Multi-organ communicated liquid core assembly and multi-organ series co-culture method

Abstract

The invention discloses a multi-organ communicating liquid core assembly. The multi-organ communicating liquid core assembly comprises an upper cover module, a transwell module and a porous cell culture base, at least one communicated perfusion channel is arranged at the bottom in the porous cell culture base, and the communicated perfusion channel is at least communicated with one culture hole; the transwell module comprises a plurality of small transwell chambers, the small transwell chambers are matched with the culture holes, the small transwell chambers are placed on the culture holes, and the upper cover module is used for sealing and covering the porous cell culture base. According to the liquid core assembly, a traditional culture plate and a micro-fluidic technology are combined, organoid high-throughput culture and multi-organ series co-culture are achieved, a series multi-organ culture mode is adopted, a dynamic culture liquid core assembly model is established in a simple and convenient mode, and the culture efficiency is improved. The bionic degree and the uniformity of the multi-organ series co-culture tissue are improved.

Description

Technical Field

[0001] This invention relates to the field of multi-organ tissue culture, and more particularly to a multi-organ connected liquid core assembly and a method for multi-organ tandem co-culture. Background Technology

[0002] Organoids are 3D cultures created through in vitro culture of adult tissues, pluripotent stem cells, or induced pluripotent stem cells. They possess similar tissue structures and some physiological functions to their corresponding organs. They hold immense potential in disease research, drug screening, drug toxicology analysis, gene and cell therapy, and other fields.

[0003] Current organoid culture methods utilize traditional cell culture plates or microfluidic chip culture devices. For multi-organ co-culture, traditional cell culture plates lack continuous dynamic perfusion capabilities, resulting in low biomimicry and failing to meet the needs of multi-organ culture. While microfluidic chips can meet these requirements, they are not only complex and costly to operate, but also present significant technical hurdles and are difficult to characterize.

[0004] Currently, the main method for culturing organoids is static culture in well plates. Operators first need to mince the acquired primary samples, then encapsulate them in Matrigel under low temperature conditions. Culture medium is then added, and the plates are placed in an incubator for cultivation. During the culture process, the medium needs to be changed three times a week, and passaged every seven to ten days. However, this culture mode has several problems that urgently need to be addressed. First, organoid growth is relatively slow, resulting in a long overall culture cycle. During the culture process, operators need to manually change the medium periodically, which not only increases the risk of contamination but also significantly increases the investment costs in manpower, equipment, and space when the number of cultured samples increases. Second, due to the static culture method in the well plates, the exchange of substances between the organoids and the culture medium is insufficient. Especially for 3D cell tissues, internal nutrients are often difficult to absorb fully. This situation ultimately limits organoid growth, thus affecting its growth rate and the similarity to human organs. Therefore, existing organoid culture methods still need further improvement and optimization.

[0005] CN117363482B discloses a method for co-culturing different types of organoids. This method uses an organoid co-culturing device, which includes a top cover, at least two second culture plates, and a co-culturing tank detachably connected to the top cover. Each second culture plate includes a second abutment with an open culture tank at its upper end, containing organoid placement holes. A second culture medium filling tank is also located on both sides of the culture tank and communicates with it. On the two side walls of the second culture plate, corresponding male and / or female splicing components are provided. The male splicing component is an outward protrusion, and the female splicing component is an inward groove matching the male splicing component. The male splicing components of two adjacent second culture plates are detachably inserted into the female splicing components, achieving connection and communication between the two second culture plates. This disclosure allows for the fusion culture of organoids within the same chip, and also enables the co-culture of different types of organoids using a male-female component splicing method. Furthermore, the first culture plate and microneedle array setup allow for the co-culture of cell tissues (such as endothelial cells) with multiple organoids, realizing various possibilities. However, this disclosed culture is a static culture, and the number of cells cultured is small, requiring the culture plate to be opened during the culture process, increasing the risk of damage. Summary of the Invention

[0006] The purpose of this invention is to provide a multi-organ connected fluid core assembly and a method for multi-organ tandem co-culture, which can achieve continuous dynamic perfusion and establish a dynamic culture fluid core assembly model in a simple way through a common cell culture medium perfusion channel to achieve multi-organ tandem co-culture.

[0007] The present invention adopts the following technical solution: A multi-organ connected fluid core assembly includes a top cover module, a transwell module, and a porous cell culture medium holder. The top cover module is sealed to the porous cell culture medium holder. The transwell module includes multiple transwell chambers with identical structures. Each transwell chamber includes a semi-permeable membrane and a transwell chamber body with an internal cavity. The sidewall of the transwell chamber body has several first through grooves evenly distributed along the circumference. The semi-permeable membrane is fitted onto the outside of the transwell chamber body. The porous cell culture medium holder includes a base body with multiple culture wells inside, the shape of which is adapted to the shape of the transwell chambers. The base body has one or more cell culture medium perfusion channels, each cell culture medium perfusion channel communicating with at least one culture well. The inlet and outlet of each cell culture medium perfusion channel are respectively connected to the cell culture medium perfusion inlet and cell culture medium perfusion outlet provided on the base body.

[0008] Furthermore, the aforementioned first through grooves extend to the top of the side wall of the transwell chamber body and evenly divide the side wall of the transwell chamber body into several mutually isolated inner pressure plates. The bottom outer wall of the transwell chamber body is connected to the inner wall of each inner pressure plate. A connecting strip is provided between adjacent inner pressure plates, and the two ends of the connecting strip are respectively connected to the middle of two adjacent inner pressure plates.

[0009] Furthermore, the transwell chamber also includes a fixing sleeve; the sidewall of the fixing sleeve is evenly distributed with several second through slots corresponding to the first through slot along the circumferential direction; the bottom of the inner wall of the fixing sleeve is provided with a locking ring, and the top of the locking ring is evenly distributed with several arc-shaped protrusions corresponding to the second through slots along the circumferential direction, and each arc-shaped protrusion has a slot between its two ends and the inner wall of the fixing sleeve; the semi-permeable membrane is attached to the inner wall of the fixing sleeve, and the lower end of the semi-permeable membrane is inserted into the slot; the lower part of the sidewall of the transwell chamber body is embedded in the semi-permeable membrane. The first through slot, the semi-permeable membrane, and the second through slot form a semi-permeable membrane window.

[0010] Furthermore, the transwell chamber body is an inverted frustum, sealed at the bottom and disc-shaped, with a cylindrical lower sidewall; the semi-permeable membrane is a cylindrical structure with openings at both ends and adapted to the lower sidewall of the transwell chamber body; the fixing sleeve is a cylindrical structure with an opening at the top and adapted to the semi-permeable membrane; the outer wall of the locking ring is fitted to the bottom of the inner wall of the fixing sleeve, and the bottom of each inner pressure plate is fitted to the top of the locking ring; the bottom of the transwell chamber body is embedded in the locking ring with an interference fit, and the top surface of the bottom of the transwell chamber body is flush with the top surface of each arc-shaped protrusion; each connecting strip is provided with a buckle, and the top of the fixing sleeve is connected to each connecting strip through the buckle.

[0011] Furthermore, the connected perfusion channel has at least two channels; the cross-sectional area of ​​the connected perfusion channel is less than or equal to the cross-sectional area of ​​the cell culture medium perfusion inlet and outlet.

[0012] Furthermore, the upper cover module includes an upper cover body and a sealing element. The bottom outer wall of the upper cover body is provided with a sealing groove that mates with the sealing element, and the sealing element is fitted into the sealing groove.

[0013] Furthermore, the porous cell culture medium seat, the cover body, and the transwell chamber are made of transparent, biocompatible materials and can be processed using 3D printing and injection molding processes.

[0014] Furthermore, both the outer surfaces of the top cover and the porous cell culture medium base are provided with tooth-like structures, which can provide an anti-slip effect and make it easy for users to pick them up.

[0015] Furthermore, the outer edge of the culture well in the porous cell culture medium body is designed with a coded identifier for identification.

[0016] Furthermore, the present invention also relates to a method for multi-organ tandem co-culture of a multi-organ connected liquid core assembly, comprising the following steps in sequence: Step 1: Place the individual transwell chambers into the culture wells of the multi-porous cell culture medium holder; Step 2: After the transwell chamber is placed, sterilize the porous cell culture medium holder and the top cover module by ultraviolet light irradiation or by ethylene oxide sterilization. Step 3: The sample tissue is chopped, digested, filtered, and centrifuged to obtain an organoid suspension of the corresponding sample tissue. The organoid suspension and matrix gel are mixed evenly according to the set ratio. Then, the organoid-matrix mixture is seeded in a Transwell chamber, and a porous cell culture medium is placed in the chamber. In a cell culture incubator, the organoid-Matrix gel mixture is solidified; Step 4: After the organoid-Matrix gel mixture has solidified, pump the culture medium into the cell culture medium perfusion channel through the cell culture medium perfusion inlet. The culture medium enters each transwell chamber through the semi-permeable membrane window on the side wall of the transwell chamber. When the culture medium flows out of the cell culture medium perfusion outlet, close the top cover module on the porous cell culture medium seat to seal the transwell chamber, and then carry out dynamic culture of the organoid model.

[0017] The multi-organ connectivity fluid core assembly disclosed in this invention has the following beneficial effects: (1) Compared with traditional cell culture well plates, this invention can not only realize the information exchange of cells or tissues in the transwell chambers of the culture well through the interconnected perfusion channel, and construct a multi-organ tandem co-culture system, but also conduct comparative experiments on different cell culture medium perfusion channels.

[0018] (2) The present invention provides a continuous perfusion channel at the bottom of the culture well inside the porous cell culture base, which not only ensures the flatness of the bottom of the base and forms a series of multiple culture wells to realize the continuous perfusion of cell culture medium in multi-organ culture, but also satisfies the self-balance and mutual communication of two or more organ cultures, thereby realizing the reproduction of the physiological and pathological characteristics of organs in vivo in vitro and creating a highly realistic biomimetic environment for multi-organ culture.

[0019] (3) The transwell chamber of the present invention has multiple semi-permeable membrane windows evenly distributed along the circumference on the side wall, which changes the traditional transwell chamber mode of setting the semi-permeable membrane at the bottom. This makes the transwell chamber with this special structure better cooperate with the culture plate with continuous dynamic perfusion channel, thereby more realistically simulating the biomimetic environment of organoid culture.

[0020] (4) The liquid core assembly of the present invention adopts a modular structure, which can be flexibly and quickly replaced and assembled, and has good characterization operability. The modular scheme is easy to implement and easy to operate. In addition, the size and structure design of the liquid core assembly can better meet the needs of high-throughput culture medium perfusion and adapt to the use of various characterization equipment, so as to realize high-throughput characterization.

[0021] (5) The liquid core assembly of the present invention has a toothed structure on the outer surface of the cover body and the porous cell culture medium seat body, which not only facilitates the operation of the test personnel, but also enhances the strength of the body structure. In addition, the outer edge of the culture well in the porous cell culture medium seat body is designed with a coding mark to distinguish and identify, reducing the risk of misoperation. Attached Figure Description

[0022] Figure 1 This is a schematic diagram of the structure of the present invention; Figure 2 This is an exploded view of the present invention; Figure 3 This is an exploded view of the upper cover module in this invention; Figure 4 This is a schematic diagram of the transwell chamber structure in this invention; Figure 5 This is an exploded view of the transwell chamber in this invention; Figure 6 This is a schematic diagram of the structure of the fixing sleeve in this invention; Figure 7 This is a schematic diagram of the structure of the porous cell culture medium seat in this invention; Figure 8 This is a top view of the porous cell culture medium holder in this invention; Figure 9 For the present invention Figure 1 A top view of the multi-organ interconnected fluid core assembly; Figure 10 yes Figure 9 A structural cross-sectional view along the AA direction; Figure 11 yes Figure 9 A structural cross-sectional view along the BB direction.

[0023] The components are as follows: 1. Top cover module; 1-1. Top cover body; 1-2. Sealing element; 1-3. Toothed structure; 2. Transwell module; 2-1. Transwell chamber body; 2-2. Semi-permeable membrane; 2-3. Fixing sleeve; 2-4. First through groove; 2-5. Inner pressure plate; 2-6. Connecting strip; 2-7. Buckle; 2-8. Second through groove; 2-9. Locking ring; 2-10. Protrusion; 2-11. Locking groove; 3. Porous cell culture medium base; 3-1. Cell culture medium inlet; 3-2. Culture well; 3-3. Toothed structure; 3-4. Cell culture medium outlet; 3-5. Cell culture medium inlet channel; 3-6. Encoded marker; 3-7. Base body. Detailed Implementation

[0024] The present invention will now be described in detail with reference to the accompanying drawings and embodiments: like Figures 1 to 11 As shown, the multi-organ connectivity fluid core assembly of the present invention includes a top cover module 1, a transwell module 2, and a porous cell culture medium seat 3.

[0025] like Figures 2 to 3 As shown, the top cover module 1 includes a top cover body 1-1 and a sealing element 1-2. The top cover body 1-1 has serrated structures 1-3 on both its front and rear sides, providing an anti-slip effect and facilitating user handling. The top cover body 1-1 is made of a transparent biocompatible material, such as PS (polystyrene), PMMA (polymethyl methacrylate), PC (polycarbonate), PET (polyethylene terephthalate), PP (polypropylene), and COC / COP (cyclic olefin polymer). The sealing element 1-2 is made of an elastic biocompatible material, such as silicone rubber or TPU (thermoplastic polyurethane elastomer). The top cover module 1 can be manufactured using 3D printing and injection molding processes. Furthermore, the bottom outer wall of the top cover body 1-1 has a sealing groove that mates with the sealing element 1-2. The sealing element 1-2 is fitted into the sealing groove to achieve a tight seal between the top cover module 1 and the porous cell culture medium holder 3.

[0026] like Figures 4 to 6As shown, the transwell module 2 includes multiple transwell chambers with identical structures. Each transwell chamber includes a transwell chamber body 2-1, a semi-permeable membrane 2-2, and a fixing sleeve 2-3. The transwell chambers are made of transparent, biocompatible material and can be processed by 3D printing and injection molding. The transwell chamber body 2-1 is an inverted frustum with an internal cavity for experiments. Unlike traditional transwell chambers with a semi-permeable membrane at the bottom, the transwell chamber body 2-1 in this invention is sealed at the bottom and is disc-shaped; the lower part of the sidewall of the transwell chamber body 2-1 is cylindrical. Several first through grooves 2-4 are evenly distributed along the circumference of the sidewall of the transwell chamber body 2-1. The first through grooves 2-4 extend to the top of the sidewall of the transwell chamber body and evenly divide the sidewall of the transwell chamber body into several mutually isolated inner pressure plates 2-5. The bottom outer wall of the transwell chamber body 2-1 is connected to the inner wall of each inner pressure plate 2-5. An arc-shaped connecting strip 2-6 is provided between adjacent inner pressure plates 2-5, with both ends of the connecting strip 2-6 connected to the middle of the two adjacent inner pressure plates 2-5 respectively. The specific number of the first through slots 2-4 can be set according to actual needs. Specifically, the number of the first through slots on the transwell chamber can be adjusted according to the size of the culture well.

[0027] The semi-permeable membrane 2-2 is a cylindrical structure with openings at both the top and bottom and adapted to the lower sidewall of the transwell chamber body 2-1. There are no particular limitations on the composition and pore size of the semi-permeable membrane 2-2; for example, it can be made of materials such as polycarbonate or polyester.

[0028] The fixing sleeve 2-3 is a cylindrical structure with an open top that fits the semi-permeable membrane 2-2. Several second through grooves 2-8, corresponding to the first through groove 2-4, are evenly distributed along the circumferential direction on the side wall of the fixing sleeve 2-3. A retaining ring 2-9 is provided at the bottom of the inner wall of the fixing sleeve 2-3. The outer wall of the retaining ring 2-9 is fitted to the bottom of the inner wall of the fixing sleeve 2-3, and the fixing sleeve 2-3 and the retaining ring 2-9 are an integral structure. Several arc-shaped protrusions 2-10, corresponding to the second through grooves 2-8, are evenly distributed along the circumferential direction on the top of the retaining ring 2-9. Each arc-shaped protrusion 2-10 has a groove 2-11 between its two ends and the inner wall of the fixing sleeve 2-3 for fixing the semi-permeable membrane 2-2. The semi-permeable membrane 2-2 is fitted to the inner wall of the fixing sleeve 2-3, and the lower end of the semi-permeable membrane 2-2 is inserted into the groove 2-11. The lower part of the side wall of the transwell chamber body 2-1 is embedded in the semi-permeable membrane 2-2, and the bottom of each inner pressure plate 2-5 is attached to the top of the locking ring 2-9. The bottom of the transwell chamber body 2-1 is embedded in the locking ring 2-9, and the bottom of the transwell chamber body 2-1 and the locking ring 2-9 are press-fitted. The top surface of the bottom of the transwell chamber body 2-1 is flush with the top surface of each arc-shaped protrusion 2-10. Each connecting strip 2-6 is provided with a buckle 2-7, and the top of the fixing sleeve 2-3 is connected to each connecting strip 2-6 through the buckle 2-7.

[0029] In this invention, the first through groove 2-4, the corresponding second through groove 2-8, and the semi-permeable membrane 2-2 form a semi-permeable membrane window for the culture medium to enter and exit.

[0030] In this embodiment, the sidewall of the transwell chamber body 2-1 may have three first through grooves 2-4 evenly distributed along the circumferential direction. Correspondingly, the fixing sleeve 2-3 is provided with three second through grooves 2-8 and three protrusions 2-10. At the same time, the first through grooves 2-4, the second through grooves 2-8 and the semi-permeable membrane 2-2 form three semi-permeable membrane windows.

[0031] Compared with traditional transwell chambers, the transwell chamber of this invention changes the traditional transwell chamber mode of placing the semipermeable membrane at the bottom by setting a semipermeable membrane window on the side. This special transwell chamber can better cooperate with the porous cell culture medium seat 3 with continuous dynamic perfusion channel to realize continuous dynamic perfusion of culture medium during organoid culture, thereby realistically simulating the biomimetic environment of organoid culture.

[0032] like Figures 7 to 11As shown, the porous cell culture medium support 3 mainly includes a base body 3-7, a cell culture medium inlet 3-1, culture wells 3-2, a toothed structure 3-3, a cell culture medium outlet 3-4, cell culture medium channels 3-5, and a coding marker 3-6. The toothed structure 3-3 is located on the front and rear surfaces of the base body 3-7. Multiple culture wells 3-2 are located within the base body 3-7, and the shape of the culture wells 3-2 is adapted to the shape of the transwell chamber. One or more cell culture medium channels 3-5 are provided within the base body 3-7. Each cell culture medium channel 3-5 communicates with at least one culture well 3-2. The inlet and outlet of each cell culture medium channel 3-5 are respectively connected to the cell culture medium inlet 3-1 and cell culture medium outlet 3-4 provided on the base body 3-7. A coding marker 3-6 is provided on one side of the base body 3-7 for each culture well 3-2.

[0033] In this embodiment, the culture wells 3-2 have 12 circular holes, i.e., a 12-well design. In actual use, the number and arrangement of the culture wells can be set according to application requirements and are not limited to this. In this embodiment, the base body 3-7 has a rectangular structure, and the culture wells 3-2 are arranged in a rectangular array. In other embodiments, the culture wells 3-2 can also be arranged in a circular array or other ways. The cell culture medium perfusion channel 3-5 is located at the bottom of the culture wells 3-2 and communicates with at least one culture well 3-2. The cell culture medium perfusion channel 3-5 can be positioned on the central axis of each row or column of culture wells 3-2, or it can be positioned off-axis. Furthermore, there is no specific limitation on the number of cell culture medium perfusion channels 3-5; it can be set to one, two, or more as needed. The cell culture medium perfusion channel 3-5 can be flexibly designed according to the arrangement of the culture wells 3-2 and the culture wells 3-2 that need to be connected. Multiple culture wells 3-2 connected by the same cell culture medium perfusion channel 3-5 can be used to construct systems such as organoid culture and multi-organ tandem co-culture. Different cell culture medium perfusion channels 3-5 can be used for control experiments. There are no specific limitations on the cross-sectional shape of the cell culture medium perfusion channel 3-5. In this embodiment, the cross-sectional shape of the cell culture medium perfusion channel 3-5 is a curved trapezoid, and its cross-sectional area is less than or equal to the cross-sectional area of ​​the cell culture medium perfusion inlet / outlet. In other embodiments, the cross-sectional shape of the cell culture medium perfusion channel 3-5 can also be circular or rectangular.

[0034] In this embodiment, each transwell chamber can form an organoid model. The four culture wells 3-2 in each row are connected by a single cell culture medium perfusion channel 3-5, thus forming a multi-organ interaction model. Different cells can be cultured in the interconnected culture wells 3-2, exchanging information and substances through the shared cell culture medium perfusion channel 3-5. Alternatively, each row of culture wells 3-2 can be designed to be connected by a single cell culture medium perfusion channel 3-5.

[0035] The base body 3-7 is also equipped with coded markers 3-6. Each coded marker 3-6 is located on the outer edge of the corresponding culture well 3-2. The culture well 3-2 is distinguished and identified by coding, which reduces the risk of misoperation.

[0036] The base body 3-7 has symmetrically arranged tooth-like structures 3-3 on the middle part of the front and rear surfaces. In this embodiment, the upper cover body 1-1 has symmetrically arranged tooth-like structures on the middle part of the front and rear surfaces. When the upper cover module covers the porous cell culture medium base and seals the transwell chamber placed in the culture well, the tooth-like structures 3-3 can prevent slipping, making it convenient for the experimenter to pick up the entire core device. At the same time, the tooth-like structures 3-3 will reduce the deformation of the porous cell culture medium base 3 to a certain extent.

[0037] In this embodiment, the porous cell culture medium holder 3 can be customized using 3D printing with transparent resin material. In other embodiments, the porous cell culture medium holder 3 can also be manufactured using biocompatible transparent materials such as PS (polystyrene) and PMMA (polymethyl methacrylate).

[0038] The multi-organ tandem co-culture method based on the above-mentioned multi-organ connectivity liquid core assembly, as described in this invention, includes the following steps: Step 1: Place each individual transwell chamber into culture well 3-2 of the porous cell culture medium holder 3; Step 2: After placement, use ultraviolet light to sterilize the porous cell culture medium holder 3 and the top cover module 1 or sterilize them with ethylene oxide. Step 3: The sample tissue is chopped, digested, filtered, and centrifuged to obtain an organoid suspension of the corresponding sample tissue. The organoid suspension and matrix gel (such as Matrigel) are mixed evenly at a set ratio. The organoid-Matrix gel mixture is then seeded in a Transwell chamber, and the porous cell culture medium seat 3 is placed inside. In a cell culture incubator, the organoid-Matrix gel mixture is solidified; Step 4: After the organoid-Matrix gel mixture has solidified, pump the culture medium into the cell culture medium perfusion channel 3-5 through the cell culture medium perfusion inlet 3-1. The culture medium enters each transwell chamber through the semi-permeable membrane window on the side wall of the transwell chamber. When the culture medium flows out of the cell culture medium perfusion outlet 3-4, close the top cover module 1 on the porous cell culture medium seat 3 to seal the transwell chamber, and then carry out dynamic culture of the organoid model.

Claims

1. A multi-organ connectivity fluid core assembly, characterized in that: The system includes a top cover module, a transwell module, and a porous cell culture medium holder. The top cover module is sealed to the porous cell culture medium holder. The transwell module includes multiple transwell chambers with identical structures. Each transwell chamber includes a semi-permeable membrane and a transwell chamber body with an internal cavity. Several first through grooves are evenly distributed along the circumferential direction on the sidewall of the transwell chamber body. The semi-permeable membrane is fitted onto the outside of the transwell chamber body. The porous cell culture medium holder includes a base body with multiple culture wells inside. The shape of the culture wells is adapted to the shape of the transwell chambers. The base body has one or more cell culture medium perfusion channels. Each cell culture medium perfusion channel is connected to at least one culture well. The inlet and outlet of each cell culture medium perfusion channel are connected to the cell culture medium perfusion inlet and cell culture medium perfusion outlet on the base body, respectively.

2. The multi-organ connectivity fluid core assembly according to claim 1, characterized in that: The aforementioned first through grooves extend to the top of the side wall of the transwell chamber body and evenly divide the side wall of the transwell chamber body into several mutually isolated inner pressure plates. The bottom outer wall of the transwell chamber body is connected to the inner wall of each inner pressure plate. A connecting strip is provided between adjacent inner pressure plates, and the two ends of the connecting strip are respectively connected to the middle of two adjacent inner pressure plates.

3. The multi-organ connectivity fluid core assembly according to claim 2, characterized in that: The transwell chamber further includes a fixing sleeve; the side wall of the fixing sleeve is evenly distributed with several second through grooves corresponding to the first through groove along the circumferential direction; the bottom of the inner wall of the fixing sleeve is provided with a locking ring, and the top of the locking ring is evenly distributed with several arc-shaped protrusions corresponding to the second through grooves along the circumferential direction, and each arc-shaped protrusion has a locking groove between its two ends and the inner wall of the fixing sleeve; the semi-permeable membrane is attached to the inner wall of the fixing sleeve, and the lower end of the semi-permeable membrane is inserted into the locking groove; the lower part of the side wall of the transwell chamber body is embedded in the semi-permeable membrane.

4. The multi-organ connectivity fluid core assembly according to claim 3, characterized in that: The transwell chamber body is an inverted frustum, sealed at the bottom and disc-shaped, with a cylindrical lower side wall; the semi-permeable membrane is a cylindrical structure with openings at both the top and bottom and adapted to the lower side wall of the transwell chamber body; the fixing sleeve is a cylindrical structure with an opening at the top and adapted to the semi-permeable membrane. The outer wall of the locking ring fits against the bottom of the inner wall of the fixing sleeve, and the bottom of each inner pressure plate fits against the top of the locking ring; the bottom of the transwell chamber body is embedded in the locking ring with an interference fit, and the top surface of the bottom of the transwell chamber body is flush with the top surface of each arc protrusion; each connecting strip is provided with a buckle, and the top of the fixing sleeve is connected to each connecting strip by the buckle.

5. The multi-organ connectivity fluid core assembly according to claim 1, characterized in that: The aforementioned interconnected perfusion channels have at least two; the cross-sectional area of ​​the interconnected perfusion channels is less than or equal to the cross-sectional area of ​​the cell culture medium perfusion inlet and outlet.

6. The multi-organ connectivity fluid core assembly according to claim 1, characterized in that: The upper cover module includes an upper cover body and a sealing element. The bottom outer wall of the upper cover body is provided with a sealing groove that mates with the sealing element, and the sealing element is fitted into the sealing groove.

7. The multi-organ connectivity fluid core assembly according to claim 1, characterized in that: The porous cell culture medium holder, the top cover body, and the transwell chamber are made of a transparent, biocompatible material.

8. The multi-organ connectivity fluid core assembly according to claim 1, characterized in that: Both the outer surface of the top cover body and the porous cell culture medium base body are provided with tooth-like structures.

9. The multi-organ connectivity fluid core assembly according to claim 1, characterized in that: The outer edge of the culture well in the porous cell culture medium body is designed with a coded mark for identification.

10. A method for multi-organ tandem co-culture based on a multi-organ connectivity liquid core assembly according to any one of claims 1 to 9, comprising the following steps in sequence: Step 1: Place the individual transwell chambers into the culture wells of the multi-porous cell culture medium holder; Step 2: After the transwell chamber is placed, sterilize the porous cell culture medium holder and the top cover module by ultraviolet light irradiation or by ethylene oxide sterilization. Step 3: The sample tissue is chopped, digested, filtered, and centrifuged to obtain an organoid suspension of the corresponding sample tissue. The organoid suspension and matrix gel are mixed evenly according to the set ratio. Then, the organoid-matrix mixture is seeded in a Transwell chamber, and a porous cell culture medium is placed in the chamber. In a cell culture incubator, the organoid-Matrix gel mixture is solidified; Step 4: After the organoid-Matrix gel mixture has solidified, pump the culture medium into the cell culture medium perfusion channel through the cell culture medium perfusion inlet. The culture medium enters each transwell chamber through the semi-permeable membrane window on the side wall of the transwell chamber. When the culture medium flows out of the cell culture medium perfusion outlet, close the top cover module on the porous cell culture medium seat to seal the transwell chamber, and then carry out dynamic culture of the organoid model.

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