfluidic organ-on-a-chip of this disclosure can be externally connected to achieve pump-driven perfusion culture. Optionally, in a pump-driven perfusion culture mode, each culture module 10 has its own microfluidic perfusion culture, for example, a first reservoir I 121 serves as an inlet for fluid intake and a first reservoir II 122 serves as an outlet for fluid output; and / or, a second reservoir I 131 serves as an inlet for fluid intake and a second reservoir II 132 serves as an outlet for fluid output.

[0074] Optionally, another pump-driven perfusion culture mode connects multiple culture modules 10 (e.g., connected in series) to achieve co-culture of multiple culture modules. For example, the first reservoir II 122, which serves as the outlet of one culture module in the series path, is connected to the first reservoir I 121, which serves as the inlet of the next adjacent culture module, and / or, the second reservoir II 132, which serves as the outlet of one culture module in the series path, is connected to the second reservoir I 131, which serves as the inlet of the next adjacent culture module. In this perfusion culture mode, the number of culture modules is determined according to actual needs. The co-culture mode of this embodiment includes single-organ co-culture or multi-organ co-culture. It is understood that single-organ co-culture involves connecting multiple culture modules inoculated with the same organ cells. Multi-organ co-culture involves connecting multiple culture modules inoculated with different organ cells. In multi-organ co-culture, different numbers of culture modules inoculated with different organ cells can be connected according to the organ volume ratio or mass ratio in vivo to construct a more biomimetic organ co-culture.

[0075] For example, in multi-organ co-culture, based on the in vivo liver-kidney volume ratio, three liver units (three culture modules) are connected with one kidney unit (one culture module) to construct a more biomimetic liver-kidney co-culture model.

[0076] For example, in multi-organ co-culture, based on the physiological correlation between intestinal absorptive surface area and liver metabolic capacity, two intestinal units (two culture modules) are connected with one liver unit (one culture module) to construct an intestinal-liver co-culture model for studying the absorption and first-pass metabolism of orally administered drugs. Based on the balance between gas exposure and systemic clearance, one lung unit (one culture module) is connected with one liver unit (one culture module) to construct a lung-liver co-culture model, simulating the absorption of substances in the lungs and their entry into the liver via the bloodstream.

[0077] When microfluidic organ-on-a-chip requires external piping to achieve pump-driven perfusion culture, the connection structure between culture modules 10 is not limited. For applications of microfluidic organ-on-a-chip that require external piping to achieve pump-driven perfusion culture, combined with Figures 1 to 17 As shown, this disclosure provides a microfluidic organ-on-a-chip system (defined as a first type of microfluidic organ-on-a-chip system), including a microfluidic organ-on-a-chip (first type of microfluidic organ-on-a-chip) of any of the foregoing embodiments and a plurality of connecting connectors 200. Each connecting connector 200 includes a first connecting portion 210 and a second connecting portion 220. The first connecting portion 210 is used for sealed docking with a first liquid reservoir 12 or a second liquid reservoir 13 on the microfluidic organ-on-a-chip (specifically, the chip body 100), and the second connecting portion 220 is used for connection with an external communication component to realize pump-driven perfusion culture in the culture area 11. In this embodiment, by setting a plurality of matching connecting connectors 200, communication between different culture modules 10 is achieved.

[0078] It is understood that the external connecting components in this embodiment refer to the components required for pump-driven perfusion culture. Optionally, the external connecting components include Luer connectors, inter-module transition connectors, microfluidic pipes, and fluid pump systems, etc., as long as they can realize pump-driven perfusion culture, and can be determined according to the actual situation.

[0079] Optionally, the first connecting part 210 has an outer contour adapted to the opening or body of the liquid storage tank for sealing connection with the liquid storage tank. The sealing connection method is not limited; a sealing element can be provided on the mating surface to achieve a seal, or an interference fit can be used to achieve a seal.

[0080] In an optional example, combined Figure 10a , Figure 12 and Figure 17 As shown, the first connecting part 210 has an outer contour adapted to the body of the storage tank. That is, the first connecting part 210 is inserted entirely into the body of the storage tank, and the first connecting part 210 has a first channel hole 211, which communicates with the culture zone 11 respectively. In the structure of the first connecting part 210 in this embodiment, the perfusion fluid flows directly into the culture zone 11 and will not stagnate / store fluid in the storage tank, thus avoiding the absence of dead volume areas in the perfusion path.

[0081] Optionally, combined Figure 10b As shown, the sidewall of the first connecting part 210 is an inclined sidewall, inclined inward in the direction away from the second connecting part. This facilitates insertion, and during insertion, the contact with the inner wall of the liquid storage tank becomes increasingly tighter, resulting in a better sealing effect. Optionally, the inclination angle α is 0.5° to 5°. Optionally, the inclination angle α is 1° to 5°. Optionally, the inclination angle α is 2° to 3°.

[0082] Optionally, such as Figure 10a , Figure 10b and Figure 11 As shown, the second connecting portion 220 is used to connect with the Luer connector 230. The second connecting portion 220 is a plug-in interface used to seal and mate with one end of the Luer connector 230. It is understood that the connecting channel (e.g., the first channel hole 211) on the connecting connector 200 mates and mates with the internal channel of the Luer connector 230. It is understood that one end of the connecting channel is located on the bottom wall of the plug-in interface, and the other end is connected to the liquid storage tank or directly to the culture zone 11.

[0083] Optionally, the connecting channel on the connecting joint 200 has the same dimensions as the internal channel of the Luer joint 230. That is, the connecting channel smoothly transitions to the internal channel of the Luer joint, without creating additional dead volume areas.

[0084] Optionally, such as Figure 12 and Figure 13 As shown, in the connecting connector 200, the second connecting part 220 is constructed as a Luer connector structure. That is, the connecting connector 200 integrates a Luer connector structure, reducing connection operations and improving operational efficiency.

[0085] In other embodiments, combined with Figures 14a to 17 As shown, the microfluidic organ-on-a-chip system also includes a cover plate 300, which can be placed on the first type of microfluidic organ-on-a-chip. Multiple connecting connectors 200 are fixedly or detachably disposed on the surface 310 of the cover plate 300 in a manner corresponding to the positions of the first liquid reservoir 12 and the second liquid reservoir 13 on the chip body 100. The first connecting portion 210 of the connecting connector 200 is located on the first side of the cover plate 300 for sealing and docking with the first liquid reservoir 12 or the second liquid reservoir 13, and the second connecting portion 220 of the connecting connector 200 is located on the second side of the cover plate 300 for connecting with external components, thereby realizing pump-driven perfusion culture in the culture area 11. In this embodiment, the cover plate 300 and the multiple connecting connectors 200 constitute a connecting cover plate (defined as the first type of connecting cover plate) for realizing communication between different culture modules 10. The multiple connecting connectors 200 are integrated on the cover plate 300, allowing multiple connecting connectors 200 to be sealed and docked to the chip at once, improving operational efficiency.

[0086] Optionally, the cover plate 300 includes a plate surface 310 and a frame 311. The frame 311 is disposed on a first side of the plate surface 310. The frame 311 is consistent with the outer contour of the microfluidic chip body and can surround the outer side of the chip body. In this embodiment of the present disclosure, the cover plate 300 can cover the microfluidic organ chip. By setting the frame 311, the accuracy of positioning and the stability of the cover are improved.

[0087] Optionally, the connecting connector 200 is detachably disposed on the cover plate 300. In this embodiment, the required number and location of connecting connectors can be disposed on the cover plate 300 as needed, offering good flexibility and allowing for easy determination of the culture modules to be connected. Figure 14a As shown, the connecting connector 200 is detachably mounted on the cover plate 300, and the second connecting part 220 is a plug-in interface. It can be understood that the cover plate 300 has multiple mounting through holes on its surface 310 for the detachable mounting of the connecting connector 200.

[0088] Optionally, the connecting joint 200 is fixedly disposed on the cover plate 300. For example, the connecting joint 200 is fixedly disposed on the cover plate 300 by integral molding. Figure 15a As shown, the connecting joint 200 is fixedly disposed on the cover plate 300, and the first connecting part 210 is located on the first side of the cover plate 300.

[0089] When the second connecting part 220 of the connector 200 is a plug-in interface, only the plug-in interface structure of the second connecting part 220 can be seen on the plate surface 310 on the second side of the cover plate 300.

[0090] When the second connection portion 220 of the connecting structure 200 is an insertion interface, the microfluidic organ-on-a-chip system also includes multiple Luer connectors 230, which are detachably inserted into the second connection portion 220 of the connecting connector 200. In this embodiment, the connecting connector 200 is integrated on the cover plate 300 and the Luer connectors 230 are detachably connected, resulting in a simple structure and good flexibility. Figure 15b and Figure 15c The diagram shown is a schematic diagram of a Luer connector 230 inserted into the second connecting part 220 of the connecting connector 200.

[0091] Of course, when the second connecting portion 220 of the connecting connector 200 is constructed as a Luer connector, Figure 15b This can be considered as a solution where the connecting joint 200 is detachably mounted on the cover plate 300; Figure 15c This can be considered as a solution where the connecting joint 200 is fixedly installed on the cover plate 300, for example, it is integrally formed and fixedly installed on the cover plate 300.

[0092] Combination Figures 1 to 23 As shown, this disclosure also provides a microfluidic organ-on-a-chip, including a support frame 400 and a plurality of chip units 500. The support frame 400 is provided with a plurality of support positions 401. The chip units 500 are movably disposed on the support positions 401. The chip unit 500 includes a unit body 510 and one or more culture modules disposed on the unit body 510. The culture modules adopt the culture module 10 in the first microfluidic organ-on-a-chip of any of the foregoing embodiments.

[0093] In the microfluidic organ-on-a-chip (referred to as the second type of microfluidic organ-on-a-chip) of this disclosure, the carrier position 401 is sized to match the chip unit 500 and is used to support and fix the chip unit 500. The chip unit 500, as an independent and detachable structure, can be freely combined and flexibly arranged in the carrier position 401 of the carrier frame 400 according to experimental needs (supporting independent experiments with a single chip unit or parallel experiments with multiple chip units). During cell seeding, a single chip unit 500 can be removed for seeding, which is faster and has a higher cell seeding success rate compared to seeding an entire high-throughput plate. Furthermore, different organ models require different seeding and culture methods and culture cycles. Traditional fixed high-throughput chips are less compatible with parallel culture of different organ models. For example, lung chips require several weeks to construct and require a specific gas-liquid interface culture environment, while liver and intestinal models need to complete drug evaluation within one week after seeding. Therefore, in this embodiment, different organ models can be seeded and constructed independently on each chip unit. When co-culture is required, the chip unit 500 representing the required organ can be placed on the support frame 400, which solves the compatibility problem of culture conditions for different cell types.

[0094] In some embodiments, the outer contour dimensions of the support frame 400 are consistent with those of existing conventional well plates. It is adaptable to existing laboratory automation equipment (such as multiwell plate incubators and fluorescence imaging platforms), exhibiting good compatibility. Figure 18 and Figure 3 As shown, the length L3 and width L4 of the support frame 400 are the same as the length L1′ and width L2′ of a conventional well plate, respectively. Optionally, the minimum distance D between two adjacent culture modules 10 on the same chip unit 500 is the same as the minimum distance D′ between adjacent culture wells in a conventional well plate. It can be understood that when the chip unit 500 is placed on any of the adapting support positions 401 on the support frame 400, the culture area 11, the two first liquid reservoirs 12, and the two second liquid reservoirs 13 of the culture module 10 on the chip unit 500 correspond to the array well arrangement of the well plate.

[0095] In this embodiment of the disclosure, the specifications of the multiple chip units 500 can be the same or different, as long as the corresponding carrier bits 401 are set accordingly.

[0096] In some embodiments, a plurality of culture modules 10 are disposed on the chip unit 500, and the plurality of culture modules 10 are arranged in an array. Optionally, as Figure 18 , Figure 19a and Figure 20 As shown, multiple culture modules 10 on the chip unit 500 are arranged in a single row or column.

[0097] In some embodiments, the sidewalls of the chip unit 500 are inclined sidewalls, which taper downwards along the thickness direction of the chip unit 500. This facilitates the placement of the chip unit 500 within the support position 401, and the narrower bottom and wider top design allows the chip unit 500 to easily enter the support position 401 and gradually make closer contact with the inner side of the support position 401, improving the placement stability of the chip unit 500. The engaging design between the chip unit and the support position ensures placement stability and facilitates quick disassembly, cleaning, or replacement after experiments, significantly improving operational convenience.

[0098] Optionally, the angle between the inclined sidewall of the chip unit 500 and the vertical direction is 0.5° to 5°. Optionally, the angle between the inclined sidewall of the chip unit 500 and the vertical direction is 1° to 5°. Optionally, the angle between the inclined sidewall of the chip unit 500 and the vertical direction is 2° to 3°.

[0099] In some embodiments, a plurality of bearer bits 401 are arranged side by side. For example, as Figure 21 , Figure 22 and Figure 23 As shown, they are arranged side-by-side along the length of the load-bearing frame 400. The number of load-bearing positions 401 is unlimited and determined according to design requirements. For example, as... Figure 21 and Figure 22 The five carrier positions 401 shown; for example, as... Figure 23 The six carrier positions 401 are shown.

[0100] In some embodiments, the support frame 400 includes a frame 410 and a partition 420. The partition 420 is disposed within the frame 410 to divide the space within the frame 410 into multiple unit spaces. The unit spaces serve as support positions 401 for adapting and setting chip units 500.

[0101] Optionally, the bottom of at least the support position 401 of the support frame 400 is hollowed out. Due to the working distance limitations of commonly used fluorescence imaging lenses (most high-magnification objectives have a working distance of <10mm), the support frame 400 adopts a hollow base plate structure design—at least the solid obstruction of the traditional base plate in the support position 401 is removed, and only the edge support structure is retained. This can minimize the distance between the bottom of the chip unit 500 and the lens, ensuring that the lens can clearly focus on the inside of the chip during fluorescence imaging, avoiding imaging blur or limited field of view problems caused by excessive distance.

[0102] In an optional example, such as Figure 22 As shown, four partitions 420 are arranged along the width direction inside the support frame 400, dividing the space inside the frame 410 into five support positions 401. Each support position 401 can be equipped with a chip unit 500, and five culture modules 10 are arranged on the chip unit 500 in a single row.

[0103] In another optional example, such as Figure 23 As shown, five partitions 420 are arranged along the width of the support frame 400, dividing the space inside the frame 410 into six support positions 401. Each support position 401 can be equipped with a chip unit 500, and five culture modules 10 are arranged in a single row on the chip unit 500.

[0104] Optionally, the support frame 400 is also provided with an operation clearance position 402, which is adjacent to the support position 401. This facilitates operation of the chip unit 500. The operation clearance position 402 can be a notch to allow tools (e.g., tweezers) that hold the chip unit 500 to be placed in position.

[0105] In an optional example, such as Figure 19a As shown, there are gaps between adjacent chip units 500 and between chip units 500 and the frame 410 of the supporting frame 400. These gaps serve as operation clearance positions 402, facilitating the placement and removal of chip units 500.

[0106] Optionally, combined Figure 19b As shown, a support frame 400 has an upper surface of a partition 420 that is lower than the upper surface of a frame 410, and a lower surface of the partition 420 that is higher than the lower surface of the frame 410; and a protruding edge boss 411 is provided on the inner sidewall of the frame 410. This creates a gap between adjacent chip units 500, with a gap between each chip unit 500 and the frame 410, and ensures that the bottom of the chip unit 500 does not extend beyond the lower surface of the frame 410. For example, a protruding edge boss 411 is provided on the inner sidewall of the frame 410 along the length direction of the chip unit 500.

[0107] The culture module 10 in the second type of microfluidic organ chip adopts the culture module 10 in the first type of microfluidic organ chip of any of the aforementioned embodiments. Therefore, the structure and technical effects of the aforementioned culture module 10 are applicable to the second type of microfluidic organ chip.

[0108] In an optional example, combined Figure 19b As shown, the chip unit 500 includes a second channel unit layer 503 (with the same structure as the second channel layer 103), a porous membrane unit layer 502 (with the same structure as the porous membrane layer 102), a first channel unit layer 501 (with the same structure as the first channel layer 101), and a liquid storage unit layer 504 (with the same structure as the liquid storage layer 104) that are stacked and connected in sequence.

[0109] Optionally, the chip unit 500 also includes a base plate unit layer 505 (with the same structure as the base plate 105), disposed on the lower surface of the second channel unit layer 503. The second culture area 112 suitable for the second channel unit layer 503 is a through structure.

[0110] The second type of microfluidic organ-on-a-chip in this disclosure can be gravity-driven perfusion culture; alternatively, a microfluidic component can be externally connected to the reservoir to achieve fluid pump-driven perfusion culture, thus achieving compatibility with both gravity-driven and fluid pump-driven perfusion culture.

[0111] For the second type of microfluidic organ-on-a-chip application scenario that requires external piping to achieve pump-driven perfusion culture, combined with Figures 10a to 24 As shown, this disclosure provides a microfluidic organ-on-a-chip system, including a second type of microfluidic organ-on-a-chip according to any of the foregoing embodiments and a plurality of connecting connectors 200. Each connecting connector 200 includes a first connecting portion 210 and a second connecting portion 220. The first connecting portion 210 is used for sealed docking with a first liquid reservoir 12 or a second liquid reservoir 13 on the second type of microfluidic organ-on-a-chip (specifically, a plurality of chip units 500), and the second connecting portion 220 is used for connection with an external communication component to realize pump-driven perfusion culture in the culture area 11. In this embodiment, by setting a plurality of matching connecting connectors 200, communication between different culture modules 10 is achieved.

[0112] In the microfluidic organ-on-a-chip system of this embodiment, the structure of the connector 200 is the same as that of the connector 200 in the first microfluidic organ-on-a-chip system described above, and will not be repeated here.

[0113] In some embodiments, combined with Figure 24 As shown, the second type of microfluidic organ-on-a-chip system also includes a cover plate 300, which can be placed on the second type of microfluidic organ-on-a-chip; a plurality of connecting connectors 200 are fixedly or detachably disposed on the surface 310 of the cover plate 300 in a manner corresponding to the positions of the first liquid reservoir 12 and the second liquid reservoir 13 on each chip unit 500; wherein, the first connecting portion 210 of the connecting connector 200 is located on the first side of the cover plate 300 for sealing connection with the first liquid reservoir 12 or the second liquid reservoir 13, and the second connecting portion 220 of the connecting connector 200 is located on the second side of the cover plate 300 for connection with external components to realize pump-driven perfusion culture in the culture area 11. In this embodiment, the cover plate 300 and the plurality of connecting connectors 200 constitute a connecting cover plate (defined as the second type of connecting cover plate, such as...). Figure 24 As shown, this is used to achieve connectivity between different culture modules 10. Multiple connecting connectors 200 are integrated on the cover plate 300, allowing multiple connecting connectors 200 to be sealed and connected to the chip at one time, improving operational efficiency.

[0114] In this embodiment, the number of connecting connectors 200 on the second type of connecting cover plate is reduced according to the number of culture modules 10 on the second type of microfluidic organ chip, such as... Figure 24 As shown. The arrangement of the connecting connector 200 and the cover plate 300 is the same as the relevant content of the first type of connecting cover plate mentioned above, and will not be repeated here.

[0115] In some embodiments, the second microfluidic organ-on-a-chip system further includes a unit cover (not shown), and a connector 200 is fixedly or detachably disposed on the unit cover in a manner corresponding to the positions of the first liquid reservoir 12 and the second liquid reservoir 13 on the chip unit 500. The first connecting portion of the connector is located on the first side of the unit cover for sealing connection with the first or second liquid reservoir, and the second connecting portion of the connector is located on the second side of the unit cover for connection with external components, enabling pump-driven perfusion culture in the culture area. In this embodiment, the unit cover and multiple connectors 200 constitute a connector (defined as a third type of connector). The third type of connector in this embodiment can be understood as obtaining the second type of connector by dividing the chip unit 500. The arrangement of the unit cover and the connectors refers to the relevant content of the first type of connector described above, and will not be repeated here.

[0116] In some embodiments, the second microfluidic organ-on-a-chip system includes a second microfluidic organ-on-a-chip, a second connecting cover plate, and a third connecting cover plate. That is, the second microfluidic organ-on-a-chip can be equipped with both a single second connecting cover plate and a third connecting cover plate divided into multiple units. The appropriate type can be flexibly selected based on actual culture requirements.

[0117] It is understandable that the second type of microfluidic organ-on-a-chip is obtained by modularizing the first type of microfluidic organ-on-a-chip to make cell seeding and culture more flexible and applicable. Therefore, the structure of the first type of microfluidic organ-on-a-chip can be applied to the second type of microfluidic organ-on-a-chip, and the technical effects of the first type of microfluidic organ-on-a-chip can also be applied to the second type of microfluidic organ-on-a-chip. This will not be elaborated further. For an understanding of the second type of microfluidic organ-on-a-chip, please refer to the relevant content of the first type of microfluidic organ-on-a-chip mentioned above.

[0118] This disclosure also provides an application of microfluidic organ-on-a-chip in any of the foregoing embodiments for constructing single-organ models, single-organ co-culture models, and multi-organ co-culture models in vitro.

[0119] Combination Figure 25 As shown, this disclosure also provides a method for multi-organ co-culture using the aforementioned second type of microfluidic organ-on-a-chip, comprising the following steps: S1. Organ models with different culture cycles are cultured on different chip units respectively; wherein, the organ models with different culture cycles are controlled to reach the cell state required for co-culture when they are cultured to the same time point.

[0120] S2. When the organ modules on different chip units reach the cell state required for co-culture, the chip units are placed on the support frame, and then the different chip units are connected to perform multi-organ co-culture.

[0121] In this embodiment of the disclosure, the organ models used in step S1 for different culture cycles are not limited and are determined according to actual needs. Optionally, organ models for different culture cycles include lung models, liver models, intestinal models, kidney models, blood vessel models, blood-brain barrier models, and skin models.

[0122] In this embodiment of the disclosure, Figure 1 , Figure 2 , Figures 4 to 7 , Figures 8a to 8e ,as well as Figure 19a In the diagram, the cultivation module 10 is drawn using a top-down perspective. The red lines represent the first cultivation area 111, the first liquid storage tank 12, and the first connecting channel 141; the blue lines represent the second cultivation area 112, the second liquid storage tank 13, and the second connecting channel 142. The spatial relationships of each structure can be understood by combining... Figure 8f To understand.

[0123] In this embodiment, a 384-well plate is specifically used as an example for benchmark design. It is understood that, based on the content disclosed in this embodiment, those skilled in the art can implement the design of other conventional well plates, such as 96-well plates, 24-well plates, etc.

[0124] The foregoing description and accompanying drawings fully illustrate embodiments of the present disclosure to enable those skilled in the art to practice them. Other embodiments may include structural and other changes. The embodiments represent only possible variations. Individual components and functions are optional unless explicitly required, and the order of operation may vary. Parts and features of some embodiments may be included or substituted for parts and features of other embodiments. Embodiments of the present disclosure are not limited to the structures described above and shown in the accompanying drawings, and various modifications and changes may be made without departing from its scope. The scope of the present disclosure is limited only by the appended claims.

Claims

1. A microfluidic organ-on-a-chip, characterized in that, include: The chip body has multiple cultivation modules mounted on it; each cultivation module includes: The culture zone includes a first culture zone and a second culture zone that are connected together, with a porous membrane installed between the first culture zone and the second culture zone to facilitate material exchange. Two first liquid storage tanks are respectively connected to the first culture zone, and the connection points are located on opposite sides of the first culture zone; Two second liquid storage tanks are connected to the second culture zone, and the connection points are located on opposite sides of the second culture zone; In this context, the culture area, two first liquid storage tanks, and two second liquid storage tanks of each culture module correspond to the array of well positions of the well plate.

2. The microfluidic organ-on-a-chip according to claim 1, characterized in that, The culture area includes a channel-type culture area or a well-shaped culture area; and / or The culture zone includes a channel-type culture zone, the length-to-width ratio of which is 3–6:1; and / or, the length of which is 5–10 mm; and / or, the width of which is 1.5–3 mm; and / or The culture zone includes well-shaped culture areas, with the distance between the opposite edges of the well-shaped culture areas ranging from 3 to 5 mm; and / or The depths of the first and second culture zones are 200–800 μm, respectively.

3. The microfluidic organ-on-a-chip according to claim 1, characterized in that, The two first liquid storage tanks are connected to the first culture zone at opposite ends in a first direction of the first culture zone; the two second liquid storage tanks are connected to the second culture zone at opposite ends in a first direction of the second culture zone; and / or The first storage tank and the first culture zone are connected by a first connecting channel, wherein the angle α1 between the first connecting channel and the first direction of the first culture zone is greater than 90° and less than 180°; and / or The second storage tank and the second culture zone are connected by a second connecting channel, wherein the angle α2 between the second connecting channel and the first direction of the second culture zone is greater than 90° and less than 180°; and / or The line connecting the two first liquid storage tanks intersects the line connecting the two second liquid storage tanks; and / or Two first liquid storage tanks are arranged on both sides of a first direction in the first culture zone, and two second liquid storage tanks are arranged on both sides of a first direction in the second culture zone; and / or A first liquid storage tank and a second liquid storage tank are disposed on one side of the culture zone in a first direction, and another first liquid storage tank and another second liquid storage tank are disposed on the other side of the culture zone in the first direction; and / or The distance between the first and second liquid storage tanks, which are set on the same side along the first direction of the culture area, is 0.5 to 1 mm.

4. The microfluidic organ-on-a-chip according to any one of claims 1 to 3, characterized in that, Each culture module has a 3×3 array of well positions corresponding to the culture zone, two first reservoirs, and two second reservoirs, where the culture zone corresponds to well position (2,2), and the two first reservoirs and two second reservoirs are arranged with well positions (1,1), (1,3), (3,1), and (3,3); and / or The vertical projections of the first and second reservoirs fall outside the culture area; and / or An observation window is provided above the culture area.

5. A microfluidic organ-on-a-chip, characterized in that, include: A support frame, on which multiple support positions are provided; Multiple chip units are movably disposed in a carrier position; each chip unit includes a unit body and multiple culture modules disposed on the unit body, wherein the culture modules adopt the culture modules in the co-culture chip as described in any one of claims 1 to 4.

6. The microfluidic organ-on-a-chip according to claim 5, characterized in that, Multiple bearer positions are arranged side by side; and / or The supporting frame includes a frame and a partition frame. The partition frame is disposed within the frame and divides the space within the frame into multiple unit spaces. Each unit space serves as a support location for accommodating and mounting chip units; and / or The bottom of at least the load-bearing position of the supporting frame is hollowed out; and / or The sidewalls of the chip cell are sloping sidewalls, which taper downwards along the thickness direction of the chip cell; the angle between the sloping sidewalls and the vertical direction is 0.5° to 5°; and / or Multiple culture modules on the chip unit are arranged in a single row or column.

7. A microfluidic organ-on-a-chip system, characterized in that, include: The microfluidic organ-on-a-chip as described in any one of claims 1 to 4 or the microfluidic organ-on-a-chip as described in claim 5 or 6; Multiple connecting joints, each connecting joint including a first connecting part and a second connecting part, the first connecting part being used for sealing connection with a first liquid storage tank or a second liquid storage tank, and the second connecting part being used for connection with external components to realize pump-driven perfusion culture in the co-culture zone.

8. The microfluidic organ-on-a-chip system according to claim 7, characterized in that, The second connecting portion of the connector is used for connection with a Luer connector; or, the second connecting portion is configured as a Luer connector; and / or The microfluidic organ-on-a-chip system further includes: a cover plate capable of covering the microfluidic organ-on-a-chip; and multiple connecting connectors fixedly or detachably disposed on the surface of the cover plate in a manner corresponding to the positions of a first and a second liquid reservoir on the microfluidic organ-on-a-chip; wherein, a first connecting portion of the connecting connector is located on a first side of the cover plate for sealing connection with the first or second liquid reservoir, and a second connecting portion of the connecting connector is located on a second side of the cover plate for connection with external components to realize pump-driven perfusion culture in the culture zone; and / or When the microfluidic chip uses the microfluidic organ-on-a-chip as described in claim 5 or 6, the microfluidic organ-on-a-chip system further includes: a unit cover plate, and a connecting connector fixedly or detachably disposed on the surface of the unit cover plate in a manner corresponding to the positions of the first and second liquid reservoirs on the chip unit; wherein, the first connecting portion of the connecting connector is located on the first side of the unit cover plate for sealing and docking with the first or second liquid reservoir, and the second connecting portion of the connecting connector is located on the second side of the unit cover plate for connecting with external components to realize pump-driven perfusion culture in the culture area.

9. The application of the microfluidic organ-on-a-chip system as described in any one of claims 1 to 6 or the microfluidic organ-on-a-chip system as described in claim 7 or 8 for the in vitro construction of single-organ models, single-organ co-culture models and multi-organ co-culture models.

10. A method for multi-organ co-culture using a microfluidic organ-on-a-chip as described in claim 5 or 6, characterized in that, include: Organ models with different culture cycles were cultured on different chip units; among them, the organ models with different culture cycles were controlled to reach the cell state required for co-culture when they were cultured to the same time point; When the organ modules on different chip units reach the cell state required for co-culture, the chip units are placed on the support frame, and then the different chip units are connected to perform multi-organ co-culture.

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