Decellularization reactor

Abstract

The decellularization reactor according to the present invention includes a mesh chamber for accommodating tissue of a xenogeneic organ to be decellularized, at least one reaction vessel for storing a decellularization solution, and a first reactor for circulating the decellularization solution between the inside and outside of the reaction vessel and analyzing various physical properties of the circulating decellularization solution in situ.

Description

[Technical Field]

[0001] The present invention relates to a decellularization reactor, and more particularly to a decellularization reactor for producing bioink. [Background technology]

[0002] Recently, as the global trend toward aging has led to an increase in the number of people with chronic diseases, interest has been growing in artificial organ manufacturing technology using 3D bioprinting.

[0003] Bioink is a term used to describe ink materials used to print artificial organs using 3D bioprinters, and is manufactured based on the extracellular matrix obtained after decellularizing xenogeneic organs.

[0004] The decellularization process of such xenogeneic organs is the most crucial process in the production of bioink, and refers to a process of removing all impurities and cells from the xenogeneic organs to prevent antigen-antibody reactions.

[0005] The decellularization process for applying 3D bioprinting technology involves immersing the xenogeneic organ in a solution. Until now, there has been no specialized equipment for decellularization, so simple laboratory glass materials such as beakers have been used as the equipment.

[0006] Furthermore, since there is no specific indicator for discharging the existing solution during the process of immersion in various decellularization solutions and transferring to the process of injecting the next solution, there is a problem of deviations in the physical properties and quality between final products. Summary of the Invention [Problem to be solved by the invention]

[0007] The object of the present invention is to provide a decellularization reaction device that can derive indicators such as the processing time of the decellularization solution, reinjection or transfer to the next solution, and completion of the decellularization process by measuring the electrical conductivity, turbidity, absorbance (UV-Vis), etc. of the solution during which decellularization is in progress. [Means for solving the problem]

[0008] A decellularization reaction device according to one embodiment of the present invention may include a mesh chamber for accommodating tissue of a heterologous organ to be decellularized therein, and at least one reaction vessel for storing a decellularization solution for carrying out a decellularization reaction with the tissue of the heterologous organ in the mesh chamber that is inserted therein.

[0009] The decellularization reaction device may also include a first reactor for discharging the decellularization solution undergoing a decellularization reaction in the reaction vessel from the reaction vessel and then circulating it back into the reaction vessel, and analyzing various physical properties of the circulating decellularization solution in situ.

[0010] The decellularization reaction device may further include a second reactor for extracting the decellularization solution undergoing a decellularization reaction inside the reaction vessel to the outside of the reaction vessel and analyzing various physical properties of the decellularization solution ex-situ.

[0011] Hereinafter, the first reactor may refer to a reactor to which an in-situ analysis device is applied, in which an analysis device such as a turbidity meter, an electrical conductivity meter, or an absorbance meter is installed in the reactor piping.

[0012] In addition, the second reactor may refer to a reactor in which an analytical device such as a turbidity meter, an electrical conductivity meter, or an absorbance meter is not installed in the reactor piping, and a method of sampling a solution and measuring the corresponding physical properties ex situ is applied.

[0013] The first reactor can include a solution circulation unit that is connected between the reaction vessel and the cover of the reaction vessel so that they communicate with each other, and that circulates the decellularization solution between the reaction vessel and the cover.

[0014] The first reactor is installed in the solution circulation section and can include a measurement section for measuring in situ various physical properties of the decellularization solution circulating in the solution circulation section during the decellularization reaction.

[0015] The mesh chamber may be coupled with a drive unit for rotating the mesh chamber so as to stir the contents within the reaction vessel.

[0016] The cover may be coupled with a lifting unit for vertically lifting and lowering the cover relative to the upper end of the reaction vessel.

[0017] The mesh chamber may have a cylindrical shape with an open interior.

[0018] The mesh chamber may be made of or coated with a chemical-resistant material depending on the physical properties of the decellularization solution.

[0019] The mesh chamber may have a mesh arranged in a lattice pattern on its outer periphery.

[0020] The size of the mesh may be set to be smaller than the tissue of the xenogeneic organ to be decellularized.

[0021] A lid for covering the mesh chamber may be attached to the upper end of the mesh chamber.

[0022] The reaction vessel may be made of a transparent material or have a transparent window so that the inside can be observed.

[0023] The reaction vessel may be configured in a double jacket configuration including an inner tube and an outer tube to control the temperature of the decellularization process.

[0024] The reaction vessel can have a solution outlet for discharging the decellularization solution.

[0025] A solution injection tube for injecting a decellularization solution into the reaction vessel may be connected to the cover.

[0026] The driving unit may include a rotation shaft coupled to the lid of the mesh chamber, and a driving motor installed on the cover of the reaction vessel and coupled to the rotation shaft to rotate the rotation shaft.

[0027] The circulation unit can include a circulation tube connected between the cover and the solution outlet for circulating the decellularization solution in the reaction vessel, and a transfer pump installed in the circulation tube for quantitatively transferring the decellularization solution circulating in the circulation tube.

[0028] The circulation pipe may be made of an expandable and contractible flexible pipe.

[0029] The measuring unit can include an absorbance analyzer, an electrical conductivity meter, and a turbidity meter for measuring the absorbance, electrical conductivity, and turbidity, respectively, of the decellularization solution circulating in the circulation tube.

[0030] The measuring unit can include a thermometer for measuring the temperature of the decellularization solution circulating in the circulation tube.

[0031] The lifting unit may include a binding plate coupled to the cover so as to be able to lift and lower, and a lifting cylinder having a lifting rod coupled to the binding plate for lifting and lowering the binding plate.

[0032] An air supply pipe for supplying air into the lifting cylinder is connected to the lifting cylinder, and a regulator for adjusting the pressure of the air supplied to the air supply pipe may be installed on the air supply pipe.

[0033] The second reactor may be installed in the circulation pipe and include a solution sampling pipe for extracting and sampling the decellularization solution circulating through the circulation pipe.

[0034] The solution sampling pipe may be provided with a solution sampling pipe control valve for controlling opening and closing of the solution sampling pipe.

[0035] A decellularization reaction device according to another embodiment of the present invention may include a mesh chamber for accommodating tissue of a heterologous organ to be decellularized therein, and at least one reaction vessel for storing a decellularization solution for carrying out a decellularization reaction with the tissue of the heterologous organ in the mesh chamber that is inserted therein.

[0036] The decellularization reaction device may also include a second reactor for extracting the decellularization solution undergoing a decellularization reaction inside the reaction vessel to the outside of the reaction vessel and analyzing various physical properties of the decellularization solution ex-situ.

[0037] A solution circulation section may be included that is connected between the reaction vessel and the cover of the reaction vessel so as to communicate with each other and that circulates the decellularization solution between the reaction vessel and the cover.

[0038] The mesh chamber may be coupled with a drive unit for rotating the mesh chamber so as to stir the contents within the reaction vessel.

[0039] The cover may be coupled with a lifting unit for vertically lifting and lowering the cover relative to the upper end of the reaction vessel.

[0040] The mesh chamber may have a mesh arranged in a lattice pattern on its outer periphery.

[0041] The size of the mesh may be set to be smaller than the tissue of the xenogeneic organ to be decellularized.

[0042] The circulation unit can include a circulation tube connected between the cover and the reaction vessel for circulating the decellularization solution in the reaction vessel, and a transfer pump installed in the circulation tube for quantitatively transferring the decellularization solution circulating in the circulation tube.

[0043] The circulation pipe may be made of an expandable and contractible flexible pipe.

[0044] The circulation tube may be provided with a solution sampling tube for extracting and sampling the decellularization solution circulating through the circulation tube.

[0045] The reaction vessels are arranged at set intervals on the base and supported by a first support frame arranged vertically to the base, and an alignment groove for aligning the positions of the reaction vessels and the cover may be arranged on the upper surface of the base.

[0046] A second elastic member may be coupled to the upper end of the support frame to absorb impact from the reaction vessel. [Effects of the Invention]

[0047] According to an embodiment of the present invention, by measuring the electrical conductivity, turbidity, absorbance (UV-Vis), etc. of a solution in which decellularization is in progress, it is possible to derive indicators such as the processing time of the decellularization solution, reinjection or transfer to the next solution, and the completion of the decellularization process.

[0048] In addition, by using a reaction vessel, mesh chamber, and solution ring structure, various physical properties of the solution (electrical conductivity, turbidity, absorbance, etc.) can be analyzed during the decellularization process, thereby making it possible to derive the conditions for discharging and re-injecting the decellularization solution and for completing the decellularization process, thereby obtaining high-quality decellularized materials while also having an outstanding effect in ensuring homogeneity between materials. [Brief explanation of the drawings]

[0049] [Figure 1] FIG. 1 is a schematic diagram illustrating the configuration of a first reactor of a decellularization reaction apparatus according to a first embodiment of the present invention. [Figure 2] FIG. 1 is a schematic side view of a first reactor of a decellularization reactor according to a first embodiment of the present invention. [Figure 3] FIG. 2 is a schematic diagram illustrating the configuration of a mesh chamber of a first reactor of a decellularization reactor according to a first embodiment of the present invention. [Figure 4] FIG. 10 is a schematic side view of a second reactor of a decellularization reactor according to a second embodiment of the present invention. [Figure 5] 1 is a graph showing the analysis results of the electrical conductivity of a decellularization solution measured during a porcine placenta decellularization process using the first reactor of the decellularization reactor according to the first embodiment of the present invention. [Figure 6] 1 is a graph showing the analysis results of the turbidity of a decellularization solution measured during a porcine placenta decellularization process using the first reactor of the decellularization reactor according to the first embodiment of the present invention. [Figure 7] 1 is a graph showing the results of analyzing the absorbance (UV-Vis) of a decellularization solution measured during a porcine placenta decellularization process using the first reactor of the decellularization reactor according to the first embodiment of the present invention. [Figure 8] 3A and 3B are photographs showing the state of a porcine placenta before and after a decellularization process using the first reactor of the decellularization reaction apparatus according to the first embodiment of the present invention. DETAILED DESCRIPTION OF THE INVENTION

[0050] Hereinafter, with reference to the accompanying drawings, embodiments of the present invention will be described so that those skilled in the art can easily implement the present invention. As will be easily understood by those skilled in the art, the embodiments described below can be modified in various ways without departing from the concept and scope of the present invention. Whenever possible, the same or similar parts in the drawings will be designated by the same reference numerals.

[0051] The terminology used below is merely for the purpose of referring to particular embodiments and is not intended to limit the present invention. As used herein, the singular form includes the plural form unless the context clearly dictates otherwise. As used in the specification, the meaning of "comprising" embodies certain properties, regions, integers, steps, operations, elements, components and / or groups, and does not exclude the presence or addition of other specific properties, regions, integers, steps, operations, elements, components and / or groups.

[0052] All terms, including technical and scientific terms, used below have the same meaning as commonly understood by a person of ordinary skill in the art to which the present invention belongs. Terms defined in dictionaries are additionally interpreted to have a meaning consistent with the relevant technical literature and the presently disclosed content, and are not interpreted in an ideal or very formal sense unless otherwise defined.

[0053] FIG. 1 is a schematic diagram of the first reactor of the decellularization reaction apparatus according to the first embodiment of the present invention, FIG. 2 is a schematic side view of the first reactor of the decellularization reaction apparatus according to the first embodiment of the present invention, and FIG. 3 is a schematic diagram of the mesh chamber of the first reactor of the decellularization reaction apparatus according to the first embodiment of the present invention.

[0054] 1 to 3, a decellularization reaction apparatus according to a first embodiment of the present invention may include a mesh chamber 100, a reaction vessel 200, and a first reactor.

[0055] The mesh chamber 100 has a set size and shape and can accommodate tissue of a xenogeneic organ to be decellularized inside.

[0056] In addition, the reaction vessel 200 can have the mesh chamber 100 inserted therein and can store a decellularization solution for carrying out a decellularization reaction with the tissue of a heterologous organ in the mesh chamber 100, and can be equipped with at least one or more.

[0057] The first reactor discharges the decellularization solution undergoing a decellularization reaction with the tissue of the heterologous organ to be decellularized within the reaction vessel 200 from the reaction vessel 200 and then circulates it back into the reaction vessel 200, allowing various physical properties of the circulating first decellularization solution to be analyzed in situ.

[0058] The first reactor may also include a driving unit 300 , a solution circulation unit 400 , and a measurement unit 500 .

[0059] The driving unit 300 is coupled to the mesh chamber 100 and can rotate the mesh chamber 100 so as to stir the contents in the reaction vessel 200 .

[0060] In addition, the solution circulation unit 400 is connected between the reaction vessel 200 and the cover 210 that covers the reaction vessel 200 so that they communicate with each other, and can circulate the decellularization solution through the reaction vessel 200 and the cover 210.

[0061] The measurement unit 500 is installed in the solution circulation unit 400 and can measure various physical properties of the decellularization solution circulating in the solution circulation unit 400 during the decellularization reaction in order to enable in-situ analysis.

[0062] A cover 210 for covering the reaction vessel 200 may be placed on top of the reaction vessel 200 .

[0063] Also, a lifting unit 600 may be included which is coupled to the cover 210 and which lifts and lowers the cover 300 in the vertical direction (Y direction in FIG. 1) so as to couple or detach the cover 210 to or from the reaction vessel 200.

[0064] The mesh chamber 100 may be configured in a hollow form with an open interior so that tissue of a xenogeneic organ to be decellularized can be accommodated therein.

[0065] The mesh chamber 100 may have a cylindrical shape so that it can be easily rotated by the driving unit 300 .

[0066] In addition, if the mesh chamber 100 is rectangular, the tissue of the foreign organ may get caught or remain at the corners when the mesh chamber 100 is rotated, which may prevent the tissue of the foreign organ from being mixed evenly. Therefore, the mesh chamber 100 is formed into a cylindrical shape, etc., in order to mix the tissue of the foreign organ evenly and efficiently.

[0067] A mesh 101 is arranged in a lattice pattern on the outer periphery of the mesh chamber 100, and the mesh 101 may have a set size, for example, about 1 mm, for decellularization of tissue of a heterologous organ contained within the mesh chamber 100.

[0068] The mesh chamber 100 may be configured to be larger than the mesh 100A of the second embodiment, and may be configured as a large type capable of accommodating, for example, 500 g (maximum 1 kg) of a xenogeneic organ to be decellularized.

[0069] Furthermore, the size of the mesh 101 may be set to be slightly smaller than the tissue of the xenogeneic organ contained within the mesh chamber 100 so that the tissue of the xenogeneic organ does not escape to the outside of the mesh 101 during the decellularization process.

[0070] A lid 110 may be included that is detachably coupled to the upper end of the mesh chamber 100 and that covers the mesh chamber 100 .

[0071] The cover 110 may have a buckle 111 so that it can be easily attached to the mesh chamber 100 .

[0072] A driving unit 300 may be coupled to the upper end of the cover 110 .

[0073] The mesh chamber 100 may be made of or coated with a material with good chemical resistance, such as Teflon, depending on the physical properties of the decellularization solution.

[0074] In addition, the reaction vessel 200 may be made of a transparent material or have a transparent window (not shown) so that the progress of the decellularization process taking place within the reaction vessel 200 can be visually observed from outside the reaction vessel 200.

[0075] The reaction vessels 200 may be arranged at set intervals on the base 10 and supported by a first support frame 20 arranged in a direction perpendicular to the base 10 (Y direction in FIG. 1).

[0076] The reaction vessel 200 may be made of a material that is resistant to corrosion by the decellularization solution to be used.

[0077] Furthermore, the reaction vessel 200 may have a double jacket configuration including an inner tube 201 and an outer tube 203 to control the temperature of the decellularization process.

[0078] The outer pipe 203 may be provided with a refrigerant inlet 205 for injecting a refrigerant into the space between the outer pipe 203 and the inner pipe 201, and a refrigerant outlet 206 for discharging the refrigerant to the outside.

[0079] The reaction vessel 200 can have a solution outlet 207 at its lower end for discharging the decellularization solution.

[0080] The cover 210 may be made of a cushioning material such as rubber and may have an O-ring shape so that it can seal the reaction vessel 200 while cushioning impact when it descends by the lifting unit 600 and comes into contact with the upper end of the reaction vessel 200.

[0081] The cover 210 may be provided with at least one injection port 211 to which a solution injection tube 220 for injecting various decellularization solutions into the reaction vessel 200 is connected.

[0082] The various decellularization solutions may consist of, for example, ultrapure water, organic solvents, etc., and each decellularization solution may be injected through a respective injection port 211, and a pump (not shown) may be installed in a pipe (not shown) 0 connected to each injection port 211 to supply a fixed amount of each decellularization solution.

[0083] In addition, the cover 210 is connected to the circulation unit 400 so that the decellularization solution circulating in the circulation unit 400 can be injected into the reaction vessel 200 .

[0084] The cover 210 may be provided with a water level measuring device 213 such as an ultrasonic type for measuring the water level of the decellularization solution in the reaction vessel 200 .

[0085] The driving unit 300 may include a rotating shaft 310 connected to the lid 110 of the mesh chamber 100, and a driving motor 320 installed on the cover 210 of the reaction vessel 200 and connected to the rotating shaft 310 to rotate the rotating shaft 310.

[0086] The driving motor 320 may be a magnetic type motor or the like so that the rotating shaft 310 can be easily rotated.

[0087] The rotation shaft 310 may be disposed at the center of the upper end of the lid 110 in a direction perpendicular to the upper end of the lid 110 (Y direction in FIG. 2), and may be coupled to the lid 110 by fastening a bolt 301 or the like.

[0088] The rotating shaft 310 can be rotated, for example, clockwise or counterclockwise, by the drive motor 320 to rotate the mesh chamber 100 and agitate the tissue of the xenogeneic organ to be decellularized contained within the mesh chamber 100, thereby facilitating decellularization using the decellularization solution injected into the reaction vessel 200.

[0089] When stirring by rotating the mesh chamber 100 at a set rotation speed (e.g., 0 to 300 rpm), stirring can be alternately performed from the forward direction (clockwise) to the reverse direction (counterclockwise) to prevent overloading of the drive motor 320 and to prevent clumping between the different organ tissues, and a stop time can be set in between.

[0090] The circulation unit 400 may include a circulation pipe 410 and a transfer pump 420 .

[0091] The circulation tube 410 is connected between the cover 210 and the solution outlet 207 and is capable of circulating the decellularization solution within the reaction vessel 200 .

[0092] In addition, the transfer pump 420 is installed in the circulation pipe 410 and can quantitatively transfer the decellularization solution circulating in the circulation pipe 410.

[0093] The circulation pipe 410 may be made of a flexible pipe so that it can be easily expanded and contracted and deformed when the cover 210 is raised and lowered by the lifting unit 600 .

[0094] In addition, a waste liquid discharge pipe 430 for discharging waste liquid of the decellularization solution may be connected to the circulation pipe 410, and a waste liquid discharge pipe control valve 431 for controlling the discharge of the decellularization solution may be installed in the waste liquid discharge pipe 430.

[0095] The measuring unit 500 can include an absorbance analyzer 510 for analyzing the absorbance (UV-Vis) of the decellularization solution circulating in the circulation tube 410.

[0096] Furthermore, the measuring unit 500 can include an electrical conductivity meter 520 for measuring the electrical conductivity of the decellularization solution circulating in the circulation tube 410 .

[0097] The measuring unit 500 can include a turbidity measuring device 530 for measuring the turbidity of the decellularization solution circulating in the circulation tube 410.

[0098] The measuring unit 500 can include a thermometer 540 for measuring the temperature of the decellularization solution circulating in the circulation tube 410.

[0099] The lifting unit 600 may include a connecting plate 610 that is connected to the cover 210 so that it can be raised and lowered integrally, and a lifting cylinder 620 that is connected to the lower end of the connecting plate 610 and has a lifting rod 621 for raising and lowering the connecting plate 620.

[0100] The lifting cylinder 620 may be installed and supported on the upper end surface of the base 10 in a direction perpendicular to the base 10 (Y direction in FIG. 2).

[0101] An air supply pipe 623 for supplying air into the lifting cylinder 620 is connected to the lifting cylinder 620, and a regulator 625 for adjusting the pressure of the air supplied to the air supply pipe 623 may be installed in the air supply pipe 623.

[0102] In addition, a second support frame 30 may be installed on the base 10, arranged parallel to the first support frame 20 and supported by the base.

[0103] A guide rod 630 may be coupled to the second support frame 30 via a connecting frame 31 to guide the lifting rod 621 in its vertical movement.

[0104] Hereinafter, the operation of the decellularization reaction device according to the first embodiment of the present invention will be described with reference to FIGS.

[0105] First, tissue of a xenogeneic organ to be decellularized and a decellularization solution for the xenogeneic organ tissue and decellularization reaction are prepared.

[0106] In order to inject the tissue of the xenogeneic organ to be decellularized and the decellularization solution into the mesh chamber 100 and the reaction vessel 200, respectively, the lid 110 is opened from the mesh chamber 100 and the cover 210 is opened from the reaction vessel 200, as shown in Figure 1.

[0107] In this state, tissue of the xenogeneic organ to be decellularized is placed inside the mesh chamber 100, the upper end of the mesh chamber is covered with the lid 110, and the lid 110 is then attached to the mesh chamber 100 using the buckle 111.

[0108] When the lifting rod 621 of the lifting cylinder 620 of the lifting unit 600 is contracted, the lifting rod 621 descends a set length, causing the cover 210 to cover the top of the reaction vessel 200 and seal the reaction vessel 200 .

[0109] In this manner, the mesh chamber 100 is inserted into the reaction vessel 200 by the cover 210 sealing the reaction vessel 200 .

[0110] At this time, the decellularization solution is injected into the reaction vessel 200 through an injection port 211 that is installed in the cover 210 and connected to a solution injection tube 220 .

[0111] Then, when the injection of the decellularization solution into the reaction vessel 200 is completed, the drive motor 320 of the drive unit 300 is driven to rotate the rotation shaft 310 clockwise or counterclockwise at a set speed, thereby rotating the mesh chamber 100 and stirring the solution.

[0112] In this way, by rotating and stirring the mesh chamber 100, the tissue of the heterologous organ in the mesh chamber 100 is stirred and undergoes a decellularization reaction with the decellularization solution in the reaction vessel 200.

[0113] At this time, the rotation of the mesh chamber 100 stirs the tissue of the heterologous organ, and the decellularization solution moves vertically from the cover 210 to the inside of the reaction vessel 200 through the circulation tube 410, allowing the tissue of the heterologous organ and the decellularization solution to mix well, thereby making it possible to obtain a decellularization solution of a homogeneous concentration in real time.

[0114] In this state, the transfer pump 420 is operated to circulate the decellularization solution undergoing a decellularization reaction in the reaction vessel 200 through the circulation pipe 410 at a set speed.

[0115] The physical properties of the decellularization solution circulating through the circulation pipe 410 from the solution outlet 207 of the reaction vessel 410 to the cover 210 can be measured in real time by a measuring unit 500 installed in the circulation pipe 410 .

[0116] That is, the absorbance (UV-Vis) of the decellularization solution is measured and analyzed using an absorbance analyzer 510, the electrical conductivity of the decellularization solution is measured using an electrical conductivity meter 520, the turbidity of the decellularization solution is measured using a turbidity meter 530, and the temperature of the decellularization solution is measured using a thermometer 540.

[0117] Furthermore, by measuring a decellularization solution of a uniform concentration, it becomes easier to analyze various physical properties of the decellularization solution, such as absorbance (UV-Vis), electrical conductivity, and turbidity, making it possible to accurately grasp the progress of the decellularization process in real time. [Example]

[0118] Hereinafter, a porcine placenta decellularization experiment will be described as an example of a xenogeneic organ to be decellularized.

[0119] First, 500 g of porcine placenta that has been pretreated, such as by removing foreign matter, is placed into the mesh chamber 100 in the decellularization reaction vessel 200, and the lid 110 of the mesh chamber 100 is closed. In the case of porcine placenta, decellularization can be performed at room temperature of around 20°C, so no specific temperature is set.

[0120] Then, gas (air) is pressurized into the lifting cylinder 620 to cover and seal the reaction vessel 200 with the cover 210.

[0121] After injecting the decellularization solution into the reaction vessel 200, the speed of the rotating shaft 310 driven by the drive motor 320 is set to 80 rpm, and the mesh chamber 100 is rotated and stirred, for example, with settings of 10 seconds clockwise, 8 seconds pause, and 10 seconds counterclockwise.

[0122] At the same time, the decellularization solution inside the reaction vessel 200 is circulated through the circulation pipe 410 using the metering pump 420. At this time, the circulation speed of the decellularization solution is set to 1,000 ml / min.

[0123] The decellularization solution in the reaction vessel 200 can be circulated through the transfer pump 420 and stirred by rotating the mesh chamber 100 to achieve a uniform concentration.

[0124] Then, in the case of the reaction vessel 100 according to the first embodiment, various physical property data of the decellularization solution can be analyzed in real time during the decellularization process using an absorbance (UV-Vis) analyzer 510, an electrical conductivity meter 520, and a turbidity meter 530 installed in the circulation pipe 410 (see FIG. 1).

[0125] In addition, in the case of the reaction vessel 100A according to the second embodiment, various physical properties of the decellularization solution can be analyzed by sampling the decellularization solution using the solution sampling pipe 550 (see FIG. 4) installed in the circulation pipe 410.

[0126] FIG. 5 is a graph showing the results of analyzing the electrical conductivity of the decellularization solution measured during the porcine placenta decellularization process, and FIG. 6 is a graph showing the results of analyzing the turbidity of the decellularization solution measured during the porcine placenta decellularization process.

[0127] Figures 5 and 6 show the results of analyzing the decellularized solution after three ultrapure water processes, where [Circle 1] indicates the result of the first ultrapure water process, [Circle 2] indicates the result of the second ultrapure water process, and [Circle 3] indicates the result of the third ultrapure water process.

[0128] FIG. 7 is a graph showing the results of analyzing the absorbance (UV-Vis) of the decellularization solution measured during the porcine placenta decellularization process.

[0129] In Figure 7, [Circle 1], [Circle 2], [Circle 3], [Circle 4], [Circle 5], and [Circle 6] represent the absorbance data over time during the first ultrapure water process.

[0130] Here, [circle 1] is the measurement data after 10 minutes, [circle 2] is the measurement data after 20 minutes, [circle 3] is the measurement data after 30 minutes, [circle 4] is the measurement data after 40 minutes, [circle 5] is the measurement data after 50 minutes, and [circle 6] is the measurement data after 60 minutes.

[0131] During the decellularization process, the physical property values ​​within the reactor tend to initially increase over time and then gradually decrease. When the decellularization solution is discharged and then reinjected, a significant change in the physical properties can be seen, as shown by [Circle 1]-[Circle 2]-[Circle 3] in Figures 5 and 6.

[0132] In this way, while monitoring the various physical properties of the decellularization solutions, it is possible to determine whether reinjection of each decellularization solution is necessary or the conditions for completing the process, and this procedure ensures stable decellularization as shown in Figure 8. Therefore, the final decellularized placental tissue material is of high quality and homogeneous.

[0133] (Test results for performance, etc.) When evaluating the quality of bioink, the most basic analysis is of DNA and collagen content. Table 1 shows the results of comparing the DNA content of pig placentas before and after using a decellularization reactor. Each pig placenta is supplied from a different supplier, so there is a large variation in the DNA content of each pig placenta. [Table 1]

[0134] As can be seen from the above results, the DNA content, which was 87-244 ng / mg before using the decellularization reactor, was reduced to an extremely low level of 0.37-0.48 ng / mg after using the decellularization reactor.

[0135] What is significant is that the DNA content was not simply low, but that high quality and uniform results were obtained despite the use of placentas from pigs raised in different locations. [Table 2]

[0136] Table 2 compares the collagen content of porcine placenta before and after using the decellularization reactor. As can be seen from the results, collagen was well preserved at a level of 22-23 μg / mg even after using the decellularization reactor.

[0137] Therefore, after using the decellularization reactor, the DNA content of the tissue was reduced to an extremely low level, and the collagen in the tissue was very well preserved, indicating that high-quality bioink material was produced.

[0138] FIG. 4 is a schematic side view of the second reactor of the decellularization reactor according to the second embodiment of the present invention.

[0139] The second reactor of the decellularization reactor according to the second embodiment of the present invention is identical to the first reactor of the decellularization reactor according to the first embodiment of the present invention, except for the details specifically described below, and therefore a detailed description thereof will be omitted.

[0140] Referring to FIG. 4, the decellularization reaction apparatus according to the second embodiment of the present invention may include a second reactor for extracting the decellularization solution undergoing a decellularization reaction with the tissue of a heterologous organ to be decellularized within the reaction vessel 200 to the outside of the reaction vessel 200 and analyzing various physical properties of the decellularization solution ex-situ.

[0141] The second reactor is installed in the circulation pipe 410 of the circulation unit 400 and may include a solution sampling pipe 550 for extracting and sampling the decellularization solution circulating through the circulation pipe 410.

[0142] The decellularization solution sampled by this solution sampling tube 550 is placed in a small container (not shown) such as a vial, and various physical properties (absorbance, electrical conductivity, turbidity, etc.) of the decellularization solution contained in the small container can then be measured and analyzed ex-situ.

[0143] A solution sampling pipe control valve 551 for controlling opening and closing of the solution sampling pipe 550 may be installed in the solution sampling pipe 550 .

[0144] The mesh chamber 100A may be configured to be smaller than the mesh chamber 100 of the first embodiment, that is, it may be configured as a small type that can accommodate a maximum amount of xenogeneic organ to be decellularized of less than 25 g, and therefore the size of the mesh 101A of the mesh chamber 100A may also be small, that is, set to be smaller than 1 mm.

[0145] The reaction vessels 200 may be arranged at set intervals on the base 10 and supported by a first support frame 20 arranged in a direction perpendicular to the base 10 (Y direction in FIG. 1).

[0146] The reaction vessel 200 and the first support frame 20 are configured to be movable and detachable for ease of cleaning, and an alignment groove 11 may be arranged on the upper surface of the base 10 for aligning the positions of the reaction vessel 200 and the cover 210.

[0147] An elastic member 21 may be attached to the upper end of the first support frame 20 to absorb the shock of the reaction vessel 200 generated when the cover 210 descends to close the reaction vessel 200 .

[0148] Although the present disclosure has been described above through preferred embodiments, it should be readily apparent to those skilled in the art to which the present invention pertains that the present invention is not limited thereto and that various modifications and variations are possible without departing from the scope of the claims set forth below. [Explanation of symbols]

[0149] 100: Mesh chamber 200: Reaction vessel

Claims

1. a mesh chamber for accommodating tissue of the xenogeneic organ to be decellularized therein; At least one reaction vessel for storing a decellularization solution for carrying out a decellularization reaction with the tissue of the xenogeneic organ in the mesh chamber to be inserted therein; and A first reactor for discharging a decellularization solution undergoing a decellularization reaction in the reaction vessel from the reaction vessel and then circulating it back into the reaction vessel, and analyzing various physical properties of the circulating decellularization solution in situ. A decellularization reactor comprising:

2. The decellularization reaction apparatus of claim 1, further comprising a second reactor for extracting the decellularization solution undergoing a decellularization reaction inside the reaction vessel to the outside of the reaction vessel and analyzing various physical properties of the decellularization solution ex-situ.

3. The decellularization reaction apparatus according to claim 2, wherein the first reactor includes a solution circulation unit connected between the reaction vessel and the cover of the reaction vessel so as to be in communication with each other, for circulating a decellularization solution between the reaction vessel and the cover.

4. The decellularization reaction apparatus according to claim 3, wherein the first reactor is installed in the solution circulation unit and includes a measurement unit for measuring in situ various physical properties of the decellularization solution circulating in the solution circulation unit during the decellularization reaction.

5. The decellularization reaction apparatus according to claim 4 , wherein a drive unit is coupled to the mesh chamber for rotating the mesh chamber so as to stir the contents in the reaction vessel.

6. The decellularization reaction apparatus according to claim 5 , wherein the cover is coupled to a lifting unit for vertically lifting and lowering the cover relative to an upper end of the reaction vessel.

7. The decellularization reactor according to claim 6 , wherein the mesh chamber has a cylindrical shape with an open interior.

8. The decellularization reaction device according to claim 7 , wherein the mesh chamber is made of or coated with a chemical-resistant material depending on the physical properties of the decellularization solution.

9. The decellularization reactor according to claim 7 , wherein a mesh is arranged in a lattice pattern on the outer periphery of the mesh chamber.

10. The decellularization reaction apparatus according to claim 9 , wherein the size of the mesh is set to be smaller than the tissue of the xenogeneic organ to be decellularized.

11. The decellularization reactor according to claim 10 , wherein a lid for covering the mesh chamber is attached to an upper end of the mesh chamber.

12. The decellularization reaction apparatus according to any one of claims 1 to 11, wherein the reaction vessel is made of a transparent material or has a transparent window so that the inside can be observed.

13. The decellularization reactor according to claim 12, wherein the reaction vessel is configured in a double jacket form including an inner tube and an outer tube for controlling the temperature of the decellularization process.

14. The decellularization reactor according to claim 13 , wherein the reaction vessel has a solution outlet for discharging the decellularization solution.

15. The decellularization reaction apparatus according to claim 14 , wherein a solution injection tube for injecting a decellularization solution into the reaction vessel is connected to the cover.

16. The drive unit is a rotating shaft coupled to the lid of the mesh chamber; The decellularization reaction apparatus according to claim 11, further comprising a drive motor that is installed on a cover of the reaction vessel and is coupled to the rotation shaft to rotate the rotation shaft.

17. The circulation section includes: a circulation tube connected between the cover and the solution outlet for circulating the decellularization solution in the reaction vessel; and The decellularization reaction apparatus according to claim 14 , further comprising a transfer pump installed in the circulation pipe for quantitatively transferring the decellularization solution circulating in the circulation pipe.

18. The decellularization reaction apparatus according to claim 17 , wherein the circulation tube is made of an expandable and contractible flexible tube.

19. The decellularization reaction apparatus according to claim 18, wherein the measuring unit includes an absorbance analyzer, an electrical conductivity meter, and a turbidity meter for measuring the absorbance, electrical conductivity, and turbidity, respectively, of the decellularization solution circulating in the circulation tube.

20. The decellularization reaction apparatus according to claim 19 , wherein the measurement unit includes a thermometer for measuring the temperature of the decellularization solution circulating in the circulation pipe.

21. The lifting unit is a coupling plate coupled to the cover so as to be able to rise and fall; and The decellularization reactor according to claim 6 , comprising a lifting cylinder having a lifting rod coupled to the binding plate for lifting and lowering the binding plate.

22. an air supply pipe for supplying air into the lifting cylinder is connected to the lifting cylinder; The decellularization reaction apparatus according to claim 21 , wherein the air supply pipe is provided with a regulator for adjusting the pressure of the air supplied to the air supply pipe.

23. The second reactor comprises: The decellularization reaction device according to claim 2 , further comprising a solution sampling tube installed in the circulation tube for extracting and sampling the decellularization solution circulating through the circulation tube.

24. The decellularization reaction apparatus according to claim 23, wherein the solution sampling pipe is provided with a solution sampling pipe control valve for controlling opening and closing of the solution sampling pipe.

25. a mesh chamber for accommodating tissue of the xenogeneic organ to be decellularized therein; At least one reaction vessel for storing a decellularization solution for carrying out a decellularization reaction with the tissue of the xenogeneic organ in the mesh chamber to be inserted therein; and A second reactor for extracting the decellularized solution undergoing a decellularization reaction in the reaction vessel to the outside of the reaction vessel and analyzing various physical properties of the decellularized solution ex-situ. A decellularization reactor comprising:

26. The decellularization reaction apparatus according to claim 25, comprising a solution circulation unit connected between the reaction vessel and the cover of the reaction vessel so as to be in communication with each other, for circulating a decellularization solution through the reaction vessel and the cover.

27. The decellularization reaction apparatus according to claim 26, wherein a drive unit is coupled to the mesh chamber for rotating the mesh chamber so as to cause agitation within the reaction vessel.

28. The decellularization reaction apparatus according to claim 27, wherein the cover is coupled to a lifting unit for vertically raising and lowering the cover relative to the upper end of the reaction vessel.

29. The decellularization reactor according to claim 25 , wherein a mesh is arranged in a lattice pattern on the outer circumferential surface of the mesh chamber.

30. The decellularization reactor according to claim 29 , wherein the size of the mesh is set to be smaller than the tissue of the xenogeneic organ to be decellularized.

31. The circulation section includes: a circulation tube connected between the cover and the reaction vessel for circulating a decellularization solution in the reaction vessel; and The decellularization reaction apparatus according to any one of claims 25 to 30, comprising a transfer pump installed in the circulation pipe for quantitatively transferring the decellularization solution circulating in the circulation pipe.

32. The decellularization reaction apparatus according to claim 31 , wherein the circulation tube is made of an expandable and contractible flexible tube.

33. The decellularization reaction apparatus according to claim 32, wherein the second reactor includes a solution sampling tube installed in the circulation tube for extracting and sampling the decellularization solution circulating through the circulation tube.

34. the reaction vessels are arranged on a base at set intervals and supported by a first support frame arranged vertically to the base; The decellularization reaction apparatus according to claim 31 , wherein an alignment groove for positional alignment between the reaction vessel and the cover is arranged on the upper end surface of the base.

35. The decellularization reaction apparatus according to claim 34, wherein an elastic member for cushioning impact of the reaction vessel is coupled to an upper end of the first support frame.

Images