CN115232750B - Sterile culture bag and method of use - Google Patents

Sterile culture bag and method of use Download PDF

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Publication number
CN115232750B
CN115232750B CN202211155855.2A CN202211155855A CN115232750B CN 115232750 B CN115232750 B CN 115232750B CN 202211155855 A CN202211155855 A CN 202211155855A CN 115232750 B CN115232750 B CN 115232750B
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joint
connector
connecting pipe
movable member
adapter tube
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CN115232750A (en
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邱雪峰
郭莹
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Haimai Medical Technology Suzhou Co ltd
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Haimai Medical Technology Suzhou Co ltd
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Abstract

The invention discloses an aseptic culture bag and a using method thereof, wherein the aseptic culture bag comprises: the bag body is provided with a culture space, and a first connector for inputting a first fluid and a second connector for outputting the first fluid are arranged on the bag body; the connection pipe is positioned in the culture space and is used for communicating the first joint and the second joint, and the pipe wall of the connection pipe can generate pulsation in the process that the first fluid passes through the connection pipe; the degradable bracket is positioned in the culture space, sleeved on the periphery of the connecting pipe and used for planting, adhering and growing cells; the sealing connection structure is arranged at the connection positions of the first joint, the second joint and the connecting pipe so as to form sealing connection between the first joint and the first end of the connecting pipe and between the second joint and the second end of the connecting pipe; wherein, the bag body is also provided with at least one inflow port and at least one outflow port.

Description

Sterile culture bag and method of use
Technical Field
The invention belongs to the technical field of medical instruments, and particularly relates to a sterile culture bag for tissues or organs and a using method thereof.
Background
Cell culture is an essential step in tissue and organ regeneration and the process can begin with seeding cells onto a scaffold and then submerging the scaffold in liquid cell culture medium. The culture bag is a carrier for tissue or organ culture, the culture bag is provided with a culture space, a bracket is arranged in the culture space and can contain liquid cell culture medium, and the bracket is immersed in the culture medium.
In the process of vascular tissue culture, an environment simulating the pulsation of human pulse is also required, and the pulsation environment can promote cells to generate tubular tissue objects, which are indispensable parts of vascular tissue culture. In the prior art, the culture bag is usually provided with a connecting tube capable of generating the above-mentioned pulsation, the connecting tube is used for introducing fluid, and the fluid generates pulsation on the tube wall of the connecting tube when passing through the connecting tube. In the process of the vascular tissue culture, the fluid in the connecting pipe is ensured not to leak into the culture space. However, in the prior art, the sealing effect of the joint of the adapter tube and the culture bag is poor, fluid in the adapter tube easily enters the culture space, the culture of vascular tissues is influenced, and the using effect is poor. Accordingly, there is a need for improvements in the art that overcome the deficiencies in the prior art.
Disclosure of Invention
Therefore, the technical problem to be solved by the invention is to provide a sterile culture bag with good use effect and a use method thereof.
In order to solve the above technical problems, the present invention provides an aseptic culture bag comprising: the bag body is provided with a culture space, and a first connector for inputting a first fluid and a second connector for outputting the first fluid are arranged on the bag body; an adapter tube positioned in the culture space and used for communicating the first connector and the second connector; the degradable bracket is positioned in the culture space, sleeved on the periphery of the connecting pipe and used for planting, adhering and growing cells; the sealing connection structure is arranged at the connection positions of the first joint, the second joint and the connecting pipe so as to form sealing connection between the first joint and the first end of the connecting pipe and between the second joint and the second end of the connecting pipe; wherein, still be equipped with at least one inflow port and at least one outflow port on the bag body.
Preferably, the sealing connection structure includes a first movable member and a second movable member that are in threaded connection with the first joint, the first movable member and the second movable member are both annular, in the axial direction of the first joint, the first movable member abuts against the second movable member, and the first movable member is distributed closer to the first joint than the second movable member;
a first external thread and a second external thread are arranged on the outer circumferential surface of the first joint, the first external thread is matched with the first moving piece, the second external thread is matched with the second moving piece, the first external thread and the second external thread are adjacently distributed, and the rotation directions of the first external thread and the second external thread are opposite;
in the axial direction of the first joint, the inner ring of the second movable member comprises an internal thread area and a unthreaded hole area, the internal thread area is distributed closer to the first joint than the unthreaded hole area, and a seam between the first joint and the first end of the connecting pipe is positioned at the junction of the internal thread area and the unthreaded hole area;
the internal thread area is matched with the second external thread, and the unthreaded hole area is abutted to the outer wall of the first end of the connection pipe.
Preferably, an accommodating groove which is opened towards the second movable piece is formed in the first movable piece, and the second movable piece is partially accommodated in the accommodating groove;
the first moving part comprises a side plate and an annular wall which is arranged on the side plate and distributed along the circumferential direction of the first joint, the side plate and the annular wall are arranged in a surrounding mode to form the accommodating groove, a threaded hole is formed in the side plate, and the first moving part is connected with the first external thread through the threaded hole; the inner ring of the annular wall is abutted against the outer peripheral wall of the second movable piece.
Preferably, the inner ring of the annular wall is a conical surface, and the inner diameter of the annular wall is gradually reduced in the direction from the connecting pipe to the first joint;
the second movable piece comprises a conical part matched with the accommodating groove and a holding part arranged on the conical part and positioned outside the accommodating groove, a through hole penetrating through the conical part and the holding part along the axial direction of the first joint is further formed in the second movable piece, and the hole wall of the through hole is an inner ring of the second movable piece.
Preferably, the holding part abuts against the notch of the accommodating groove and forms a sealing connection with the notch of the accommodating groove;
and a sealing element is arranged on the end face of the holding part, which is abutted against the notch of the accommodating groove.
Preferably, a seam between the first joint and the first end of the adapter tube is located in the receiving groove, the distance from the seam to the bottom of the receiving groove is L1, the distance from the seam to the notch of the receiving groove is L2, where L1: the value range of L2 is 0.2-1.
Preferably, a plug part extending along the axial direction of the connecting pipe is arranged at the first end of the connecting pipe, and the plug part can be inserted into the first joint;
the outer diameter of the inserting portion is smaller than that of the connecting pipe, a step-shaped limiting portion is formed between the inserting portion and the first end of the connecting pipe, and one end of the first joint abuts against the step-shaped limiting portion.
Preferably, a sealing convex part extending along the circumferential direction of the connecting pipe is arranged on the outer wall of the connecting part, and the sealing convex part is abutted against the inner wall of the first joint;
the outer circumferential surface of the sealing convex part sequentially comprises a first guide surface, an abutting surface and a second guide surface in the direction from the first end of the connecting pipe to the connecting part, wherein the abutting surface abuts against the inner wall of the first joint and connects the first guide surface with the second guide surface;
the outer diameter of the first guide surface gradually increases in a direction from the first end of the adapter tube to the socket, the outer diameter of the abutment surface remains unchanged, and the outer diameter of the second guide surface gradually decreases.
Preferably, a gap is formed between the outer wall of the connecting pipe and the inner wall of the degradable bracket, and the value range of the gap is 0-5 cm.
Preferably, the material of the connecting pipe is silica gel; or,
the material of the connecting pipe is ethylene-vinyl acetate copolymer; or,
the material of the connecting pipe is fluorinated ethylene propylene copolymer; or,
the connecting pipe is made of Teflon; or,
the material of the connecting pipe is polyvinyl chloride; or,
the material of the connecting pipe is nylon.
Preferably, the first connector is externally connected with a pulsating pump for leading the first fluid to enter the adapter tube, and the pulsating pump is connected with the second connector through a pipeline, so that a circulation loop is formed among the first connector, the adapter tube, the second connector and the pulsating pump;
wherein the pressure of the pulsating pump increases with the increase of the culture time, and the value range of the pressure of the pulsating pump is 0-500 mmHg.
Preferably, the bag body is further provided with a sampling port, the sampling port is connected with a detection unit, and the detection unit is configured to detect the pH value, the temperature, the dissolved oxygen concentration and the carbon dioxide concentration of the liquid in the culture space in real time or discontinuously, and judge whether the liquid in the culture space has pathogens or not;
wherein, a sampling port is also arranged on the bag body, and the liquid in the culture space can be manually extracted from the sampling port; or,
the liquid in the culture space can be extracted from the sampling opening in an automatic manner; or,
the liquid in the culture space can be extracted from the sampling port in a manner of combining manual operation and automatic operation.
Preferably, the tube wall of the adapter tube is capable of pulsating during the passage of the first fluid through the adapter tube; wherein, the flow mode of the first fluid in the connecting pipe is continuous or pulsating.
The invention also provides a using method of the sterile culture bag, which comprises the following steps:
sleeving the degradable support on the connecting pipe;
the first end of the connecting pipe is connected to the first joint of the bag body in a sealing way by using a sealing connection structure, and the second end of the connecting pipe is connected to the second joint of the bag body in a sealing way, so that the connecting pipe and the degradable support are arranged in the bag body;
inputting a first fluid into the connecting pipe, and simultaneously conveying a culture medium, a cell suspension and carbon dioxide gas into the culture space of the bag body through the inflow port;
after the cells are successfully inoculated, introducing a cell removing solution into the culture space through the inflow port;
after the cell removal operation, introducing a cleaning solution into the culture space through the inflow port;
successfully cultured tissue on the adapter tube was removed.
Preferably, in the process of connecting the connecting pipe to the first connector and the second connector of the pouch body using the sealing connection structure, the steps of:
firstly, a first moving part of a sealing connection structure is screwed on a first external thread of a first joint, and a second moving part is sleeved at a first end and a second end of an adapting pipe;
then inserting the inserting parts at the two ends of the connecting pipe into the first joint and the second joint respectively;
and then, the second moving piece is pushed to enable the second moving piece to be partially inserted into the accommodating groove of the first moving piece, and the second moving piece is rotated to enable the internal thread area on the second moving piece to be matched with the second external thread on the first joint, so that the second moving piece is abutted against the first moving piece.
Preferably, the process of removing the successfully cultured tissue from the adapter tube comprises the following steps:
firstly, cutting off the bag body;
then, rotating the first movable piece on the first joint to enable the first movable piece to move towards the direction far away from the second movable piece;
then, the patch part at the first end of the connecting pipe is pulled out of the first connector;
and finally, rotating the first moving part on the second joint to enable the first moving part to move towards the direction far away from the second moving part, and then rotating the second moving part to enable the second moving part to be separated from the second joint.
The technical scheme provided by the invention has the following advantages:
1. the first connector and the first end of the connecting pipe and the second connector and the second end of the connecting pipe can be in sealed connection through the sealed connection structure, so that the space in the connecting pipe and the culture space of the bag body are two mutually independent spaces, the first fluid in the connecting pipe is prevented from influencing the culture of blood vessels in the culture space, and the sealed connection structure has the advantage of good use effect;
2. when the second moving part rotates, under the matching action of the inner thread area and the second outer thread, the second moving part can prop against the first moving part in the process of moving along the connecting pipe to the first joint direction, and under the action of friction force, the first moving part has a trend of rotating in the same direction as the second moving part, but because the directions of the two threads matched with the first joint are opposite, the rotating trend of the first moving part can cause the first moving part and the second moving part to tightly abut against each other in the axial direction of the first joint, and the sealing performance is good.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings needed to be used in the embodiments or the technical solutions in the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 is a schematic structural view of a sterile culture bag provided by the present invention;
FIG. 2 is a schematic view of the connection between the first connector, the sealing connection structure and the connecting tube;
FIG. 3 is an exploded view of FIG. 2;
FIG. 4 is an exploded view of FIG. 3;
FIG. 5 is a schematic cross-sectional view of FIG. 2;
FIG. 6 is an enlarged schematic view of region A in FIG. 5;
FIG. 7 is a schematic view showing that the outer diameter of the first joint is larger than that of the adapter tube;
FIG. 8 is a perspective view of the second movable member;
FIG. 9 is a schematic cross-sectional view of the second movable member;
FIG. 10 is a schematic perspective view of the first movable member;
FIG. 11 is a schematic view of the construction of the adapter tube;
fig. 12 is an enlarged schematic view of a region B in fig. 11.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the accompanying drawings, and it is to be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. The invention will be described in detail hereinafter with reference to the drawings and embodiments. It should be noted that the embodiments and features of the embodiments may be combined with each other without conflict.
It should be noted that the terms "first," "second," and the like in the description and claims of the present invention and in the drawings described above are used for distinguishing between similar elements and not necessarily for describing a particular sequential or chronological order.
In the present invention, unless stated to the contrary, the use of directional terms such as "upper, lower, top, bottom" or the like, generally refers to the orientation of the components as shown in the drawings, or to the vertical, perpendicular, or gravitational orientation of the components themselves; likewise, for ease of understanding and description, "inner and outer" refer to the inner and outer relative to the profile of the components themselves, but the above directional words are not intended to limit the invention.
Example 1
The invention provides an aseptic culture bag, which is a culture bag subjected to sterilization operation, and is aseptic without viable bacteria. Only under the precondition that the culture bag is sterile, the cells can be adhered, grown and proliferated to finally obtain the required tissue.
In one illustrative scenario, the sterile culture bag may be used for tissue engineering vascular culture. The sterile culture bag in the above example is used for culturing human blood vessels, and is only one possible application scenario. In other possible and not explicitly excluded scenarios, the sterile culture bag may also be used as a culture for other tubular tissues including, but not limited to, human and mammalian ureters, trachea, esophagus, bile ducts, intestinal ducts, nerve ducts, vas deferens, fallopian tubes, lymphatic vessels, and the like; the sterile culture bag can also be used for culturing solid organs such as heart, liver, lung, kidney and the like. The following description will mainly describe the scenario where the sterile culture bag is used for tissue engineering blood vessel culture. It will nevertheless be understood that no limitation of the scope of the embodiments of the invention is thereby intended, as indicated by the foregoing description.
Referring to FIG. 1, the sterile culture bag comprises: bag body 100, connecting pipe 200, degradable support 300 and sealing connection structure 400. The bag body 100 has a culture space in which the adapter tube 200, the degradable holder 300 and the sealing connection structure 400 are accommodated. The degradable stent 300 is sleeved on the periphery of the connection tube 200 and used for cell planting, adhesion and growth.
Considering that the bag 100 is not only a tissue culture container but also a tissue decellularization container, the bag 100 needs to have a resistance to corrosion or destruction by agents such as detergents, decellularization agents, sterilization agents, and preservatives.
In this embodiment, the pouch 100 is made of a biocompatible material that is flexible. For example, the material of the bag body 100 may be ultra-low density polyethylene, linear low density polyethylene, fluorinated ethylene polymer, or the like. Of course, the material of the bag body 100 includes, but is not limited to, the above materials, and other medical materials that can resist corrosion or damage by agents such as detergents, decellularization agents, sterilization agents, preservation agents, etc.
The bag 100 is provided with a first connector 110 for inputting the first fluid and a second connector 120 for outputting the first fluid. The first fluid may be a liquid or a gas. The adapter tube 200 is used to communicate the first connector 110 with the second connector 120. Wherein a first end of the adapter tube 200 is connected to the first connector 110 and a second end of the adapter tube 200 is connected to the second connector 120. The adaptor tube 200 is made of non-degradable material, such as silica gel, polyvinyl chloride, nylon, teflon, fluorinated ethylene propylene copolymer, ethylene-vinyl acetate copolymer, ultra-low density polyethylene, polyvinylidene fluoride, ethyl vinyl acetate, etc. It is understood that the material of the connecting tube 200 includes, but is not limited to, the above materials, which are not described in detail herein.
In this embodiment, the first connector 110 and the second connector 120 are respectively disposed at both ends of the bag body 100 in the longitudinal direction. The line connecting the first connector 110 and the second connector 120 is parallel to the adapter tube 200. Further, the first joint 110, the second joint 120 and the adapter tube 200 are coaxially arranged.
The first tab 110 and the second tab 120 are both partially located in the culture space of the bag 100 and partially located outside the bag 100. The portion of the first connector 110 located outside the pouch 100 is connected to a pump (not shown) located outside the pouch 100, and the pump is used for delivering the first fluid located outside the pouch 100 into the adapter tube 200.
When the first fluid enters the adapter tube 200, the first fluid impacts the tube wall of the adapter tube 200, causing the tube wall to vibrate, thereby generating pulsations. That is, the tube wall of the adapter tube 200 can generate pulsation during the first fluid passes through the adapter tube 200.
The type of pump body determines the flow pattern of the first fluid in the adapter tube 200, wherein the pump body may be a pulsating pump or a flat flow pump. When the pump body is a pulsating pump, the flow mode of the first fluid in the connecting pipe 200 is in a pulsating mode; when the pump body is a constant flow pump, the flow pattern of the first fluid in the connection pipe 200 is continuous.
In this embodiment, the pump body is preferably a pulsatile pump. The pulsating pump is located between the first connector 110 and the second connector 120, and selectively communicates the first connector 110 with the second connector 120, so that a circulation loop is formed between the first connector 110, the connecting tube 200, the second connector 120 and the pulsating pump. Thereby, the first fluid can be circulated in the joint pipe 200. Of course, the circulation circuit also comprises a storage structure for providing said first fluid, which storage structure is part of said circulation circuit. The above "selectively communicating the first connector 110 with the second connector 120" can be understood as: when the pulsation pump works, the first connector 110 is communicated with the second connector 120; when the pulsation pump stops working, the first joint 110 is not communicated with the second joint 120.
The parts of the circulation circuit located outside the bag body 100 are connected to each other through a pipe, for example, the pulse pump is connected to the reservoir structure through a pipe, the first joint 110 is connected to the pulse pump through a pipe, and the second joint 120 is connected to the reservoir structure through a pipe. It will be appreciated that when there are a plurality of bags, the lines of the plurality of bags may be connected in parallel. In this embodiment, under the action of the pulsatile pump, the first fluid in the storage structure enters the adapter tube 200, flows back to the storage structure after passing through the adapter tube 200, and is continuously circulated.
In one embodiment, the adaptor tube 200 is preferably a flexible thin-walled teflon tube, whereby the pulsations generated by the adaptor tube 200 are better transferred to the degradable stent 300. The pulsations generated by the above-described adapter tube 200 serve to act as a mechanical stimulus, causing the cells on the degradable scaffold 300 to produce extracellular matrix, forming a strong tubular tissue containing cells.
The process of culturing blood vessels with cells takes several weeks, generally about 6-12 weeks. The pressure of the pulsating pump varies with time throughout the culture, and specifically, the pressure of the pulsating pump increases with the increase of the culture time, wherein the pressure of the pulsating pump has a value ranging from 0 to 500mmHg.
For example, in the first week, 3 to 5 days, the pressure of the pulse pump is 10 to 20mmHg; in the second week, at the stage of 8-10 days, the pressure of the pulsation pump is 20-30 mmHg; in the third week, at the stage of 17-19 days, the pressure of the pulsation pump is 30-45 mmHg; in the fourth week, at the stage of 23-25 days, the pressure of the pulsating pump is 45-60 mmHg; in the fifth week, at the stage of 31-33 days, the pressure of the pulsation pump is 60-80 mmHg; in the sixth week, at the stage of 37-39 days, the pressure of the pulsation pump is 80-110 mmHg; in the seventh week, at the stage of 45-47 days, the pressure of the pulsation pump is 110-150 mmHg; in the eighth week, the pressure of the pulse pump is 150 to 200mmHg at 53 to 55 days. From the above, the pulsating pump pressure was increased after the culture stage.
There is a gap between the outer wall of the adapter tube 200 and the inner wall of the degradable scaffold 300. The gap between the outer wall of the adapter tube 200 and the inner wall of the degradable stent 300 has a value ranging from 0 to 5cm. Under the above value range, the existence of the gap does not affect the transmission of the pulsation generated by the connection tube 200 to the degradable stent 300.
Specifically, the value of the gap may be 0cm, 1cm, 2cm, 3cm, 4cm, 5cm, or the like, or may be an increase in the interval between 0 and 5cm by 0.1cm, 0.2cm, 0.3cm, 0.4cm, 0.5cm, 0.6cm, 0.7cm, 0.8cm, 0.9 cm. These are only examples of what is intended to be explicitly recited, and all possible combinations of numerical values between the lowest value and the highest value that are explicitly recited in the specification in a similar manner are to be considered.
In this embodiment, the bag 100 is further provided with at least one inlet port 600 and at least one outlet port 700, the inlet port 600 is used for allowing the fluid to enter the culture space of the bag 100, and the outlet port 700 is used for allowing the fluid to be discharged from the culture space of the bag 100. It should be noted that, in this embodiment, the fluid in the culture space of the bag 100 is referred to as a second fluid. The second fluid is liquid or a mixed fluid of liquid and a small amount of gas.
The second fluid introduced into the culture space through the inflow port 600 may be a culture medium, a cell suspension, a cell-removing solution, a cleaning solution, carbon dioxide gas, or the like. Similarly, the fluid flowing out of the culture space through the outflow port 700 may be a culture medium, a cell suspension, a decellularization solution, a cleaning solution, carbon dioxide gas, or the like.
The total number of ports and connectors on the bag 100 may be 4, 5, 6, or other numbers. Taking the case that the total number of ports and connectors on the bag 100 is 5 as an example, please refer to fig. 1, where the number of connectors on the bag 100 is two, that is, the first connector 110 and the second connector 120; the number of inflow ports 600 is two, and the number of outflow ports 700 is one. One of the two inflow ports 600 is an inflow port for the culture medium, and the other is an inflow port for the carbon dioxide gas. The inflow and outflow ports 700 are disposed opposite to each other, and the "opposing arrangement" can be understood as the inflow and outflow ports 700 are disposed back-to-back. Wherein the inflow and outflow ports 700 for the culture medium are provided at the edge of the bag body 100 under the degradable stent 300, which will help to drain the culture medium and remove the residual cells and tissues not attached to the degradable stent 300 during the culture.
Wherein the inflow port of the culture medium is a multi-functional port through which the cell suspension can be delivered into the culture space of the bag body 100. The second fluid in the culture space is discharged through the outflow port 700. It is to be noted that the decellularization solution and the washing solution also enter the culture space through the inflow port of the medium and then flow out through the outflow port.
In some cases, the inflow port of the carbon dioxide gas may be replaced by a breather hole provided in the bag body 100. Particularly, the bag body 100 is provided with a through hole, the through hole is covered with a waterproof breathable film, and the through hole and the waterproof breathable film form the breathing hole. The breathing hole can guarantee the gas exchange between bag body 100 and the surrounding culture environment to can guarantee that bag body 100 is interior to be aseptic environment.
When the bag 100 is used, it is placed in a culture apparatus (not shown) filled with a predetermined carbon dioxide concentration, and carbon dioxide gas enters the culture space of the bag 100 through the breathing holes. In this example, the carbon dioxide concentration in the culture space was maintained at 5%. When the inflow port of carbon dioxide gas adopts the breathing hole to replace, the total number of ports and joints on the bag body 100 is 4, and the value range of the number of the breathing holes is 1-6. It should be noted that when the bag body 100 is made of waterproof and breathable material, the bag body 100 does not need to be provided with breathing holes due to the characteristics of the waterproof and breathable material.
In this embodiment, the sealing connection structure 400 is provided at the connection positions of the first joint 110, the second joint 120 and the adapter tube 200, so that the first joint 110 and the first end of the adapter tube 200, and the second joint 120 and the second end of the adapter tube 200 form a sealing connection therebetween. Thus, the first fluid in the adapter tube 200 does not leak from the connection into the culture space, affecting the normal growth and proliferation of the cells.
Considering that the connection structure between the first connector 110 and the first end of the adapter tube 200 and the connection structure between the second connector 120 and the second end of the adapter tube 200 are the same, only one of the first connector 110 and the second connector 120 will be explained in describing the sealing connection structure 400. The following description will be made mainly on the connection relationship between the first joint 110, the first end of the adapter tube 200, and the sealing connection structure 400.
Referring to fig. 2 to 6, the sealing connection structure 400 includes a first movable member 410 and a second movable member 420 threadedly coupled to the first joint 110. The first movable member 410 and the second movable member 420 are annular, and in the axial direction of the first joint 110, the first movable member 410 abuts against the second movable member 420, and the first movable member 410 is distributed closer to the first joint 110 than the second movable member 420. The above "the first movable member 410 is disposed closer to the first joint 110 than the second movable member 420" may be understood as the second movable member 420 is disposed closer to the connecting tube 200 than the first movable member 410.
In the present embodiment, please refer to fig. 6, a first external thread 111 and a second external thread 112 are disposed on an outer circumferential surface of the first joint 110. The first external threads 111 engage the first movable member 410 and the second external threads 112 engage the second movable member 420. The first external thread 111 and the second external thread 112 are disposed adjacent to each other, and the rotation directions of the first external thread 111 and the second external thread 112 are opposite.
Referring to fig. 9, in the axial direction of the first joint 110, the inner ring of the second movable member 420 includes an internal threaded region 421 and an unthreaded hole region 422, the internal threaded region 421 is distributed closer to the first joint 110 than the unthreaded hole region 422, and a seam M between the first joint 110 and the first end of the connecting tube 200 is located at a boundary between the internal threaded region 421 and the unthreaded hole region 422. The internal threaded region 421 mates with the second external thread 112 and the unthreaded region 422 abuts the outer wall of the first end of the adapter tube 200.
The first external thread 111 and the second external thread 112 not only realize structural connection, but also ensure the sealing connection between the first joint 110 and the first moving part 410 and between the first joint 110 and the second moving part 420, and have the advantages of reliable connection and good sealing performance.
When the second movable member 420 is rotated, under the cooperation of the internal thread region 421 and the second external thread 112, the second movable member 420 will abut against the first movable member 410 during the movement along the direction from the connecting pipe 200 to the first joint 110, and under the action of friction, the first movable member 410 has a tendency to rotate in the same direction as the second movable member 420. However, since the two threads are engaged with the first joint 110 in opposite directions, the above-mentioned rotation tendency of the first movable member 410 causes the first movable member 410 and the second movable member 420 to be tightly abutted in the axial direction of the first joint 110, thereby having an advantage of good sealing performance.
In the present embodiment, please refer to fig. 6, 8 and 10, an accommodating groove opened toward the second movable member 420 is disposed on the first movable member 410, a portion of the second movable member 420 is accommodated in the accommodating groove, a portion of the second movable member 420 located outside the accommodating groove forms a holding portion 424, and the holding portion 424 is used for an operator to hold, so that the operator can rotate the second movable member 420.
With regard to the structure of the first movable member 410, please refer to fig. 10, the first movable member 410 includes a side plate 411 and an annular wall 412 disposed on the side plate 411 and distributed along the circumferential direction of the first joint 110, and the side plate 411 and the annular wall 412 surround to form the above-mentioned receiving slot. Wherein, a threaded hole 413 is opened on the side plate 411, and the first movable member 410 is connected with the first external thread 111 through the threaded hole 413. The inner ring of the annular wall 412 abuts the outer peripheral wall of the second movable member 420.
In this embodiment, the annular wall 412 can constrain the second movable member 420 in the radial direction of the first joint 110, so that the second movable member 420 is tightly wrapped at the connection between the first joint 110 and the adapter tube 200 in the circumferential direction of the first joint 110, and the second fluid in the culture space can be effectively prevented from permeating through the aperture area 422 to the connection between the first joint 110 and the adapter tube 200. It can be seen that the second fluid in the culture space cannot permeate into the adapter tube 200 through the connection between the first connector 110 and the adapter tube 200. Similarly, the first fluid in the adapter tube 200 cannot permeate into the culture space through the joint between the first connector 110 and the adapter tube 200, so that the space in the adapter tube 200 and the culture space are two mutually independent spaces, which is beneficial to culture blood vessels and has the advantage of good use effect.
A seam M between the first connector 110 and the first end of the connecting tube 200 is located in the receiving groove, a distance between the seam M and the bottom of the receiving groove is L1, and a distance between the seam M and the notch of the receiving groove is L2. Since the seam M is located at the intersection of the female threaded region 421 and the unthreaded hole region 422, L1 can also be understood as the length of the female threaded region 421 in the axial direction of the first joint 110.
In the present embodiment, L1: the value range of L2 is 0.2-1, preferably, L1: the ratio of L2 is 0.5. In the present embodiment, the length of the light hole region 422 in the axial direction of the first connector 110 is the sum of L2 and the length of the holding portion 424. The above-mentioned "length of the grip portion 424" refers to the length of the grip portion 424 in the axial direction of the first joint 110. As can be seen from the above, the length of the unthreaded hole region 422 is greater than the length of the internal threaded region 421. The length of the light hole region 422 is set to be greater than the length of the internal thread region 421 to mean: in order to improve the sealing between the perforated region 422 and the adapter tube 200.
When the ratio of L1: when the ratio of L2 is 0.5, the ratio of the lengths of the female screw region 421, the unthreaded hole region 422, and the through-hole 425 in the axial direction along the first joint 110 is 1:2.5:3.5, thereby, the connection strength of the internal thread region 421 and the second external thread 112 can be ensured, and simultaneously, the sufficient contact area between the unthreaded region 422 and the outer wall of the adapter tube 200 can be ensured, so that the sealing effect can be achieved. When the aperture area 422 abuts against the outer wall of the adapter tube 200, the annular wall 412 restricts the second movable member 420, so that the second movable member 420 is tightly wrapped at the joint between the first joint 110 and the adapter tube 200, and the sealing effect is further improved.
The inner circumference of the annular wall 412 is a conical surface, and the inner diameter of the annular wall 412 is gradually reduced in the direction from the adapter tube 200 to the first joint 110. The conical surface of the annular wall 412 is provided to facilitate the second movable member 420 entering the receiving groove, thereby facilitating the connection of the internally threaded region 421 with the second external thread 112.
Referring to fig. 8, the second movable member 420 includes a conical portion 423 engaged with the receiving groove, and the holding portion 424 disposed on the conical portion 423 and located outside the receiving groove.
The second movable member 420 is further provided with a through hole 425 penetrating the conical portion 423 and the holding portion 424 in the axial direction of the first joint 110, and a hole wall of the through hole 425 is an inner ring of the second movable member 420. That is, the wall of the through hole 425 is provided with the female screw region 421 and the unthreaded hole region 422.
Referring to fig. 6, when the outer diameter of the first joint 110 is equal to the outer diameter of the connection tube 200, the through hole 425 is a circular hole. Referring to fig. 7, when the outer diameter of the first connector 110 is larger than that of the adapter tube 200, the through hole 425 is a stepped hole.
In the present embodiment, please refer to fig. 6 and 7, the holding portion 424 abuts against the notch of the receiving groove, a sealing member 500 is disposed on an end surface of the holding portion 424 abutting against the notch of the receiving groove, and the sealing member 500 enables the holding portion 424 and the notch of the receiving groove to form a sealing connection. Thereby, the sealing performance between the first movable member 410 and the second movable member 420 is further improved. Wherein, the sealing member 500 is a rubber sealing ring.
Referring to fig. 11, a first end of the connection tube 200 is provided with a patch portion 210 extending in an axial direction of the connection tube 200, an outer diameter of the patch portion 210 is smaller than an outer diameter of the connection tube 200, and the patch portion 210 can be inserted into the first connector 110.
A step-shaped limiting portion is formed between the plug portion 210 and the first end of the connecting tube 200, and one end of the first connector 110 abuts against the step-shaped limiting portion. The abutting contact between the first joint 110 and the step-shaped limiting portion is the position of the seam M.
In order to improve the sealing performance between the patch section 210 and the first connector 110, a sealing protrusion 220 extending in the circumferential direction of the adapter tube 200 is provided on the outer wall of the patch section 210, and the sealing protrusion 220 abuts against the inner wall of the first connector 110.
With regard to the structure of the sealing convex portion 220, referring to fig. 12, the outer circumferential surface of the sealing convex portion 220 sequentially includes a first guide surface 221, an abutment surface 222, and a second guide surface 223 in a direction from the first end of the adapter tube 200 to the connection portion 210, wherein the abutment surface 222 abuts against the inner wall of the first joint 110 and connects the first guide surface 221 and the second guide surface 223. In the direction from the first end of the adapter tube 200 to the socket 210, the outer diameter of the first guide surface 221 gradually increases, the outer diameter of the abutment surface 222 remains unchanged, and the outer diameter of the second guide surface 223 gradually decreases.
The first guide surface 221 and the second guide surface 223 are tapered surfaces, the contact surface 222 is a cylindrical surface, and the outer diameter of the contact surface 222 is substantially the same as the outer diameter of the first joint 110. In this embodiment, the first guide surface 221 and the second guide surface 223 are tapered surfaces, which can facilitate the insertion or extraction of the first connector 110 into or from the patch panel 210, and the first connector 110 is not easily damaged, which has the advantage of easy assembly and disassembly.
In addition, when the plug 210 is inserted into the first connector 110 by the sealing protrusion 220, the plug 210 and the first connector 110 are not easily displaced relative to each other in the axial direction even when an external force is applied thereto, and the plug 210 is not easily detached from the first connector 110, which provides an advantage of stable and reliable connection.
In this embodiment, the pouch 100 is any size and shape structure capable of containing tissue, tubular tissue structures, organs, and the like. The pouch 100 may be transparent or include a transparent portion to allow an operator to visually observe the inside of the pouch 100 during the incubation period.
The bag 100 is further provided with a sampling port (not shown) for sampling the liquid in the culture space. Wherein the sampling port may be one of the outflow ports 700, i.e. one of the outflow ports 700 is the above-mentioned sampling port; alternatively, the sampling port may be a separate port independent from the outflow port 700.
The sampling port is connected to a detection unit (not shown) configured to detect the pH, temperature, dissolved oxygen, and carbon dioxide concentration of the liquid in the culture space in real time or intermittently, and determine the period and frequency of medium replacement by determining the presence or absence of a pathogen in the liquid in the culture space. The "pathogen" refers to bacteria, fungi, mycoplasma, chlamydia, etc.
As for the way of sampling, the first way, the liquid in the culture space can be manually extracted from the sampling port; in a second mode, the liquid in the culture space can be extracted from the sampling port in an automatic mode; in a third way, the liquid in the culture space can be extracted from the sampling opening in a manual and automatic combined manner.
Regarding the mode of culture medium change, likewise, there are also manual mode, automatic mode and manual and automatic mode that combines, and the operator can select according to actual demand for use. During the cultivation process, the operator periodically drains all or part of the culture medium from the bag 100 and replaces it with fresh culture medium.
In this embodiment, the bag body 100 is further provided with a liquid feeding tube (not shown) located outside the bag body 100, and the liquid feeding tube can accelerate the culture medium to flow into or out of the bag body 100, which helps to improve the work efficiency.
The bag body 100 is further provided with a first connection pipe (not shown) connected to the breathing hole and a second connection pipe (not shown) for cell seeding. Wherein, the liquid feeding pipe, the first connecting pipe and the second connecting pipe are aseptic pipes.
Example 2
The invention also provides a using method of the sterile culture bag, which is the using method of the sterile culture bag in the embodiment 1. The following description will be given taking as an example a case where the total number of ports and connectors on the bag body 100 is 5.
Referring to fig. 1, the hollow arrows in fig. 1 represent the flow direction of the fluid. Specifically, the use method of the sterile culture bag comprises the following steps:
1) Sleeving the degradable stent 300 on the connecting pipe 200;
2) The first end of the connecting pipe 200 is hermetically connected to the first connector 110 of the bag body 100 by using the sealing connection structure 400, and the second end of the connecting pipe 200 is hermetically connected to the second connector 120 of the bag body 100, so that the connecting pipe 200 and the degradable support 300 are installed in the bag body 100;
3) Feeding a first fluid into the adapter tube 200 while feeding a culture medium, a cell suspension and carbon dioxide gas into the culture space of the bag body 100 through the inflow port 600;
4) After the cells are successfully inoculated, introducing a decellularization reagent into the culture space through the inflow port 600;
5) After the cell removal operation, a cleaning solution is introduced into the culture space through the inflow port 600;
6) The tissue that has been successfully cultured on the adapter tube 200, which is a cultured blood vessel, is removed.
In the process of connecting the connecting pipe 200 to the first connector 110 and the second connector 120 of the pouch body 100 using the sealing connection structure 400, the following steps are further included:
first, the first movable member 410 of the sealing connection structure 400 is screwed on the first external thread 111 of the first joint 110, and the second movable member 420 is sleeved at the first end and the second end of the connection pipe 200;
then, the patch parts 210 at both ends of the connecting tube 200 are respectively inserted into the first joint 110 and the second joint 120;
then, the second movable member 420 is pushed to partially insert the second movable member 420 into the receiving groove of the first movable member 410, and the second movable member 420 is rotated to match the internal thread region 421 with the second external thread 112, so that the second movable member 420 abuts against the first movable member 410.
In this embodiment, the following steps are also included in the process of removing the successfully cultured tissue from the adaptor tube 200:
first, the bag body 100 is cut;
then, the first movable member 410 on the first joint 110 is rotated to make the first movable member 410 move towards the direction away from the second movable member 420, at this time, the circumferential direction of the second movable member 420 is not restrained, and the second movable member 420 is not tightly wrapped on the outer wall of the connecting pipe 200;
then, the inserting portion 210 of the connecting tube 200 is pulled out from the first connector 110, and since the second movable member 420 is not tightly wrapped on the outer wall of the connecting tube 200, the operator can easily pull out the inserting portion 210 from the first connector 110;
finally, the first movable member 410 on the second joint 120 is rotated to move the first movable member 410 toward a direction away from the second movable member 420, and then the second movable member 420 is rotated to separate the second movable member 420 from the second joint 120. During the process of disengaging the second movable member 420 from the second connector 120, the second movable member 420 can push the blood vessel on the adapter tube 200 to slide toward the first connector 110 due to the movement of the second movable member 420 in a direction away from the second connector 120. That is, the second movable member 420 also has a function of facilitating the separation of the blood vessel from the engaging tube 200, thereby facilitating the removal of the blood vessel, which is advantageous in terms of convenience of use.
It is to be understood that the above-described embodiments are only a few, but not all, embodiments of the present invention. Based on the embodiments of the present invention, those skilled in the art may make other variations or modifications without creative efforts, and shall fall within the protection scope of the present invention.

Claims (9)

1. An aseptic culture bag, comprising:
a bag body (100) having a culture space, the bag body (100) being provided with a first connector (110) for inputting a first fluid and a second connector (120) for outputting the first fluid;
an adapter tube (200) located in the culture space for communicating the first connector (110) with the second connector (120);
the degradable bracket (300) is positioned in the culture space, sleeved on the periphery of the connecting pipe (200) and used for planting, adhering and growing cells;
a sealing connection structure (400) located in the culture space, wherein the sealing connection structure (400) is arranged at the connection positions of the first connector (110), the second connector (120) and the adapter tube (200) so as to form a sealing connection between the first connector (110) and the first end of the adapter tube (200) and between the second connector (120) and the second end of the adapter tube (200);
wherein, the bag body (100) is also provided with at least one inflow port (600) and at least one outflow port (700);
the sealing connection structure (400) comprises a first movable part (410) and a second movable part (420) which are in threaded connection with the first joint (110), the first movable part (410) and the second movable part (420) are annular, in the axial direction of the first joint (110), the first movable part (410) abuts against the second movable part (420), and the first movable part (410) is distributed closer to the first joint (110) than the second movable part (420);
a first external thread (111) and a second external thread (112) are arranged on the outer circumferential surface of the first joint (110), the first external thread (111) is matched with the first movable piece (410), the second external thread (112) is matched with the second movable piece (420), the first external thread (111) and the second external thread (112) are adjacently distributed, and the rotation directions of the first external thread (111) and the second external thread (112) are opposite;
in the axial direction of the first joint (110), the inner ring of the second movable part (420) comprises an internally threaded region (421) and a unthreaded region (422), the internally threaded region (421) being distributed closer to the first joint (110) than the unthreaded region (422), the seam between the first joint (110) and the first end of the adapter tube (200) being located at the intersection of the internally threaded region (421) and the unthreaded region (422);
the internally threaded region (421) mates with the second externally threaded region (112), the unthreaded region (422) abutting an outer wall of the first end of the adapter tube (200);
an accommodating groove which is opened towards the second movable piece (420) is formed in the first movable piece (410), and the second movable piece (420) is partially accommodated in the accommodating groove;
the first movable piece (410) comprises a side plate (411) and an annular wall (412) which is arranged on the side plate (411) and distributed along the circumferential direction of the first joint (110), the side plate (411) and the annular wall (412) are arranged in a surrounding mode to form the accommodating groove, a threaded hole (413) is formed in the side plate (411), and the first movable piece (410) is connected with the first external thread (111) through the threaded hole (413);
the inner ring of the annular wall (412) abuts against the outer peripheral wall of the second movable member (420);
the inner circle of the annular wall (412) is a conical surface, and the inner diameter of the annular wall (412) is gradually reduced in the direction from the adapter tube (200) to the first joint (110);
the second movable piece (420) comprises a conical part (423) matched with the accommodating groove and a holding part (424) arranged on the conical part (423) and positioned outside the accommodating groove, a through hole (425) penetrating through the conical part (423) and the holding part (424) along the axial direction of the first joint (110) is further formed in the second movable piece (420), and the hole wall of the through hole (425) is an inner ring of the second movable piece (420);
the holding part (424) is abutted against the notch of the containing groove and is in sealing connection with the notch of the containing groove;
wherein, a sealing element (500) is arranged on the end surface of the holding part (424) which is abutted against the notch of the containing groove;
a plug-in portion (210) extending along an axial direction of the connection tube (200) is arranged at a first end of the connection tube (200), and the plug-in portion (210) can be inserted into the first connector (110);
the outer diameter of the connecting part (210) is smaller than that of the connecting pipe (200), a step-shaped limiting part is formed between the connecting part (210) and the first end of the connecting pipe (200), and one end of the first joint (110) abuts against the step-shaped limiting part;
a sealing convex part (220) extending along the circumferential direction of the connecting pipe (200) is arranged on the outer wall of the connecting part (210), and the sealing convex part (220) is abutted with the inner wall of the first joint (110);
the outer peripheral surface of the sealing convex part (220) sequentially comprises a first guide surface (221), an abutting surface (222) and a second guide surface (223) in the direction from the first end of the adapter tube (200) to the plug part (210), wherein the abutting surface (222) abuts against the inner wall of the first joint (110) and connects the first guide surface (221) and the second guide surface (223);
the outer diameter of the first guide surface (221) is gradually increased in a direction from the first end of the adapter tube (200) to the plug portion (210), the outer diameter of the abutment surface (222) is kept unchanged, and the outer diameter of the second guide surface (223) is gradually decreased;
a gap is formed between the outer wall of the linking pipe (200) and the inner wall of the degradable bracket (300), and the value range of the gap is 0-5 cm;
wherein, the using method of the sterile culture bag comprises the following steps: sleeving the degradable bracket (300) on the connecting pipe (200);
the first end of the connecting pipe (200) is connected to the first connector (110) of the bag body (100) in a sealing mode through the sealing connection structure (400), the second end of the connecting pipe (200) is connected to the second connector (120) of the bag body (100) in a sealing mode, and therefore the connecting pipe (200) and the degradable support (300) are installed in the bag body (100);
feeding a first fluid into the adapter tube (200), and simultaneously feeding a culture medium, a cell suspension and carbon dioxide gas into the culture space of the bag body (100) through the inflow port (600);
after the cells are successfully inoculated, introducing a cell removal solution into the culture space through the inflow port (600);
after the cell removal operation, a washing solution is introduced into the culture space through the inflow port (600);
removing the successfully cultured tissue from the adapter tube (200);
wherein, in the process of connecting the connecting pipe (200) to the first connector (110) and the second connector (120) of the bag body (100) by using the sealing connection structure (400), the method comprises the following steps:
firstly, screwing a first movable member (410) of a sealing connection structure (400) on a first external thread (111) of a first joint (110), and sleeving a second movable member (420) at a first end and a second end of a connection pipe (200);
then, inserting the inserting parts (210) at the two ends of the connecting pipe (200) onto the first joint (110) and the second joint (120) respectively;
then, the second movable member (420) is pushed to enable the second movable member (420) to be partially inserted into the accommodating groove of the first movable member (410), and the second movable member (420) is rotated to enable the internal thread area (421) on the second movable member (420) to be matched with the second external thread (112) on the first joint (110), so that the second movable member (420) abuts against the first movable member (410).
2. The sterile culture bag of claim 1,
a seam between the first joint (110) and the first end of the adapter tube (200) is located in the receiving groove, a distance between the seam and a groove bottom of the receiving groove is L1, a distance between the seam and a groove opening of the receiving groove is L2, wherein L1: the value range of L2 is 0.2-1.
3. The sterile culture bag of claim 1,
the material of the connecting pipe (200) is silica gel; or,
the material of the connecting pipe (200) is ethylene-vinyl acetate copolymer; or,
the material of the connecting pipe (200) is fluorinated ethylene propylene copolymer; or,
the material of the connecting pipe (200) is Teflon; or,
the material of the connecting pipe (200) is polyvinyl chloride; or,
the material of the connecting pipe (200) is nylon.
4. The sterile culture bag of claim 1,
the first connector (110) is externally connected with a pulsating pump for leading the first fluid to enter the adapter tube (200), and the pulsating pump is connected with the second connector (120) through a pipeline, so that a circulation loop is formed among the first connector (110), the adapter tube (200), the second connector (120) and the pulsating pump;
wherein the pressure of the pulsation pump increases with the increase of the culture time, and the pressure of the pulsation pump has a value ranging from 0 to 500mmHg.
5. The sterile culture bag of claim 1,
the bag body (100) is also provided with a sampling port, the sampling port is connected with a detection unit, and the detection unit is configured to detect the pH value, the temperature, the dissolved oxygen concentration and the carbon dioxide concentration of the liquid in the culture space in real time or discontinuously and judge whether the liquid in the culture space has pathogens or not;
wherein the liquid in the culture space can be manually extracted from the sampling port; or,
the liquid in the culture space can be extracted from the sampling opening in an automatic manner; or,
the liquid in the culture space can be extracted from the sampling opening in a manual and automatic combined manner.
6. The sterile culture bag according to claim 1 wherein the tube wall of the adapter tube (200) is capable of pulsating during the passage of the first fluid through the adapter tube (200);
wherein the flow of the first fluid in the adapter tube (200) is continuous or pulsating.
7. The method of using the sterile culture bag of claim 1, comprising:
sleeving the degradable support (300) on the connecting pipe (200);
the first end of the connecting pipe (200) is connected to the first connector (110) of the bag body (100) in a sealing mode through the sealing connection structure (400), the second end of the connecting pipe (200) is connected to the second connector (120) of the bag body (100) in a sealing mode, and therefore the connecting pipe (200) and the degradable support (300) are installed in the bag body (100);
feeding a first fluid into the adapter tube (200), and simultaneously feeding a culture medium, a cell suspension and carbon dioxide gas into the culture space of the bag body (100) through the inflow port (600);
after the inoculation of the cells is successful, a cell removal solution is introduced into the culture space through the inflow port (600);
after the cell removal operation, a washing solution is introduced into the culture space through the inflow port (600);
successfully cultured tissue on the adapter tube (200) is removed.
8. The method of using an aseptic culture bag as defined in claim 7,
in the process of connecting the connecting pipe (200) to the first connector (110) and the second connector (120) of the bag body (100) by using the sealing connection structure (400), the method comprises the following steps:
firstly, a first movable piece (410) of a sealing connection structure (400) is screwed on a first external thread (111) of a first joint (110), and a second movable piece (420) is sleeved at a first end and a second end of a connecting pipe (200);
then, inserting the inserting parts (210) at the two ends of the connecting pipe (200) onto the first joint (110) and the second joint (120) respectively;
then, the second movable member (420) is pushed to enable the second movable member (420) to be partially inserted into the accommodating groove of the first movable member (410), and the second movable member (420) is rotated to enable the internal thread area (421) on the second movable member (420) to be matched with the second external thread (112) on the first joint (110), so that the second movable member (420) abuts against the first movable member (410).
9. The method of using a sterile culture bag according to claim 7,
in the process of removing the successfully cultured tissue from the adapter tube (200), the method comprises the following steps:
firstly, cutting off a bag body (100);
then, rotating the first movable member (410) on the first joint (110) to enable the first movable member (410) to move towards a direction far away from the second movable member (420);
subsequently, the patch part (210) at the first end of the adapter tube (200) is pulled out of the first connector (110);
finally, the first movable piece (410) on the second joint (120) is rotated to enable the first movable piece (410) to move towards the direction far away from the second movable piece (420), then the second movable piece (420) is rotated to enable the second movable piece (420) to be separated from the second joint (120), and in the process that the second movable piece (420) is separated from the second joint (120), the second movable piece (420) moves towards the direction far away from the second joint (120).
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Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1067403A (en) * 1991-06-03 1992-12-30 美国管道和塑料制品公司 The method and apparatus of installing pipe liner
KR20000022284A (en) * 1996-06-27 2000-04-25 나우톤 질 케이. Apparatus and method for sterilizing, seeding, culturing, storing, shipping and testing tissue, synthetic or native, vascular grafts
CN1481906A (en) * 2002-09-09 2004-03-17 上海组织工程研究与开发中心 Method and apparatus for constructing blood vessel in vitro in the t issue project
CN2647864Y (en) * 2003-08-28 2004-10-13 中国人民解放军第三军医大学 Reactor for in vitro blood vessel culture
CN101333498A (en) * 2008-06-13 2008-12-31 东华大学 Apparatus for culturing tissue engineered blood
CN203999633U (en) * 2014-04-29 2014-12-10 中国人民解放军第四军医大学 A kind of vascular bodies external stress culture apparatus and culture systems
CN114107049A (en) * 2021-11-25 2022-03-01 上海泉众机电科技有限公司 Experimental cavity device for blood vessel culture
CN114949362A (en) * 2022-07-29 2022-08-30 海迈医疗科技(苏州)有限公司 Tissue engineering blood vessel capable of realizing in vivo rapid recellularization and preparation method thereof
CN115076476A (en) * 2022-05-25 2022-09-20 中国一冶集团有限公司 Water leakage prevention device for water supply pipeline joint

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR100496354B1 (en) * 2002-03-27 2005-06-20 서울산업대학교 산학협력단 Hybrid Grafts Including Biodegradable Polymer Supporting Layer And Manufacturing Process Thereof
US7622298B2 (en) * 2006-03-24 2009-11-24 Norits, Inc. Method for creating perfusable microvessel systems
CN103438253B (en) * 2013-08-15 2016-06-01 奇瑞汽车股份有限公司 A kind of self-locking type one-way valve

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1067403A (en) * 1991-06-03 1992-12-30 美国管道和塑料制品公司 The method and apparatus of installing pipe liner
KR20000022284A (en) * 1996-06-27 2000-04-25 나우톤 질 케이. Apparatus and method for sterilizing, seeding, culturing, storing, shipping and testing tissue, synthetic or native, vascular grafts
CN1481906A (en) * 2002-09-09 2004-03-17 上海组织工程研究与开发中心 Method and apparatus for constructing blood vessel in vitro in the t issue project
CN2647864Y (en) * 2003-08-28 2004-10-13 中国人民解放军第三军医大学 Reactor for in vitro blood vessel culture
CN101333498A (en) * 2008-06-13 2008-12-31 东华大学 Apparatus for culturing tissue engineered blood
CN203999633U (en) * 2014-04-29 2014-12-10 中国人民解放军第四军医大学 A kind of vascular bodies external stress culture apparatus and culture systems
CN114107049A (en) * 2021-11-25 2022-03-01 上海泉众机电科技有限公司 Experimental cavity device for blood vessel culture
CN115076476A (en) * 2022-05-25 2022-09-20 中国一冶集团有限公司 Water leakage prevention device for water supply pipeline joint
CN114949362A (en) * 2022-07-29 2022-08-30 海迈医疗科技(苏州)有限公司 Tissue engineering blood vessel capable of realizing in vivo rapid recellularization and preparation method thereof

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