Closed live cell tissue perfusion culture device

Abstract

The utility model relates to the field of cytology, and discloses a closed living cell tissue perfusion culture device which can ensure that liquid flow flowing through incubated living cells and in-vitro tissues is stable and a perfusion liquid is collected without loss. Comprising a perfusion cabin bottom, a perfusion cabin body, a pressurization fastening structure, an upper sealing cover and a filtering layer. The cabin bottom is funnel-shaped, the lower end of the bottom is provided with a perfusate outlet, and a filter layer is arranged between the funnel and the cabin body to prevent living cells or in-vitro tissues from leaking and ensure that a culture solution flows through; the cabin bottom accommodates the cabin body, and the cabin body is pressed on the filter layer and is in closed connection with the cabin bottom; an incubation cavity is formed in the cabin body and is an incubation culture space for living cell tissues, a filtering layer is arranged on the bottom side of the space, and the cabin body is opened upwards, so that perfusate, living cells or in-vitro tissues can be injected into the incubation cavity, and then the cabin body is hermetically connected with an upper sealing cover in which a perfusate inlet penetrates. A perfusion culture solution enters the incubation cavity through the inlet and flows out of the outlet through the filter layer.

Description

Technical Field

[0001] This application relates to the field of cell biology, specifically to a closed-loop live cell tissue perfusion culture device. Background Technology

[0002] This section is intended to provide background or context for the embodiments of this application as set forth in the claims. The content in this section is for reference only and does not constitute an admission or confirmation that it is prior art that has been disclosed.

[0003] In current live cell or ex vivo tissue perfusion experiments, the perfusion medium flows through the sample to maintain its physiological activity. By directly collecting the perfusion fluid or adding specific drug components to the perfusion medium, researchers use biochemical experiments, immunological experiments, or mass spectrometry to detect changes in the composition of the perfusion medium during the perfusion process. This allows for in-depth analysis of cell metabolites, specific products of drug responses, and their changes, ultimately revealing the mechanisms of metabolism in the sample and the effects of the drug. However, existing perfusion fluid collection methods generally employ open structures, such as ex vivo organ perfusion tanks or similar microfluidic systems. These open-structure perfusion fluid collection methods are susceptible to liquid evaporation, which can adversely affect experimental results.

[0004] Furthermore, while the closed structure of the incubation tank can reduce the evaporation of the perfusion solution to some extent and help stabilize the experimental environment, the lack of tight connections between components can easily lead to leakage and even loss of incubated cells during operation. This severely affects the success rate of accurate collection of the perfusion solution and limits its widespread application in experiments. Therefore, maintaining environmental stability, reducing systematic errors, and achieving accurate and quantitative collection of the perfusion solution during the incubation of live cells and ex vivo tissues have become key technical challenges for the success of experiments in this field. Summary of the Invention

[0005] The purpose of this application is to provide a closed-loop live cell tissue perfusion culture device that can ensure a stable incubation environment and constant fluid flow for live cells and ex vivo tissues, thereby improving the success rate of experiments and minimizing losses during perfusion fluid collection.

[0006] The closed-type live cell tissue perfusion culture device disclosed in this application includes: perfusion incubation chamber bottom 1, perfusion chamber body 2, pressure fastening structure 3, upper sealing cover 4, and filter layer 6.

[0007] The bottom inner side of the perfusion incubation chamber bottom 1 is recessed downward to form a funnel structure. The bottom of the perfusion incubation chamber bottom 1 is provided with a perfusion fluid outlet 5. There is a filter layer 6 between the funnel structure and the perfusion chamber body 2 above. The filter layer 6 is configured to prevent the leakage of live cells or ex vivo tissue and ensure that the perfusion culture medium flows smoothly. The outer edge of the perfusion incubation chamber bottom 1 extends upward to at least partially accommodate the perfusion chamber body 2.

[0008] The lower edge of the irrigation chamber 2 is pressed against the upper surface of the filter layer 6 under the vertical pressure and lateral resistance generated by the pressure fastening structure 3, forming a sealed connection with the bottom of the irrigation incubation chamber 1, and preventing the bottom of the irrigation incubation chamber 1 and the irrigation chamber 2 from displacing each other in the horizontal direction.

[0009] The perfusion chamber 2 is provided with an incubation chamber 9, which is configured as an incubation and culture space for live cell tissues. The incubation chamber 9 is connected to the perfusion fluid outlet 5 through the filter layer 6. The upward opening of the perfusion chamber 2 is configured to inject biological samples into the incubation chamber 9. After the injection is completed, the perfusion chamber 2 is sealed and connected to the upper sealing cover 4 above.

[0010] The upper sealing cap 4 has a perforation fluid inlet 13 through it, and the upper sealing cap 4 extends at least partially into the incubation chamber 9. The perforation fluid inlet 13 communicates with the incubation chamber 9.

[0011] The perfusion culture medium enters the incubation chamber 9 through the perfusion inlet 13 and flows out through the perfusion outlet 5 through the filter layer 6 below the incubation chamber 9, completing the perfusion process for living cells and tissues. The filter layer 6 is configured to prevent leakage of living cells or ex vivo tissues and ensure smooth flow of the perfusion culture medium.

[0012] In a preferred embodiment, the lower edge of the irrigation chamber 2 has a 45° inclined surface structure 8, and a sealing ring 7 (O-ring) is provided on the bottom of the irrigation incubation chamber bottom 1. After using the O-ring, the irrigation chamber 2 can achieve a sealing effect between the irrigation chamber 2, the irrigation incubation chamber bottom 1, and the filter layer 6 when subjected to downward pressure. The sealing ring 7 is configured to effectively prevent leakage.

[0013] In a preferred embodiment, the upper sealing cap 4 has a screw-like structure, with an inlet 13 penetrating through the screw portion. The screw portion extends into the incubation chamber 9 of the infusion chamber 2, and the inlet 13 communicates with the incubation chamber 9. The infusion chamber 2 and the upper sealing cap 4 are connected by a nut and screw structure. The upper surface of the infusion chamber 2 is provided with a sealing ring groove 11 that mates with a sealing ring 7 (O-ring). The sealing ring 7 is configured to ensure a tight connection between the infusion chamber 2 and the upper sealing cap 4, preventing liquid leakage.

[0014] In a preferred embodiment, the structure of the pressure fastening structure 3 and the downward pressure generated, combined with the sealing ring 7 (O-ring), can achieve the effect of sealing the periphery of the filter layer 6 and prevent the bottom 1 of the perfusion incubation chamber and the perfusion chamber body 2 from displacing relative to each other in the horizontal direction. The pressure fastening structure 3 is configured to effectively ensure that the filter layer 6 is flat, undamaged, and wrinkle-free, and to prevent leakage of ex vivo perfused cells.

[0015] In another preferred embodiment, the pressure fastening structure 3 changes shape when combined with the bottom 1 of the irrigation incubation chamber and the body 2 of the irrigation chamber. The pressure fastening structure 3 becomes a long positioning screw 14 structure. The body 2 of the irrigation chamber has multiple symmetrical positioning screw holes 15 (at least 2) at multiple locations. The positioning screw 14 can pass through the positioning screw holes 15. The outer side of the bottom 1 of the irrigation incubation chamber has a positioning screw hole with internal thread to act as a nut. Alternatively, the bottom 1 of the irrigation incubation chamber has the positioning screw hole 15, through which the positioning screw 14 can pass, and a nut is provided on the lower surface for fastening. Combined with the use of a sealing ring 7 (O-ring), the periphery of the filter layer 6 is sealed to prevent the bottom 1 of the irrigation incubation chamber and the body 2 of the irrigation chamber from displacing each other in the horizontal direction.

[0016] In another preferred embodiment, the upper edge of the funnel structure of the bottom 1 of the irrigation incubation chamber and the lower edge of the irrigation chamber body 2 are still circular, matching the sealing ring 7 (O-ring). The inner upper edge of the bottom 1 of the irrigation incubation chamber and the outer side of the middle part of the irrigation chamber body 2 are polygonal in shape, which cannot rotate between each other. When the pressure fastening structure 3 generates downward pressure, it achieves the purpose of preventing the bottom 1 of the irrigation incubation chamber and the irrigation chamber body 2 from displacing each other in the horizontal direction. The pressure fastening structure 3 can achieve the purpose of sealing the periphery of the filter layer 6 as long as it generates pressure.

[0017] In another preferred embodiment, the structures of the irrigation incubation chamber bottom 1 and the irrigation chamber body 2 are modified to achieve a pressurized and secure connection: the irrigation chamber body 2 is first separated into two parts: a fixing cap 17 and an incubation chamber shell 16. The outer edge of the irrigation incubation chamber bottom 1 extends upward, completely enclosing the incubation chamber shell 16. The incubation chamber shell 16 is a cylindrical structure, with its outer diameter being approximately equal to the inner diameter of the upwardly extending portion of the irrigation incubation chamber bottom 1, leaving only a small gap between them. The upper edge of the incubation chamber shell 16 is smooth, and the outer side of the lower edge of the incubation chamber shell 16 is a 45° inclined surface structure 8. When pressure is applied to the upper part, its lower edge makes a tight seal with the sealing ring 7 (O-ring) to ensure the sealing effect between the lower edge of the incubation chamber shell 16 and the filter layer 6 at the bottom of the infusion incubation chamber 1, preventing liquid leakage and ensuring that the filter layer 6 is flat without wrinkles or damage; the inner wall side of the upper end of the infusion incubation chamber 1 has an internal thread 10, the fixing cap 17 is a screw-like structure, the outer wall side of its lower screw part has an external thread 12, and the outer side of the lower edge of the screw part is a 45° inclined structure 8. After adding the sealing ring 7 (O-ring) between the fixing cap 17 and the incubation chamber shell 16, it can be connected to the bottom of the infusion incubation chamber. The upper internal thread 10 is screwed tightly closed under pressure. This sealing structure is configured to completely seal the edges of the upper and lower ends of the incubation chamber shell 16 by applying pressure to prevent side leakage. The fixing cap 17 has a cylindrical cavity running vertically through its center. The screw portion at the lower end of the upper sealing cover 4 can extend into this cylindrical cavity and is tightly closed by the internal thread 10 on the inner wall of the cylindrical cavity and the external thread 12 on the outer wall of the screw portion of the upper sealing cover 4. At the same time, an O-ring is placed in the sealing groove 11 on the fixing cap 17 to prevent liquid leakage. This is to ensure the seal between the incubation chamber shell 16 and the bottom 1 of the infusion incubation chamber. The filter layers 6 do not undergo horizontal displacement. The outer side of the upper edge of the incubation chamber shell 16 has a laterally protruding ear 19 structure. The upper inner surface of the infusion incubation chamber bottom 1 has a vertically recessed positioning groove 18 structure. When the ear 19 is inserted into the positioning groove 18, the incubation chamber shell 16 and the infusion incubation chamber bottom 1 cannot undergo horizontal displacement. In particular, when the fixing cap 17 is tightened downwards, the pressurized incubation chamber shell 16 can only move vertically downwards until it presses the O-ring 7 and the filter layer 6, thereby preventing liquid leakage and ensuring that the filter layer is flat and wrinkle-free.

[0018] In another preferred embodiment, the filter layer 6 is filter paper or nylon mesh.

[0019] In another preferred embodiment, the irrigation fluid outlet 5 is connected to the irrigation fluid collection device.

[0020] In the embodiments of this application, the closed design includes a perfusion incubation chamber bottom 1, a filter layer 6, a perfusion chamber body 2, an upper sealing cover 4, and a pressure fastening structure 3, which can effectively prevent the evaporation of the perfusion culture medium and overcome the problem of unstable temperature and incubation environment in open perfusion systems. The filter layer 6 at the bottom of the perfusion incubation chamber bottom 1 separates live cells or ex vivo tissues from the perfusion medium, ensuring the purity of the perfusion medium collection and improving the accuracy and reliability of the experiment. The connection design between the perfusion medium inlet / outlet and the incubation chamber 9 ensures the stability of the perfusion medium flow during the experiment. With the help of multiple sealing rings 7 (O-rings), the problem of liquid leakage caused by loose structure and the resulting systematic error is solved, thereby achieving stable and accurate collection of perfusion medium samples and meeting the high precision and high reliability requirements of the experiment.

[0021] Furthermore, the lower edge of the perfusion chamber 2 has a 45° inclined surface structure 8. After using an O-ring, the perfusion chamber 2 can achieve a sealing effect between the perfusion chamber 2, the bottom of the perfusion incubation chamber 1, and the filter layer 6 when the perfusion chamber 2 is subjected to downward pressure, thereby effectively preventing leakage and further improving the overall sealing performance and experimental reliability of the device. At the same time, it simplifies the mechanical structure and facilitates the machining of the device.

[0022] Furthermore, the structure of the pressurized fastening structure 3 and the downward pressure it generates prevent horizontal displacement between the bottom 1 of the perfusion incubation chamber and the body 2 of the perfusion chamber. This maintains the stable positioning of the two structures and effectively ensures that the filter layer 6 is flat, undamaged, and wrinkle-free, preventing cell leakage.

[0023] Furthermore, the upper sealing cover 4 has a screw-like structure, with an inlet for the irrigation fluid penetrating through the screw section at its lower end. The screw section extends into the incubation chamber 9 of the irrigation chamber 2, and the inlet 13 for the irrigation fluid communicates with the incubation chamber 9. Here, the irrigation chamber 2 and the upper sealing cover 4 are connected by a nut and screw structure. A sealing groove 11 is formed on the upper surface of the irrigation chamber 2 around the central axis, and a sealing ring 7 (O-ring) is embedded inside, thereby ensuring a tight connection between the irrigation chamber 2 and the upper sealing cover 4 and preventing liquid leakage.

[0024] The specification of this application contains numerous technical features distributed across various technical solutions. Listing all possible combinations of these technical features (i.e., technical solutions) would make the specification excessively lengthy. To avoid this problem, the various technical features disclosed in the above-described invention, the various technical features disclosed in the following embodiments and examples, and the various technical features disclosed in the accompanying drawings can be freely combined to form various new technical solutions (all of which are considered to have been described in this specification), unless such a combination of technical features is technically infeasible. For example, one example discloses feature A+B+C, and another example discloses feature A+B+D+E. Features C and D are equivalent technical means that serve the same function, and technically only one needs to be used; they cannot be used simultaneously. Feature E can technically be combined with feature C. Therefore, the solution A+B+C+D should not be considered as described because it is technically infeasible, while the solution A+B+C+E should be considered as described. Attached Figure Description

[0025] Figure 1 This is a schematic diagram of a closed-loop live cell tissue perfusion culture device according to one embodiment of this application.

[0026] Figure 2 This is a schematic diagram of a closed-loop live cell tissue perfusion culture device according to one embodiment of this application.

[0027] Figure 3 This is a schematic diagram of a closed-loop live cell tissue perfusion culture device according to one embodiment of this application.

[0028] Explanation of reference numerals in the attached figures:

[0029] 1. Bottom of the perfusion incubation chamber; 2. Perfusion chamber body; 3. Pressurization and fastening structure; 4. Upper sealing cover; 5. Perfusion fluid outlet; 6. Filter layer; 7. Sealing ring; 8. 45° inclined structure; 9. Incubation chamber; 10. Internal thread; 11. Sealing ring groove; 12. External thread; 13. Perfusion fluid inlet; 14. Positioning screw; 15. Positioning screw hole; 16. Incubation chamber shell; 17. Fixing cap; 18. Positioning groove; 19. Ear; Detailed Implementation

[0030] In the following description, many technical details are presented to help the reader better understand this application. However, those skilled in the art will understand that the technical solutions claimed in this application can be implemented even without these technical details and various variations and modifications based on the following embodiments.

[0031] As used in this article, the terms "sealing ring" and "O-ring" are used interchangeably.

[0032] To make the objectives, technical solutions, and advantages of this application clearer, the embodiments of this application will be described in further detail below with reference to the accompanying drawings.

[0033] This application relates to a closed-loop live cell tissue perfusion culture device, the structural diagram of which is shown below. Figure 1 As shown, it includes: the bottom of the irrigation incubation chamber 1, the irrigation chamber body 2, the pressurization and fastening structure 3, the upper sealing cover 4, and the filter layer 6.

[0034] The bottom inner side of the perfusion incubation chamber 1 is recessed downward to form a funnel structure, and the bottommost end of the bottom is provided with a perfusion fluid outlet 5. There is a filter layer 6 between the funnel structure and the perfusion chamber body 2 above. The filter layer 6 is configured to prevent the leakage of live cells or ex vivo tissue and ensure that the perfusion culture medium flows smoothly. The outer edge of the perfusion incubation chamber 1 extends upward to at least partially accommodate the perfusion chamber body 2 above.

[0035] The lower edge of the irrigation chamber 2 is pressed against the upper surface of the filter layer 6 under the vertical pressure and lateral resistance generated by the pressure fastening structure 3, and forms a sealed connection with the bottom of the irrigation incubation chamber 1, preventing horizontal displacement between the bottom of the irrigation incubation chamber 1 and the irrigation chamber 2.

[0036] The perfusion chamber 2 is equipped with an incubation chamber 9, which is configured as an incubation and culture space for live cell tissues. The incubation chamber 9 communicates downward through the filter layer 6 with the perfusion fluid outlet 5. The upward opening of the perfusion chamber 2 is set to inject biological samples (such as perfusion fluid, live cells or ex vivo tissues and other experimental materials) into the incubation chamber 9. After injection, it is sealed with the upper sealing cover 4 with a screw and nut structure.

[0037] The upper sealing cap 4 has a screw-shaped structure, with a perfusion liquid inlet 13 penetrating through the screw section at its lower end. The screw section extends into the incubation chamber, and the perfusion liquid inlet 13 communicates with the incubation chamber 9.

[0038] The perfusion culture medium enters the incubation chamber 9 through the perfusion inlet 13 and flows out through the perfusion outlet 5 through the filter layer 6 below the incubation chamber 9, completing the perfusion process for living cells and tissues. The filter layer 6 here is configured to prevent leakage of living cells or ex vivo tissues and ensure smooth flow of the perfusion culture medium.

[0039] In an optional embodiment, the lower edge of the irrigation chamber 2 has a 45° inclined structure 8. After using the sealing ring 7 (O-ring), when the irrigation chamber 2 is subjected to downward pressure, the irrigation chamber 2 can achieve a sealing effect between itself, the bottom of the irrigation incubation chamber 1, and the filter layer 6, which can effectively prevent leakage.

[0040] In an optional embodiment, the upper sealing cover 4 has a screw-like structure, with an inlet 13 for the irrigation fluid penetrating through the screw portion at its lower end. The screw portion extends into the incubation chamber 9, and the inlet 13 communicates with the incubation chamber 9. The irrigation chamber 2 and the upper sealing cover 4 are connected by a nut and screw structure. A sealing ring groove 11 is formed on the upper surface of the irrigation chamber 2 around the central axis. The sealing ring 7 is embedded in the sealing ring groove 11 to ensure a tight connection between the irrigation chamber 2 and the upper sealing cover 4 and prevent liquid leakage.

[0041] In an optional embodiment, the structure of the pressure fastening structure 3 and the resulting downward pressure, combined with the sealing ring 7 (O-ring), can achieve a tight seal around the edge of the filter layer 6 and prevent the bottom 1 of the perfusion incubation chamber and the perfusion chamber body 2 from shifting relative to each other in the horizontal direction. This effectively ensures that the filter layer 6 is flat and free of wrinkles or damage, and can better prevent leakage of ex vivo perfused cells.

[0042] In another optional embodiment, the pressure fastening structure 3 changes shape during its combination with the bottom 1 of the irrigation incubation chamber and the body 2 of the irrigation chamber. The pressure fastening structure 3 becomes a long positioning screw 14 structure. Symmetrical positioning screw holes 15 (at least 2) are provided at multiple positions on the body 2 of the irrigation chamber. The positioning screw 14 can be inserted into the positioning screw hole 15. The positioning screw hole 15 with internal thread is provided on the edge of the bottom 1 of the irrigation incubation chamber to act as a nut. Alternatively, the positioning screw hole 15 is provided on the edge of the bottom 1 of the irrigation incubation chamber, through which the positioning screw 14 can pass and a nut is provided on the lower surface for fastening. Combined with the use of the sealing ring 7 (O-ring), the periphery of the filter layer 6 is sealed to prevent the bottom 1 of the irrigation incubation chamber and the body 2 of the irrigation chamber from displacing each other in the horizontal direction.

[0043] In another optional embodiment, the upper edge of the funnel at the bottom of the incubation chamber 1 and the lower edge of the incubation chamber 2 are still circular, matching the sealing ring 7 (O-ring). The upper inner edge of the bottom of the incubation chamber and the outer side of the middle part of the incubation chamber are polygonal in shape, which cannot rotate with each other. When the pressure fastener structure 3 generates downward pressure, it achieves the purpose of preventing mutual displacement between the bottom of the incubation chamber 1 and the incubation chamber 2 in the horizontal direction. The pressure fastener structure 3 can achieve the purpose of sealing the periphery of the filter layer 6 as long as it generates pressure.

[0044] In another optional embodiment, the bottom of the irrigation incubation chamber 1 has a cavity that fully accommodates the incubation chamber shell 16. The height of the bottom of the irrigation incubation chamber 1 is greater than the height of the incubation chamber shell 16. The bottom of the irrigation incubation chamber 1 is provided with a positioning groove 18 for mounting the protruding ear 19 of the upper edge of the incubation chamber shell 16 into the positioning groove 18, so as to prevent horizontal displacement between the two structures, the incubation chamber shell 16 and the bottom of the irrigation incubation chamber 1.

[0045] In another optional embodiment, a fixing cap 17 is also included. The lower protrusion of the fixing cap 17 presses against the upper surface of the incubation chamber shell 16. A cylindrical cavity runs through the fixing cap 17. The upper screw structure of the fixing cap 17 is screwed tightly to the upper nut structure of the incubation chamber bottom 1, and a sealed connection is formed between the fixing cap 17, the incubation chamber bottom 1, and the incubation chamber shell 16 by means of an O-ring.

[0046] In another optional embodiment, the structures of the irrigation incubation chamber bottom 1 and the irrigation chamber body 2 are modified and combined to achieve a pressurized and secure effect: the irrigation chamber body 2 is first separated into two parts, a fixing cap 17 and an incubation chamber shell 16. The upper edge of the irrigation incubation chamber bottom 1 extends upward and completely encloses and accommodates the incubation chamber shell 16. The incubation chamber shell 16 has a cylindrical structure with a smooth upper edge and a 45° beveled structure 8 on the outer side of the lower edge. When pressure is applied to the upper part, it makes a tight seal with the sealing ring 7 (O-ring) to ensure that the lower part is in close contact with the inside of the irrigation incubation chamber bottom 1. The airtightness of filter layer 6 prevents liquid leakage and ensures the flatness of filter layer 6 inside the bottom 1 of the incubation chamber. The inner wall of the upper end of the bottom 1 of the incubation chamber has internal threads 10, and the fixing cap 17 has a screw-like structure (the upper part is a screw and the lower part is a threaded rod). The outer wall of the threaded part of the fixing cap 17 has external threads 12. The outer side of the lower edge of the fixing cap 17 has a 45° bevel structure 8. After the sealing ring 7 (O-ring) is embedded in the sealing ring groove 11, it can be tightly screwed into contact with the upper part of the bottom 1 of the incubation chamber and the upper edge of the incubation chamber shell 16. The sealing ring 7 can completely seal the upper edge of the incubation chamber shell 16 to prevent side leakage; while the fixing cap 17 has a cylindrical cavity running vertically through its central axis, through which the screw portion of the upper sealing cover 4 can pass, and the fixing cap 17 relies on the internal thread 10 on the inner wall of its cylindrical cavity to tighten with the external thread 12 on the outer wall of the screw portion of the upper sealing cover 4 to form a sealed structure and prevent liquid leakage; to ensure that there is no horizontal mutual displacement between the incubation chamber shell 16 and the filter layer 6 of the bottom 1 of the perfusion incubation chamber, the incubation chamber shell 16 The upper outer edge has a laterally protruding ear 19 structure, and the upper inner side of the bottom of the incubation chamber 1 has a vertically recessed positioning groove 18 structure. When the ear 19 is embedded in the positioning groove 18, the incubation chamber shell 16 and the bottom of the incubation chamber 1 cannot be horizontally displaced. In particular, when the fixing cap 17 is tightened downwards, the incubation chamber shell 16, which is subjected to downward pressure, can only move vertically downwards until it presses the sealing ring 7 (O-ring) and the filter layer 6, thereby preventing liquid leakage and ensuring that the filter layer 6 is flat without wrinkles or damage.

[0047] In an optional embodiment, the perfusion outlet 5 is connected to a perfusion collection device. Depending on different experimental requirements, the perfusion outlet 5 can be designed to be compatible with various types of collection devices, such as sealed bottles, test tubes, microfluidic chips, or online analysis devices. This connection can be a detachable flexible hose connection, a locking interface, or a fixed rigid pipe to adapt to different experimental environments. Furthermore, the perfusion outlet 5 can be configured as a multi-way distribution system, allowing the perfusion to flow simultaneously to multiple different collection devices for parallel analysis or segmented experiments. In some applications, the connection between the perfusion outlet 5 and the collection device can also be configured with a flow control valve or an automated sampling device to achieve precise fluid control and sample collection, further improving experimental efficiency and data reliability.

[0048] To better understand the technical solution of this application, several specific examples are provided below. The details listed in these examples are mainly for ease of understanding and are not intended to limit the scope of protection of this application.

[0049] Example 1

[0050] like Figure 1 As shown, this invention proposes a closed-loop live cell tissue perfusion culture device, which can ensure a stable incubation environment and constant fluid flow for live cells and ex vivo tissues, thereby improving the success rate of experiments and minimizing losses during perfusion fluid collection. This closed-loop live cell tissue perfusion culture device consists of the following parts: a perfusion incubation chamber bottom 1, a perfusion chamber body 2, a pressure-locking structure 3, an upper sealing cover 4, and a filter layer 6;

[0051] The bottom of the perfusion incubation chamber 1 is recessed downwards into a shallow funnel shape, and the bottommost end of the bottom is provided with a perfusion fluid outlet 5. There is a filter layer 6 between the funnel structure and the perfusion chamber above, which is configured to prevent the leakage of live cells or ex vivo tissue and ensure that the perfusion culture medium flows smoothly. The outer edge of the perfusion incubation chamber bottom 1 extends upwards to at least partially accommodate the perfusion chamber 2 above.

[0052] The lower edge of the irrigation chamber 2 is pressed against the upper surface of the filter layer 6 under the vertical pressure and lateral resistance generated by the pressure fastening structure 3, forming a sealed connection with the bottom of the irrigation incubation chamber 1, and preventing horizontal displacement between the bottom of the irrigation incubation chamber 1 and the irrigation chamber 2.

[0053] The perfusion chamber 2 is equipped with an incubation chamber 9, which is configured as an incubation and culture space for live cell tissues. The incubation chamber 9 is connected to the perfusion fluid outlet 5 through the filter layer 6. The upward opening of the perfusion chamber 2 is set to inject perfusion fluid, live cells or ex vivo tissues and other experimental materials into the incubation chamber 9. After injection, it is sealed with the upper sealing cover 4 with a screw and nut structure.

[0054] The upper sealing cap 4 has a screw-shaped structure, with an inlet 13 for the perfusion fluid channel penetrating through its screw section. The screw section extends into the incubation chamber 9, and the perfusion fluid inlet 13 communicates with the incubation chamber 9.

[0055] The perfusion culture medium enters the incubation chamber 9 through the perfusion inlet 13 and flows out through the perfusion outlet 5 through the filter layer 6 below the incubation chamber 9, completing the perfusion process for living cells and tissues. The filter layer 6 here is configured to prevent leakage of living cells or ex vivo tissues and ensure smooth flow of the perfusion culture medium.

[0056] To further ensure a good seal, the lower edge of the infusion chamber 2, where it contacts the sealing ring 7 (O-ring), is designed with a 45° bevel structure 8. Additionally, in the connection structure between the infusion chamber 2 and the upper sealing cover 4, a sealing ring groove 11 is formed on the upper surface of the infusion chamber 2 around the central axis. Both of these designs help optimize the sealing performance of the sealing ring 7 (O-ring), ensuring a tight connection between the infusion chamber 2 and the upper sealing cover 4 and preventing liquid leakage.

[0057] To further ensure the structural and functional integrity of the filter layer 6, in addition to the 45° inclined structure 8 at the lower edge of the irrigation chamber 2 and the sealing ring 7 (O-ring) to achieve a tight connection, the inner diameters of the contact points between the irrigation chamber bottom 1 and the irrigation chamber 2 are required to be similar, with only a small gap between them. In addition, the pressure fastening structure 3 ensures that the irrigation chamber bottom 1 and the irrigation chamber 2 are vertically pressed together and prevents horizontal displacement.

[0058] In summary, this embodiment provides an improved closed-loop live cell tissue perfusion culture device. Through reasonable structural design and sealing measures, it can effectively improve the stability and reliability of perfusion experiments and is suitable for perfusion culture experiments of various cells or tissues.

[0059] Example 2

[0060] like Figure 2 As shown, this embodiment provides a closed live cell tissue perfusion culture device. Based on the previous embodiment, the pressure-fastening structure is further replaced with positioning screws 14 (at least two), and corresponding positioning screw holes 15 are symmetrically arranged at the edges of the perfusion incubation chamber bottom 1 and the perfusion chamber body 2. By tightening the positioning screws 14, the perfusion chamber body 2 is pressed tightly downwards against the inner bottom of the perfusion incubation chamber bottom 1, thereby improving the overall stability and sealing performance of the perfusion culture device. It is suitable for live cell or ex vivo tissue perfusion experiments where higher device stability is required.

[0061] The overall structure of the device remains basically unchanged, but the location of the positioning screw hole 15 should be selected at the edge as much as possible. At the same time, in order to ensure the balance and stability of the device structure, the positioning screw hole 15 should be set at a position that is symmetrical to each other on the plane, with at least two.

[0062] Example 3

[0063] like Figure 3 As shown, this embodiment provides a closed-loop live cell tissue perfusion culture device improved based on Embodiment 1. Compared with Embodiment 1, this embodiment has undergone structural modifications, redesigning the perfusion incubation chamber bottom 1 and perfusion chamber body 2 in the original perfusion chamber structure to enhance the device's sealing, stability, and ease of operation: the perfusion chamber body 2 is first separated into a fixing cap 17 and an incubation chamber shell 16, with the upper edge of the perfusion incubation chamber bottom 1 extending upwards to completely enclose the incubation chamber shell 16; the incubation chamber shell 16 is a cylindrical structure, with its outer diameter being approximately equal to the inner diameter of the upwardly extending portion of the perfusion incubation chamber bottom 1, leaving only a small gap between them; the upper edge of the incubation chamber shell 16 is smooth, and the outer side of the lower edge is a 45° inclined structure 8, subjected to downward... After pressure, it makes a tight seal with the sealing ring 7 (O-ring) to ensure the sealing effect between the lower edge and the filter layer 6 inside the incubation chamber bottom 1, preventing liquid leakage and ensuring that the filter layer 6 is flat without wrinkles or damage; the fixing cap 17 is a screw-like structure (the upper part is a screw and the lower part is a screw rod), with external threads 12 on the screw rod part and a 45° inclined structure 8 on the outer side of the lower edge. The inner wall side of the upper end of the incubation chamber bottom 1 has internal threads 10. After the screw structure of the fixing cap 17 is tightened with the screw and the sealing ring 7 (O-ring) is added, it can be sealed to the incubation chamber bottom 1 and the incubation chamber shell 16, so that the edges of the upper and lower ends of the incubation chamber shell 16 are completely sealed to prevent side leakage;

[0064] The fixed cap 17 has a cylindrical cavity running vertically through its central axis, through which the screw part of the upper sealing cover 4 can pass. The internal thread 10 on the inner surface of the cylindrical cavity engages with the external thread 12 of the screw of the upper sealing cover 4 to tighten and seal the cavity, preventing liquid leakage.

[0065] To prevent horizontal displacement between the incubation chamber shell 16 and the filter layer 6 of the incubation chamber bottom 1, there is a laterally protruding ear 19 structure on the outer side of the upper edge of the incubation chamber shell 16, and a vertically recessed positioning groove 18 structure on the upper inner surface of the incubation chamber bottom 1. When the ear 19 is inserted into the positioning groove 18 of the incubation chamber bottom 1, horizontal displacement between the incubation chamber shell 16 and the incubation chamber bottom 1 is impossible. In particular, when the fixing cap 17 is tightened downwards, the pressurized incubation chamber shell 16 can only move vertically downwards until it presses the sealing ring 7 (O-ring) and the filter layer 6. The pressure is evenly distributed at all positions on the edge of the filter layer 6, which achieves the purpose of preventing liquid leakage and ensuring that the filter layer 6 is flat without wrinkles or damage.

[0066] By redesigning the structure of the perfusion incubation chamber bottom 1 and the perfusion chamber body 2, this embodiment not only improves the sealing and stability of the perfusion culture device, but also enhances its flexibility and ease of operation in adapting to different experimental conditions. Furthermore, it reduces the difficulty of machining the entire device, improves the quality of batch processing, and ensures that reliable experimental data can still be provided under high precision requirements.

[0067] It should be noted that in this patent application, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Furthermore, the terms "comprising," "including," or any other variations thereof are intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitations, an element defined by the phrase "comprising one" does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element. In this patent application, if it refers to performing an action according to an element, it means performing the action at least according to that element, including two cases: performing the action only according to that element, and performing the action according to that element and other elements. Expressions such as "multiple," "repeatedly," and "various" include two, two times, two kinds, and more than two, more than two times, and more than two kinds.

[0068] All documents mentioned in this application are considered to be incorporated in their entirety into the disclosure of this application so that they can serve as a basis for modifications if necessary. Furthermore, it should be understood that after reading the foregoing disclosure of this application, those skilled in the art can make various alterations or modifications to this application, and these equivalent forms also fall within the scope of protection claimed in this application.

Claims

1. A closed-loop live cell tissue perfusion culture device, characterized in that, include: The bottom of the irrigation incubation chamber (1), the irrigation chamber body (2), the pressurization and fastening structure (3), the upper sealing cover (4), and the filter layer (6); The bottom inner side of the perfusion incubation chamber bottom (1) is recessed downward to form a funnel structure. The bottom of the perfusion incubation chamber bottom (1) is provided with a perfusion fluid outlet (5). There is a filter layer (6) between the funnel structure and the perfusion chamber body (2) above. The filter layer (6) is configured to prevent the leakage of live cells or ex vivo tissue and ensure that the perfusion culture medium flows smoothly. The outer edge of the perfusion incubation chamber bottom (1) extends upward to at least partially accommodate the perfusion chamber body (2). The lower edge of the perfusion chamber (2) is pressed against the upper surface of the filter layer (6) under the vertical pressure and lateral resistance generated by the pressure fastening structure (3), forming a sealed connection with the bottom of the perfusion incubation chamber (1), and preventing horizontal displacement between the lower edge of the perfusion chamber (2) and the bottom of the perfusion incubation chamber (1); the perfusion chamber (2) is provided with an incubation chamber (9), which is configured as an incubation and culture space for live cell tissues. The incubation chamber (9) is connected to the perfusion fluid outlet (5) through the filter layer (6). The opening of the perfusion chamber (2) is configured to inject biological samples into the incubation chamber (9). After the injection is completed, the perfusion chamber (2) is sealed with the upper sealing cover (4) above. The upper sealing cap (4) extends at least partially into the incubation chamber (9), and the upper sealing cap (4) has a perfusion liquid inlet (13) that communicates with the incubation chamber (9); The perfusion culture medium enters the incubation chamber (9) through the perfusion inlet (13) and flows out from the perfusion outlet (5) through the filter layer (6) below the incubation chamber (9), thus completing the perfusion process of living cell tissue.

2. The closed-loop live cell tissue perfusion culture device as described in claim 1, characterized in that, The lower edge of the irrigation chamber (2) has a 45° inclined surface structure (8), and a sealing ring (7) is provided on the bottom of the irrigation incubation chamber (1). When the irrigation chamber (2) is subjected to downward pressure, the sealing ring (7) achieves a sealing effect between the irrigation chamber (2), the irrigation incubation chamber (1), and the filter layer (6), preventing leakage.

3. The closed-loop live cell tissue perfusion culture device as described in claim 1, characterized in that, The structure of the pressurizing fastening structure (3) and the downward pressure generated prevent horizontal displacement between the bottom (1) of the irrigation incubation chamber and the body (2) of the irrigation chamber. The pressurizing fastening structure (3) is also configured to keep the filter layer (6) flat and wrinkle-free.

4. The closed-loop live cell tissue perfusion culture device as described in claim 1, characterized in that, The upper sealing cover (4) has a screw-like structure, with the inlet (13) of the perfusion liquid penetrating through the screw portion at its lower end. The screw portion extends into the incubation chamber (9) of the perfusion chamber (2). The inlet (13) of the perfusion liquid communicates with the incubation chamber (9). The perfusion chamber (2) and the upper sealing cover (4) are connected by a nut and screw structure. The upper surface of the perfusion chamber (2) is provided with a sealing ring groove (11) that cooperates with the sealing ring (7). The sealing ring is configured to ensure a tight connection between the perfusion chamber (2) and the upper sealing cover (4) to prevent liquid leakage.

5. The closed-loop live cell tissue perfusion culture device as described in claim 1, characterized in that, It also includes a positioning screw (14); The bottom (1) and body (2) of the irrigation incubation chamber are provided with a plurality of positioning screw holes (15). Each positioning screw hole (15) of the bottom (1) of the irrigation incubation chamber corresponds to the positioning screw hole (15) of the body (2). The positioning screw (14) extends from the positioning screw hole (15) of the body (2) into the positioning screw hole (15) of the bottom (1) of the irrigation incubation chamber. The positioning screw (14) is configured to fix the bottom (1) and body (2) of the irrigation incubation chamber, prevent the bottom (1) and body (2) of the irrigation incubation chamber from shifting in the horizontal direction, and make the body (2) of the irrigation incubation chamber sealed to the bottom (1).

6. The closed-loop live cell tissue perfusion culture device as described in claim 1, characterized in that, The bottom (1) of the irrigation incubation chamber completely accommodates the cavity of the incubation chamber shell (16). The height of the bottom (1) of the irrigation incubation chamber is greater than the height of the incubation chamber shell (16). The bottom (1) of the irrigation incubation chamber is provided with a positioning groove (18) for mounting the protruding ear (19) of the upper edge of the incubation chamber shell (16) into the positioning groove (18) so as to fix the two structures of the incubation chamber shell (16) and the bottom (1) of the irrigation incubation chamber to prevent horizontal displacement.

7. The closed-loop live cell tissue perfusion culture device as described in claim 6, characterized in that, It also includes a fixing cap (17), the lower end of which protrudes and presses against the upper surface of the incubation chamber shell (16). A cylindrical cavity runs through the fixing cap (17). The upper end of the fixing cap (17) is screwed together with the upper end of the nut structure of the irrigation incubation chamber bottom (1), and the fixing cap (17), the irrigation incubation chamber bottom (1), and the incubation chamber shell (16) are connected by an O-ring.

8. The closed-loop live cell tissue perfusion culture device as described in claim 1, characterized in that, The filter layer (6) is filter paper or nylon mesh.

9. The closed-loop live cell tissue perfusion culture device according to any one of claims 1-8, characterized in that, The irrigation fluid outlet (5) is connected to the irrigation fluid collection device.

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