CN103614296B - Double-chamber three-dimensional pouring bioreactor system - Google Patents

Abstract

The invention discloses a double-cavity three-dimensional perfusion bioreactor system, which comprises a bioreactor, a three-dimensional support is arranged in the bioreactor, a perfusion device is connected to the bioreactor, and the bioreactor includes a first The cavity, and the second cavity filled with the inducing culture factor, a dialysis membrane component that can pass through the components with a molecular weight smaller than that of the inducing cytokine in the inducing culture factor is arranged between the first cavity and the second cavity, and the three-dimensional support is located on the in the second chamber. In the present invention, through the double-chamber structure design of the bioreactor, the ordinary nutrient culture solution and the induced culture factor in the two chambers can be changed independently, and the induced culture factor can be retained while the common nutrient culture solution is replaced, greatly reducing the induced cytokines. The dosage can improve its effective utilization and reduce the cost.

Description

A dual-chamber three-dimensional perfusion bioreactor system

technical field

The invention belongs to the field of bioengineering equipment, in particular to a double-chamber three-dimensional perfusion bioreactor system in which cells grow and differentiate in a three-dimensional support.

Background technique

At present, in the field of life science research and medical field, there are many cell culture methods, which can basically be classified into two-dimensional and three-dimensional culture methods according to the dimensions of culture. In two-dimensional culture methods, such as glass or plastic culture bottles and culture dishes, it is necessary to provide a plane for cell attachment and growth, and to provide a better growth environment for cells by modifying the type of plane material and the contact surface, and also through an inverted microscope And ordinary microscopes can easily observe the growth status of cells. However, normal cells grow in a three-dimensional environment in both human and animal bodies, and some cells cannot adhere to any plane for growth. Therefore, providing cells with a three-dimensional environment for culture in vitro is more in line with the normal physiological state of cells and helps to improve Good simulation of the corresponding growth process in vivo.

There are many forms of three-dimensional culture in the current technical field, such as: (1) On the basis of two-dimensional culture plane, add a layer of collagen matrix components above the plane to make cells grow in the three-dimensional environment of the matrix, or Three-dimensional scaffolds are made by artificial materials and cultured in combination with a perfusion system; (2) Power is provided by the swing and rotation of propellers of various shapes to make cells grow in a three-dimensional environment suspended in the culture liquid. Multiple cells are microencapsulated to provide cells with an attachment surface and contact between cells; (3) Create a gravity-free environment through the principle of centrifugal force, so that cells can grow in all directions in three-dimensional space.

In the above-mentioned three-dimensional culture method of cells, the method of culturing through a three-dimensional support combined with a perfusion system, such as "A Rotary Perfusion Bioreactor System" disclosed in the patent document CN 1288235C, includes a bioreactor, the biological The reactor includes a base, on which there is a reaction vessel, which is filled with a culture solution and fixed with a three-dimensional support. The reaction vessel injects the culture solution into the reaction vessel through a perfusion system, and the two ends of the reaction vessel pass through a semi-permeable membrane. Perform gas exchange inside and outside the bioreactor. The rotating perfusion bioreactor system can improve the cell culture environment very well, promote a more uniform distribution of cells on the three-dimensional scaffold, promote the delivery of oxygen and nutrients to the interior of the scaffold, and maintain the activity and function of the cells inside the scaffold. Realize the real-time exchange of gases inside and outside the reactor, ensure that the content of oxygen and carbon dioxide in the culture medium is basically stable, maintain a physiological pH value, and provide a more favorable microenvironment for cell metabolism and function. In order to promote the growth and differentiation of cells, the culture medium in the above-mentioned bioreactor generally includes common nutrient culture medium and induced culture factors, and common nutrient culture medium generally includes basic nutrients such as sugar, amino acids, trace elements, etc. to maintain cell growth and proliferation , also includes buffer to adjust the appropriate physiological pH value, and inducible culture factors play a role as the key components of cell growth and differentiation, but they are extremely expensive because of their difficulty in obtaining and low yield. Since the above-mentioned bioreactor has only one reaction vessel, the growth culture medium and the induction culture factor must be mixed in the same reaction vessel, and the amount of the induction culture factor required for one cultivation is very large; the common nutrient culture medium is relatively cheap, for To meet the needs of cell growth, the culture medium needs to be replaced frequently. Every time the common nutrient culture medium is replaced, the induced culture factors will also be lost. The cost of such culture is relatively high.

Contents of the invention

In order to overcome the deficiencies of the prior art, the present invention provides a dual-chamber three-dimensional perfusion bioreactor system, which greatly improves the utilization rate of induced culture factors through the design of the double-chamber structure and dialysis membrane components , reduce the culture cost, and are more economical and affordable than the three-dimensional perfusion culture in the prior art.

In order to achieve the above object, the technical solution adopted in the present invention is: a double-cavity three-dimensional perfusion bioreactor system, including a bioreactor, a three-dimensional support is provided in the bioreactor, and a perfusion device is connected to the bioreactor, which is characterized in that: The bioreactor includes a first chamber filled with ordinary nutrient culture solution, and a second chamber filled with inducible culture factors, between the first chamber and the second chamber there is an available molecular weight less than the induced culture factor A dialysis membrane module through which cytokine-inducing components pass, and the three-dimensional scaffold is located in the second cavity.

In the above-mentioned structure, the design of the double-chamber structure makes the ordinary nutrient culture solution and the induced culture factor be placed in two chambers, and the nutrients required for cell growth and proliferation can be distributed in the two chambers through the dialysis membrane module between the double chambers. It can be exchanged at any time, while the macromolecular substances such as inducing cytokines in the inducing culture factors are isolated in the second chamber; when the ordinary nutrient culture medium is replaced, the inducing culture factors can be retained, which greatly reduces the amount of inducing cytokines and improves the production efficiency. Its effective utilization rate greatly reduces the cultivation cost.

As a further configuration of the present invention, micropores or microchannels are provided in the three-dimensional stent, and the micropores or microchannels merge to form two inlet and outlet conduits, which are respectively an arterial liquid inlet pipe and a venous liquid outlet pipe, and the arterial liquid inlet pipe The tube and the venous outlet tube are respectively connected with the perfusion device to form the internal liquid circulation mechanism of the three-dimensional stent.

In the above structure, the three-dimensional scaffold can be provided with natural or artificial three-dimensional micro or micro-channels, and finally form an internal liquid circulation mechanism similar to the circulation of human blood vessels through the micro-pores or micro-channels, which can fill the three-dimensional scaffold to form a full three-dimensional structure, so that cells can be better attached to the three-dimensional scaffold, so that the cells in all corners of the scaffold can get a good exchange of gas and metabolic substances. When using organ decellularized scaffolds and some artificial scaffolds with good biocompatibility , in the late stage of cell culture, tissues and organs with appropriate shapes can be formed, and can be directly transplanted to the corresponding blood vessels in the body through the arterial and venous inflow and outflow tubes; Arteries flow in and veins flow out of the blood circulation tube set.

As a further configuration of the present invention, the internal liquid circulation mechanism includes a needle assembly connected to the arterial liquid inlet tube and the venous liquid outlet pipe respectively, and the needle assembly is opposite to the arterial liquid inlet pipe or the venous outlet pipe. The other end of the liquid tube connection is connected to the perfusion device. The needle assembly includes at least two needles, which are respectively a first needle and a second needle. The tail of the first needle is connected to the head of the second needle through a silicone head. end-to-end serial connection.

In the above structure, the needle assembly is connected by two needles, and the silicone head of the two needle holders has a certain degree of elasticity so that the end of the first needle connected to the arterial inlet tube has better swingability, which is convenient for installing the three-dimensional support and connecting with the needle. The installation of components; in addition, the setting of the silicone head can prevent air bubbles from entering the second cavity. When the silicone head is filled with liquid, the connection between the two needles is well sealed, and when the second needle and the perfusion After the connection of the device is disconnected, if there is no silicone head, the gas in the air will enter the second cavity. In severe cases, it will block the vascular system in the three-dimensional stent and affect the cultivation of cells.

As a further configuration of the present invention, the side wall of the second cavity is provided with a through hole for the needle assembly to pass through, the side wall of the second cavity is provided with a supporting cavity protruding outward, the first The head of a needle passes through the through hole and is located in the second cavity, and the tail of the first needle is fixed on the side wall of the second cavity through the supporting cavity.

In the above structure, only the head of the needle assembly is located in the second cavity, while other parts are installed outside the second cavity through the support cavity fixed on the side wall of the second cavity, which increases the size of the second cavity. The operable space in the cavity improves the flexibility of operation, such as the operation of fixing the bracket, connecting the needle assembly and the bracket, and the like.

As a further configuration of the present invention, the two opposite side walls of the second cavity are respectively provided with an outer circulation inlet and an outer circulation outlet, and the outer circulation inlet and the outer circulation outlet are respectively connected with the filling device to form a one-way flow The external liquid circulation mechanism.

In the above structure, the external liquid circulation mechanism refers to the liquid circulation mechanism located outside the three-dimensional support and located in the second cavity. The circulation is a one-way circulation mechanism located outside the three-dimensional support. The external liquid circulation mechanism can Reduce the adhesion of cells to the inner wall or bottom of the second cavity, so that the cells adhered to it can be brought into the three-dimensional scaffold through the external liquid circulation as much as possible, so that the cells can be cultured in the three-dimensional scaffold; internal and external circulation are used together , the effect is better; in addition, it is also possible to control the perfusion device to carry out interval culture medium circulation, so that the cells in the three-dimensional scaffold can be more easily attached to the three-dimensional scaffold without being washed away.

As a further configuration of the present invention, the two side walls of the end connecting the second cavity to the first cavity are provided with slots, the dialysis membrane assembly is embedded in the slot, and the dialysis membrane assembly It includes a first splint, a second splint, and a cellulose semipermeable membrane attached between the first splint and the second splint.

In the above structure, a rectangular communication channel is provided between the first cavity and the second cavity, and the sidewalls on both sides of the communication channel are respectively provided with corresponding slots in the vertical direction, and the dialysis membrane module is inserted through the slots. It is installed in the communication channel to isolate the first cavity and the second cavity, which is easy to install and easy to replace; the above-mentioned first splint and second splint are set with a through-slot structure, and the semi-permeable membrane is sandwiched Between the two splints and in the through groove, this structure has a good sealing effect and a good dialysis isolation effect.

As a further configuration of the present invention, an end of the second cavity facing the dialysis membrane assembly is embedded with a partition that blocks the communication between the first cavity and the second cavity.

In the above structure, the spacer can completely isolate the two cavities, which facilitates the independent liquid exchange of the two cavities.

As a further setting of the present invention, the bioreactor also includes a bubble removal device, the bubble device includes a liquid storage tube, the lower end of the liquid storage tube is connected to the perfusion device, and the upper end of the liquid storage tube is connected to a negative pressure suction tube. device, the side wall of the liquid storage tube is provided with a drainage port communicating with the second cavity.

In the above structure, the negative-pressure suction device in the air-removing device can use a syringe with the needle removed, and the negative-pressure suction device can remove the air bubbles generated in the second chamber to ensure that there are no air bubbles in the liquid entering the second chamber. The vasculature within the three-dimensional stent will not be blocked by air bubbles.

As a further configuration of the present invention, the bottom area of the second cavity is 1/3 of the first cavity, and the height of the second cavity is half of the height of the first cavity.

In the above structure, the vertical height of the second cavity is only half of the first cavity; a protruding window is also provided above the second cavity, and a transparent cover is provided on the window; the present invention reduces the vertical height and The opening design of the second cavity window greatly facilitates the operation related to the three-dimensional scaffold in the second cavity and the observation of the cell growth in the three-dimensional scaffold through a stereo microscope during the cell culture process of the three-dimensional scaffold, because of the small height It can meet the requirement that the observation distance of the stereo microscope must be very close.

As a further configuration of the present invention, the above-mentioned bioreactor is made of transparent material. The transparent design of the two cavities facilitates the requirement of light irradiation at the bottom, and also facilitates the real-time imaging observation of fluorescence.

By adopting the above scheme, the present invention can allow cells to obtain good nutrient and gas exchange in the system through the double-chamber three-dimensional perfusion bioreactor system, and waste can be well discharged. It can better grow, proliferate and differentiate into the target cells; and the double-chamber structure design of the bioreactor can make the ordinary nutrient culture medium and the induced culture factor in the two chambers be changed independently, and the normal nutrient culture medium can be replaced at the same time. Retain the induced culture factors, greatly reduce the amount of induced cytokines, improve their effective utilization, and reduce costs.

The present invention will be further described below in conjunction with the accompanying drawings.

Description of drawings

Accompanying drawing 1 is the structural representation of the specific embodiment of the present invention;

Accompanying drawing 2 is the structural front view of the specific embodiment of the present invention;

Accompanying drawing 3 is the A-A sectional view of accompanying drawing 2 of the specific embodiment of the present invention;

Accompanying drawing 4 is the structural sectional view of the bioreactor of specific embodiment of the present invention;

Accompanying drawing 5 is the right view of the structure of the specific embodiment of the present invention;

Accompanying drawing 6 is a left view of the structure of a specific embodiment of the present invention;

Accompanying drawing 7 is the structural sectional view of the dialysis membrane module of the specific embodiment of the present invention;

Accompanying drawing 8 is the structural representation of the needle assembly of the specific embodiment of the present invention;

Accompanying drawing 9 is a schematic diagram of the internal structure of the communication channel in accompanying drawing 4 of a specific embodiment of the present invention;

Figure 10 is another schematic diagram of the internal structure of the communication channel in Figure 4 of the specific embodiment of the present invention.

detailed description

Specific embodiments of the present invention such as shown in Fig. 1-9 are two-chamber three-dimensional perfusion bioreactor system, comprise bioreactor 1, be provided with three-dimensional support 2 in bioreactor 1, bioreactor 1 is connected with perfusion device, bioreactor The device includes a base 3, a first cavity 11 for filling common nutrient culture fluid, and a second cavity 12 for inducing culture factor filling, the first cavity 11 and the second cavity 12 are located on the base 3, and the base 3 starts The role of the peristaltic pump to the power source of the supporting and fixed circulation perfusion device; between the first chamber 11 and the second chamber 12, there is a dialysis membrane assembly that can pass through the components with a molecular weight smaller than that of the induced cytokine in the induced culture factor 4. The three-dimensional support 2 is located in the second cavity 12 . The design of the double-chamber structure allows ordinary nutrient culture solution and induced culture factors to be placed in two chambers, and through the dialysis membrane module 4 between the double chambers, the nutrients required for cell growth and proliferation can be exchanged at any time in the two chambers. The macromolecular substances such as the inducing cytokines in the inducing culture factors are isolated in the second chamber 12; when the ordinary nutrient culture solution is replaced, the inducing culture factors can be retained, which greatly reduces the amount of inducing cytokines and improves its effectiveness. The utilization rate greatly reduces the cultivation cost.

The above-mentioned three-dimensional support 2 is provided with artificial or natural three-dimensional micropores or micropipes, and the micropores or micropipes converge to form two inlet and outlet pipelines, which are arterial liquid inlet pipe and venous liquid outlet pipe, arterial liquid inlet pipe and venous liquid outlet pipe They are respectively connected with the perfusion device to form the internal liquid circulation mechanism of the three-dimensional support 4 . The above-mentioned three-dimensional scaffold 2 has a wide range of choices, and it can be a decellularized scaffold for various tissues and organs such as liver, lung, heart, and kidney. The decellularized scaffold is to retain the extracellular matrix skeleton by removing the original cells in the organ and tissue Formation of the three-dimensional scaffold 2 . This three-dimensional stent 2 has internal blood vessels and other structures that conform to normal physiology. By connecting to the above-mentioned internal liquid circulation mechanism, it can simulate the three-dimensional environment of cell growth in the body to promote the growth and differentiation of stem cells. You can also choose some thicker synthetic scaffolds, as long as they have the above-mentioned circulation pathways similar to blood circulation, they can be applied, and the same organ 3D printing scaffolds can also be applied.

In the above structure, an internal liquid circulation mechanism similar to human blood vessel circulation is formed in the three-dimensional scaffold 2, which can fill the three-dimensional scaffold 2 to form a plump three-dimensional structure, so that cells can be better attached to the three-dimensional scaffold 2, making the three-dimensional scaffold 2. The cells in all corners of the interior are well exchanged for gas and metabolic substances. When using organ decellularized scaffolds and some artificial scaffolds with good biocompatibility, tissues and organs with appropriate shapes can be formed in the later stage of cell culture, and It can be directly transplanted to the corresponding blood vessels in the body through the arterial and venous liquid inlet and outlet pipes; the above-mentioned arterial liquid inlet pipe and venous liquid outlet pipe refer to the pipeline arrangement of the blood circulation that is similar to the blood circulation in which the liquid in the blood vessels of the human body flows in from the arteries and flows out from the veins.

The above-mentioned perfusion device includes a peristaltic pump 51 and a pump tube 52. The base 3 of the above-mentioned bioreactor 1 is provided with a corresponding peristaltic pump holder and a nozzle holder matched with the pump tube 52, and the peristaltic pump 51 is installed on the base 3 , form a whole, fit compactly, and reduce the occupied space.

As shown in Figures 3 and 8, the above-mentioned internal liquid circulation mechanism includes a needle assembly 6 connected to the arterial liquid inlet pipe and the venous liquid outlet pipe respectively, and the needle assembly 6 is opposite to the other end connected to the arterial liquid inlet pipe or the venous liquid outlet pipe. Connected to the pump tube 52 , the needle assembly 6 includes two needles, respectively a first needle 61 and a second needle 62 , the tail of the first needle 61 is connected in series with the head of the second needle 62 through a silicone head 63 . The needle assembly 6 is connected by two needles, and the silicone head 63 between the two needles has a certain degree of elasticity so that the end of the first needle 61 connected to the arterial inlet tube has better swingability, which is convenient for installing the three-dimensional support 2 and connecting with it. The installation of the needle head assembly 6; in addition, the setting of the silicone head 63 can prevent air bubbles from entering the second cavity 12. When the silicone head 63 is filled with liquid, the sealing of the connection between the two needles is good. After the connection between the second needle 62 and the perfusion device is disconnected, if there is no silicone tip 63, the gas in the air will enter the second cavity 12, and in serious cases, it will block the vascular system in the three-dimensional stent 2, affect cell culture.

The side wall of the second cavity 12 is provided with two through holes for the two needle assemblies 6 to pass through, and the side wall of the second cavity 12 is provided with an outwardly protruding supporting cavity 121, which is supported by the supporting cavity 121. The head of the first needle 61 is located in the second cavity 12 through the through hole, and the tail of the first needle 61 is fixed on the side wall of the second cavity 12 through the supporting cavity 121 . Only the head of the needle assembly 6 is located in the second cavity 12, while other parts are installed outside the second cavity 12 through the supporting cavity 121 fixed on the side wall of the second cavity 12, which increases the second cavity. The operable space in the cavity 12 improves the flexibility of operation, such as the operation of fixing the three-dimensional support 2 and connecting the needle assembly 6 and the three-dimensional support 2 .

The two opposite side walls of the second chamber 12 are respectively provided with interfaces 13 connected to the inlet and outlet of the outer circulation, and the interface 13 of the inlet of the outer circulation and the interface 13 of the outlet of the outer circulation are respectively connected with the perfusion device to form a one-way flow The external liquid circulation mechanism. The external liquid circulation mechanism refers to the liquid circulation mechanism located outside the three-dimensional support 2 and located in the second cavity 12. This circulation is a circulation mechanism that runs through the outside of the three-dimensional support 2 in one direction. The external liquid circulation mechanism can reduce When the cells adhere to the inner wall or bottom surface of the second cavity 12, the cells adhered to it are brought into the three-dimensional support 2 through the external liquid circulation as much as possible, so that the cells are cultivated in the three-dimensional support 2; the internal and external circulation When used in combination, the effect is better; in addition, by controlling the perfusion device to perform interval circulation, the cells in the three-dimensional scaffold 2 can be more easily attached to the three-dimensional scaffold 2 without being washed away.

As shown in Figures 3, 4, and 9, a rectangular communication passage 14 is provided between the second chamber 12 and the first chamber 11, and the side walls on both sides of the communication passage 14 are respectively provided with corresponding openings in the vertical direction. Card slot 141, the dialysis membrane assembly 4 is embedded in the card slot 141, the dialysis membrane assembly 4 includes a first splint 41, a second splint 42, and a cellulose half attached between the first splint 41 and the second splint 42 The permeable membrane 43 generally induces cytokines to be macromolecular proteins, so the semipermeable membrane 43 is generally less than 3.5kd. In specific experiments, it can be set according to the molecular weight of different cytokine-inducing proteins in the inducing culture factors. The dialysis membrane module 4 is embedded in the communication channel 14 through the slot 141, which is convenient to install and easy to replace; Between the two splints and in the through groove 44, this structure has a good sealing effect and a good dialysis isolation effect.

As shown in FIG. 4 , the end of the second cavity 12 facing the dialysis membrane assembly 4 is embedded with a partition 7 that blocks the communication between the first cavity 11 and the second cavity 12 . The spacer 7 can completely isolate the two cavities, which facilitates the independent liquid exchange of the two cavities. The side wall of the communication channel 14 is also provided with a groove 142 for inserting the spacer 7. The groove 142 is located at the end of the above-mentioned locking groove 141 facing the second concave cavity 12, and there is a gap between the spacer 7 and the dialysis membrane assembly 4. The gap, the two are independently installed in the communication channel 14 as shown in Figure 10. The independently installed dialysis membrane assembly 4 and the partition 7 have better sealing performance.

As shown in Figure 1, the above-mentioned bioreactor 1 also includes a bubble removal device 8, and its bubble device 8 includes a liquid storage tube 81, the lower end of the liquid storage tube 81 is connected with the pump tube 52 through a medical three-way pipe, and the liquid storage tube 81 The upper end of the upper end is connected with a negative pressure suction device 82, and the side wall of the liquid storage tube 81 is provided with a drainage port 811 that communicates with the needle assembly 6 connected to the arterial liquid inlet tube on the three-dimensional stent 2. The negative-pressure suction device 82 in the air-removing device 8 can adopt the syringe with the needle removed, and the negative-pressure suction device 82 can remove the bubbles generated in the second chamber 12 to ensure that the liquid entering the second chamber 12 has no bubbles. Make sure that the vasculature within the 3D scaffold 2 is not blocked by air bubbles.

The bottom area of the second cavity 12 is 1/3 of the first cavity 11 , and the height of the second cavity 12 is half of the height of the first cavity 11 . The vertical height of the second chamber 12 is only half of the first chamber 11; the above-mentioned second chamber 12 is also provided with a protruding window 122, and the window 122 is covered with a transparent cover 123; The vertical height of the second cavity 12 and the design of the window 13 of the second cavity 12 greatly facilitate the operation related to the three-dimensional support 2 in the second cavity 12 and the three-dimensional observation through a stereomicroscope during the cell culture process of the three-dimensional support 2 The growth of cells in the scaffold, because the smaller height can meet the requirements of the stereo microscope for the observation distance must be very close.

As shown in Figure 3, the other end of the first chamber 11 opposite to the second chamber 12 is provided with a connecting pipe 111, the connecting pipe 111 is provided with a sealing ring 114, and the connecting pipe 111 is threadedly connected with an interface cover 112, the interface cover 112 is provided with a standard interface 113 connected with the perfusion device. In the present invention, the interfaces or pipes connected with the perfusion device are all provided as standard parts, which is convenient for configuration. The first cavity 11 is designed with an openable connecting tube 111 , so that the liquid in the cavity can be easily injected or removed, and in addition, the possibility of bacterial contamination during the cultivation process is also reduced. This design can not only enter liquid but also gas. Through the combination of bubbling gas and hair, it can well prevent the cells of the culture device from being damaged by the bubbles generated by the bubbling device.

The above-mentioned bioreactor 1 is made of transparent material, and can be made of optically transparent polystyrene and other polyethylene plastic materials according to different production conditions. The transparent design of the two cavities facilitates the requirement of light irradiation at the bottom, and also facilitates the real-time imaging observation of fluorescence.

By adopting the above scheme, the present invention can allow cells to obtain good nutrient and gas exchange in the system through the double-chamber three-dimensional perfusion bioreactor system, and waste can be well discharged. It can better grow, proliferate and differentiate into target cells; and the double-chamber structure design of bioreactor 1 can make the common nutrient culture medium and induced culture factors in the two cavities change independently, while changing the common nutrient culture medium Induced culture factors can be retained, the amount of induced cytokines can be greatly reduced, the effective utilization rate of them can be improved, and the cost can be reduced.

Claims (10)

1. a two-chamber dimensional perfusion bioreactor system, comprise bio-reactor, three-dimensional rack is provided with in bio-reactor, bio-reactor is connected with device for casting, it is characterized in that: described bio-reactor comprises first cavity volume of filling for ordinary nutritional nutrient solution, and for the second cavity volume that the inducing culture factor is filled, be provided with the dialysis membrane assembly that the composition that can be less than inducing cytokine in the inducing culture factor for molecular weight passes through between first cavity volume and the second cavity volume, described three-dimensional rack is arranged in described second cavity volume.

2. two-chamber dimensional perfusion bioreactor system according to claim 1, it is characterized in that: in described three-dimensional rack, be provided with micropore or microchannel, described micropore or microchannel converge formation two inlet and outlet pipings and are respectively artery liquid-inlet pipe and vein drain pipe, and described artery liquid-inlet pipe and vein drain pipe are connected to form the internal liquid circulation mechanism of three-dimensional rack respectively with described device for casting.

3. two-chamber dimensional perfusion bioreactor system according to claim 2, it is characterized in that: described internal liquid circulation mechanism comprise respectively with described artery liquid-inlet pipe and the needle assembly that is connected with described vein drain pipe, the other end that described needle assembly is connected with described artery liquid-inlet pipe or vein drain pipe is relatively connected with described device for casting, described needle assembly comprises at least two syringe needles, be respectively the first syringe needle and the second syringe needle, the afterbody of the first syringe needle is connected in series by the head head and the tail of silica gel head and described second syringe needle.

4. two-chamber dimensional perfusion bioreactor system according to claim 3, it is characterized in that: described second cavity volume sidewall has the through hole passed for described needle assembly, described second cavity volume sidewall is provided with the support chamber of protruding outwardly, the head of described first syringe needle is arranged in described second cavity volume through described through hole, and the afterbody of described first syringe needle is fixedly arranged on described second cavity volume sidewall by described support chamber.

5. the two-chamber dimensional perfusion bioreactor system according to claim 1-4 any one, it is characterized in that: two sidewalls that described second cavity volume is relative are respectively equipped with outer circulation import and outer circulation outlet, described outer circulation import and outer circulation outlet are connected to form the outside liquid cycling mechanism of one-way flow respectively with described device for casting.

6. two-chamber dimensional perfusion bioreactor system according to claim 1, it is characterized in that: two sidewalls of one end that described second cavity volume is connected with described first cavity volume offer draw-in groove, described dialysis membrane assembly is embedded in draw-in groove, the Mierocrystalline cellulose semi-permeable membranes that described dialysis membrane assembly comprises the first clamping plate, the second clamping plate and is attached between the first clamping plate and the second clamping plate.

7. the two-chamber dimensional perfusion bioreactor system according to claim 1 or 6, is characterized in that: be embedded with the partition blocking described first cavity volume and be communicated with the second cavity volume in described second cavity volume towards one end of described dialysis membrane assembly.

8. two-chamber dimensional perfusion bioreactor system according to claim 1, it is characterized in that: described bio-reactor also comprises the bulb apparatus that degass, its bubble assembly described comprises liquid storage pipe, the lower end of liquid storage pipe is connected with described device for casting, the upper end of liquid storage pipe is connected with negative pressure suction device, and the sidewall of liquid storage pipe is provided with the draining hole be communicated with described second cavity volume.

9. two-chamber dimensional perfusion bioreactor system according to claim 1, is characterized in that: the floorage of described second cavity volume is 1/3 of the first cavity volume, and described second cavity volume height is the half of described first cavity volume height.

10. two-chamber dimensional perfusion bioreactor system according to claim 1, is characterized in that: described bio-reactor is that transparent material is made.