Decellularization reaction device

Abstract

Provided is a decellularization reaction device. A decellularization reaction device according to the present invention is provided with: a grid chamber in which tissues of a heterogeneous organ to be decellularized are accommodated; at least one reaction vessel for storing a decellularized solution; and a first reactor for circulating the decellularized solution inside and outside the reaction vessel and performing in-situ analysis of various physical properties of the circulating decellularized solution.

Description

Technical Field

[0001] The present invention relates to a decellularization reaction device, and more particularly, to a decellularization reaction device for preparing bioink. Background Art

[0002] Recently, with the global aging trend, the number of patients with chronic diseases has increased, and the technology of manufacturing artificial organs using 3D bioprinting has attracted much attention.

[0003] Bioink refers to an ink material for outputting artificial organs through a 3D bioprinter, which is manufactured based on the extracellular matrix obtained by decellularizing xenogeneic organs.

[0004] This decellularization process of xenogeneic organs is the most core process in preparing bioink, which refers to removing all impurities and cells in the xenogeneic organs to avoid antigen-antibody reactions.

[0005] The decellularization process applied to 3D bioprinting technology is a method of treating xenogeneic organs by soaking them in a solution. So far, there is no dedicated device for decellularization, but simple laboratory glassware materials such as beakers are used as the device.

[0006] In addition, during the soaking process in various decellularization solutions, there is no specific index for the process of discharging the current solution and then transferring to injecting the next solution, so there is a problem that the physical properties and quality between the final products deviate. Summary of the Invention

[0007] Technical Problem

[0008] An object of the present invention is to provide a decellularization reaction device that can obtain indexes such as the treatment time of the decellularization solution, re-injection or transfer to the next solution, and the end of the decellularization process by measuring the conductivity, turbidity, and absorbance (UV-Vis) of the solution undergoing decellularization.

[0009] Technical Solution

[0010] The decellularization reaction device according to an embodiment of the present invention may include: a grid chamber for accommodating the tissue of a xenogeneic organ to be decellularized therein; at least one reaction container for storing a decellularization solution that undergoes a decellularization reaction with the tissue of the xenogeneic organ inserted into the grid chamber inside the reaction container.

[0011] In addition, the decellularization reaction device may include: a first reactor for discharging the decellularization solution undergoing a decellularization reaction in the reaction container from the reaction container and then circulating the decellularization solution back into the inside of the reaction container, and performing in-situ analysis of various physical properties of the circulated decellularization solution.

[0012] The decellularization reaction device may further include: a second reactor, which is used to extract the decellularization solution undergoing the decellularization reaction in the reaction vessel to the outside of the reaction vessel and perform ex-situ analysis on the physical properties of the decellularization solution.

[0013] Hereinafter, the first reactor may refer to a reactor using an in-situ analysis device, and analysis devices such as a turbidity meter, a conductivity meter, and an absorbance meter are installed on the reactor pipeline.

[0014] In addition, the second reactor may refer to a reactor that does not install analysis devices such as a turbidity meter, a conductivity meter, and an absorbance meter on the reactor pipeline, but uses a method of extracting and sampling the solution and then performing off-site measurement of the corresponding physical properties.

[0015] The first reactor may include: a solution circulation part, which is connected between the reaction vessel and the cover of the reaction vessel and is communicated with each other, and is used to circulate the decellularization solution between the reaction vessel and the cover.

[0016] The first reactor may include: a measurement part, which is installed on the solution circulation part and is used to perform in-situ measurement on various physical properties of the decellularization solution that is circulating in the solution circulation part and undergoing the decellularization reaction.

[0017] The grid chamber may be combined with a driving part, and the driving part is used to rotate the grid chamber in the reaction vessel for stirring.

[0018] A lifting part may be combined with the cover, and the lifting part is used to lift the cover relative to the upper end of the reaction vessel in the vertical direction.

[0019] The grid chamber may be configured in a hollow cylindrical shape inside.

[0020] The grid chamber may be made of a chemical-resistant material or coated with this material according to the physical properties of the decellularization solution.

[0021] Grids may be arranged in a lattice pattern on the outer peripheral surface of the grid chamber.

[0022] The size of the grids may be set to a size smaller than the tissue of the decellularized target xenogeneic organ.

[0023] A lid for covering the grid chamber may be combined with the upper end of the grid chamber.

[0024] The reaction vessel may be made of a transparent material or have a transparent window so as to be able to observe the inside.

[0025] The reaction vessel may be configured in a double-layer jacket shape including an inner tube and an outer tube so as to control the temperature of the decellularization process.

[0026] The reaction vessel may have a solution outlet for flowing out the decellularized solution.

[0027] A solution injection tube for injecting the decellularized solution into the interior of the reaction vessel may be connected to the cover.

[0028] The driving unit may include: a rotating shaft coupled to the lid of the mesh chamber; and a driving motor mounted on the cover of the reaction vessel while being coupled to the rotating shaft and configured to rotate the rotating shaft.

[0029] The solution circulation unit may include: a circulation pipe connected between the cover and the solution outlet for circulating the decellularized solution in the reaction vessel; and a delivery pump mounted on the circulation pipe for quantitatively delivering the decellularized solution circulated in the circulation pipe.

[0030] The circulation pipe may be formed of a flexible pipe capable of expanding and contracting.

[0031] The measurement unit may include an absorbance analyzer, a conductivity meter, and a turbidity meter for respectively measuring the absorbance, conductivity, and turbidity of the decellularized solution circulated in the circulation pipe.

[0032] The measurement unit may include a thermometer for measuring the temperature of the decellularized solution circulated in the circulation pipe.

[0033] The lifting unit may include: a coupling plate coupled to the cover and capable of lifting; and a lifting cylinder coupled to the coupling plate and having a lifting rod for lifting the coupling plate.

[0034] An air supply pipe for supplying air into the lifting cylinder may be connected to the lifting cylinder, and a regulator for adjusting the pressure of the air supplied to the air supply pipe may be mounted on the air supply pipe.

[0035] The second reactor may include: a solution sampling pipe mounted on the circulation pipe for extracting and sampling the decellularized solution circulated through the circulation pipe.

[0036] A solution sampling pipe control valve for controlling the opening and closing of the solution sampling pipe may be mounted on the solution sampling pipe.

[0037] The decellularization reaction device according to another implementation example of the present invention may include: a mesh chamber for accommodating therein a tissue of a decellularization target xenogeneic organ; and at least one reaction vessel for storing a decellularized solution for performing a decellularization reaction with the tissue of the xenogeneic organ inserted into the mesh chamber inside the reaction vessel.

[0038] In addition, the decellularization reaction device may include: a second reactor configured to extract the decellularization solution undergoing the decellularization reaction in the reaction vessel to the outside of the reaction vessel and perform ex-situ analysis on various physical properties of the decellularization solution.

[0039] The first reactor may include: a solution circulation section connected and communicating with each other between the reaction vessel and the lid of the reaction vessel, and configured to circulate the decellularization solution between the reaction vessel and the lid.

[0040] The mesh chamber may be coupled with a driving section configured to rotate the mesh chamber within the reaction vessel for stirring.

[0041] A lifting section may be coupled to the lid and configured to lift and lower the lid relative to the upper end of the reaction vessel in the vertical direction.

[0042] Meshes may be arranged in a lattice pattern on the outer peripheral surface of the mesh chamber.

[0043] The size of the meshes may be set to a size smaller than the tissue of the decellularization target xenogeneic organ.

[0044] The solution circulation section may include: a circulation pipe connected between the lid and the reaction vessel and configured to circulate the decellularization solution in the reaction vessel; and a transfer pump installed on the circulation pipe and configured to quantitatively transfer the decellularization solution circulating in the circulation pipe.

[0045] The circulation pipe may be composed of a flexible pipe capable of stretching.

[0046] A solution sampling pipe may be installed on the circulation pipe and configured to extract and sample the decellularization solution circulating through the circulation pipe.

[0047] The reaction vessels may be arranged on the base at set intervals and supported by a support frame arranged vertically relative to the base. Alignment grooves for aligning the positions of the reaction vessels and the lids are arranged on the upper end surface of the base.

[0048] A second elastic member for buffering the impact of the reaction vessel is coupled to the upper end of the support frame.

[0049] Advantageous Effects

[0050] According to an embodiment of the present invention, by measuring the conductivity, turbidity, absorbance (UV-Vis), etc. of the solution undergoing the decellularization reaction, it is possible to obtain indicators such as the processing time of the decellularization solution, reinjection or transfer to the next solution, and the end of the decellularization process.

[0051] In addition, a reaction vessel, a mesh chamber, and a solution circulation structure can be used to analyze various physical properties (such as conductivity, turbidity, and absorbance) of the solution in the decellularization process, and to obtain the discharge and reinjection of the decellularized solution and the end conditions of the decellularization process, thereby achieving excellent effects in obtaining high-quality decellularized materials while ensuring the uniformity among the materials. BRIEF DESCRIPTION OF THE DRAWINGS

[0052] Figure 1 is a schematic structural diagram of the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0053] Figure 2 is a schematic side view of the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0054] Figure 3 is a schematic structural diagram of the mesh chamber of the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0055] Figure 4 is a schematic structural diagram of the second reactor in the decellularization reaction device of the second embodiment of the present invention.

[0056] Figure 5 is a graph showing the analysis results of the conductivity of the decellularized solution measured during the porcine placenta decellularization process using the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0057] Figure 6 is a graph showing the analysis results of the turbidity of the decellularized solution measured during the porcine placenta decellularization process using the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0058] Figure 7 is a graph showing the analysis results of the absorbance (UV-Vis) of the decellularized solution measured during the porcine placenta decellularization process using the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0059] Figure 8 is a photograph showing the states before and after the porcine placenta decellularization process using the first reactor in the decellularization reaction device of the first embodiment of the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0060] Hereinafter, embodiments of the present invention will be described with reference to the accompanying drawings so that those with ordinary knowledge in the technical field to which the present invention pertains can easily implement the present invention. As those with ordinary knowledge in the technical field to which the present invention pertains can understand, the following-described embodiments can be deformed into various forms without departing from the concept and scope of the present invention. For the same or similar parts, the same reference numerals are used as much as possible in the drawings.

[0061] The technical terms used hereinafter are only for referring to specific embodiments and are not intended to limit the present invention. Unless the sentence clearly indicates the opposite meaning, the singular form used herein also includes the plural form. The meaning of "including" used in the specification embodies specific characteristics, regions, integers, steps, operations, elements, and / or components, but does not exclude the existence or addition of other specific characteristics, regions, integers, steps, operations, elements, components, and / or groups.

[0062] All terms, including technical terms and scientific terms, used hereinafter have the same meaning as commonly understood by those with ordinary knowledge in the technical field to which the present invention pertains. Terms defined in the dictionary are further interpreted to be consistent with the relevant technical literature and the currently disclosed content. Unless otherwise defined, they should not be interpreted as ideal or very formal meanings.

[0063] Figure 1 It is a schematic structural diagram of the first reactor in the decellularization reaction device of the first embodiment of the present invention. Figure 2 It is a schematic side view of the first reactor in the decellularization reaction device of the first embodiment of the present invention. Figure 3 It is a schematic structural diagram of the grid chamber of the first reactor in the decellularization reaction device of the first embodiment of the present invention.

[0064] Referring to Figures 1 to 3 , the decellularization reaction device of the first embodiment of the present invention may include a grid chamber 100, a reaction vessel 200, and a first reactor.

[0065] The grid chamber 100 may have a set size and shape, and a xenogeneic organ tissue to be decellularized can be accommodated inside.

[0066] In addition, at least one reaction vessel 200 may be provided, the grid chamber 100 may be inserted inside the reaction vessel 200, and the reaction vessel 200 may store a decellularization solution for performing a decellularization reaction with the xenogeneic organ tissue inside the grid chamber 100.

[0067] After the first reactor discharges the decellularization solution that is undergoing a decellularization reaction with the acellularized xenogeneic organ tissue in the reaction vessel 200 from the reaction vessel 200, it can be recycled back into the interior of the reaction vessel 200, and at the same time, various physical properties of the first decellularization solution during circulation can be analyzed in-situ.

[0068] In addition, the first reactor may include a driving unit 300, a solution circulation unit 400, and a measurement unit 500.

[0069] The driving unit 300 can be coupled to the mesh chamber 100 and rotate the mesh chamber 100 within the reaction vessel 200 for stirring.

[0070] In addition, the solution circulation unit 400 is connected between the reaction vessel 200 and the cover 210 covering the reaction vessel 200 and is in communication with each other, and is used to circulate the decellularization solution between the reaction vessel 200 and the cover 210.

[0071] The measurement unit 500 can be installed on the solution circulation unit 400, and it can perform measurements so as to be able to perform in-situ analysis of various physical properties of the decellularization solution that is circulating in the solution circulation unit 400 and is undergoing a decellularization reaction.

[0072] Above the reaction vessel 200, a cover 210 for covering the reaction vessel 200 can be arranged.

[0073] In addition, the decellularization reaction device may include a lifting unit 600, which is coupled to the cover 210 and is used to lift the cover 300 in the vertical direction ( Figure 1 the Y direction in

[0074] so that the cover 210 can be coupled to or separated from the reaction vessel 200.

[0075] The mesh chamber 100 can be configured to have a hollow shape with a hollow interior so that acellularized xenogeneic organ tissue, etc. can be accommodated inside.

[0076] In addition, when the mesh chamber 100 has a quadrilateral shape, when the mesh chamber 100 rotates, the xenogeneic organ tissue will be caught or retained at the corner parts, resulting in uneven stirring of the xenogeneic organ tissue. Therefore, in order to stir the xenogeneic organ tissue evenly and efficiently, the mesh chamber 100 is designed to have a cylindrical shape, etc.

[0077] On the outer peripheral surface of the mesh chamber 100, a mesh 101 can be arranged in a lattice pattern. In order to enable the acellularized xenogeneic organ tissue, etc. accommodated in the mesh chamber 100 to undergo a decellularization reaction, the mesh 101 can have a set size, for example, a size of about 1 mm.

[0078] The mesh chamber 100 can be configured as a chamber larger than the mesh chamber 100A of the second embodiment, and the mesh chamber 100 can be configured as a large chamber capable of accommodating, for example, 500 g (up to 1 Kg) of acellularized xenogeneic organs of a xenogeneic object.

[0079] In addition, the size of the mesh 101 can be set to be slightly smaller than the size of the xenogeneic organ tissue, so that the xenogeneic organ tissue accommodated in the mesh chamber 100 does not leak outside the mesh 101 during the acellularization process.

[0080] The mesh chamber may include a lid 110, which can be detachably coupled to the upper end of the mesh chamber 100 and is used to cover the mesh chamber 100.

[0081] The lid 110 can be in the form of a buckle 111 or the like to facilitate easy coupling with the mesh chamber 100.

[0082] A drive unit 300 can be coupled to the upper end of the lid 110.

[0083] The mesh chamber 100 can be made of a material with excellent chemical resistance such as Teflon or coated with such a material according to the physical properties of the acellularization solution.

[0084] In addition, the reaction vessel 200 can be made of a transparent material or have a transparent window (not shown) so that the progress of the acellularization process occurring inside the reaction vessel 200 can be visually observed from the outside of the reaction vessel 200.

[0085] The reaction vessels 200 can be arranged on the base 10 at set intervals and are supported by a first support frame 20, and the first support frame 20 is arranged along the vertical direction ( Figure 1 the Y direction in

[0086] The reaction vessel 200 can be made of a material that is corrosion-resistant to the acellularization solution to be used.

[0087] In addition, the reaction vessel 200 can be in the form of a double jacket composed of an inner tube 201 and an outer tube 203 to control the temperature of the acellularization process.

[0088] A coolant inlet 205 and a coolant outlet 206 can be respectively provided on the outer tube 203. The coolant inlet 205 is used to inject coolant into the space between the outer tube 203 and the inner tube 201, and the coolant outlet 206 is used to discharge the coolant to the outside.

[0089] The reaction vessel 200 can have a solution outlet 207 at its lower end for the acellularization solution to flow out.

[0090] The cover 210 can be made of a shock-absorbing material such as rubber and have a shape such as an O-ring, so that when it descends through the lifting part 600 and contacts the upper end of the reaction vessel 200, it can seal the reaction vessel 200 while buffering the impact.

[0091] At least one injection port 211 for connecting the solution injection tube 220 can be installed on the cover 210, and the solution injection tube 220 is used to inject the decellularized solution into the interior of the reaction vessel 200.

[0092] Various decellularized solutions can be composed of, for example, ultrapure water, organic solvents, etc. Various decellularized solutions can be injected through each injection port 211, and a pump (not shown) for quantitatively supplying each decellularized solution can be installed on the pipeline (not shown) connected to each injection port 211.

[0093] In addition, the solution circulation part 400 can be connected to the cover 210, so that the decellularized solution circulated in the solution circulation part 400 can be injected into the interior of the reaction vessel 200.

[0094] A water level measuring instrument 213 such as an ultrasonic wave for measuring the water level of the decellularized solution in the reaction vessel 200 can be installed on the cover 210.

[0095] The driving part 300 can include: a rotating shaft 310 combined with the lid 110 of the grid chamber 100; a driving motor 320, which is installed on the cover 210 of the reaction vessel 200 and combined with the rotating shaft 310, and is used to rotate the rotating shaft 310.

[0096] The driving motor 320 can be composed of a magnetically controlled motor or the like, so that the rotating shaft 310 can be easily rotated.

[0097] The rotating shaft 310 can be arranged at the center of the upper end of the lid 110 along the vertical direction ( Figure 2 the Y direction) of the upper end of the lid 110, and can be combined with the lid 110 through combination with a bolt 301 or the like.

[0098] The rotating shaft 310 can be rotated, for example, clockwise or counterclockwise by the driving motor 320, so as to drive the grid chamber 100 to rotate to stir the decellularized target xenogeneic organ tissue accommodated in the grid chamber 100, so as to guide the xenogeneic organ tissue to smoothly achieve decellularization under the action of the decellularized solution injected into the reaction vessel 200.

[0099] When the grid chamber 100 is rotated and stirred at a set rotation speed (for example, 0 - 300 rpm), in order to prevent overload of the driving motor 320 and prevent entanglement between xenogeneic organs, it can be stirred alternately from the forward direction (clockwise direction) to the reverse direction (counterclockwise direction), and a pause time can be set in the middle.

[0100] The solution circulation unit 400 may include a circulation pipe 410 and a delivery pump 420.

[0101] The circulation pipe 410 may be connected between the cover 210 and the solution outlet 207 to circulate the decellularized solution in the reaction vessel 200.

[0102] In addition, the delivery pump 420 may be installed on the circulation pipe 410 to quantitatively deliver the decellularized solution circulating in the circulation pipe 410.

[0103] The circulation pipe 410 may be composed of a flexible pipe or the like so as to be easily stretched and deformed in shape when the cover 210 is lifted and lowered by the lifting unit 600.

[0104] In addition, a waste liquid discharge pipe 430 for discharging the waste liquid of the decellularized solution may be connected to the circulation pipe 410, and a waste liquid discharge pipe control valve 431 for controlling the discharge of the decellularized solution may be installed on the waste liquid discharge pipe 430.

[0105] The measurement unit 500 may include an absorbance analyzer 510 for analyzing the absorbance (UV-Vis) of the decellularized solution circulating in the circulation pipe 410.

[0106] In addition, the measurement unit 500 may include a conductivity meter 520 for measuring the conductivity of the decellularized solution circulating in the circulation pipe 410.

[0107] The measurement unit 500 may include a turbidity meter 530 for measuring the turbidity of the decellularized solution circulating in the circulation pipe 410.

[0108] The measurement unit 500 may include a thermometer 540 for measuring the temperature of the decellularized solution circulating in the circulation pipe 410.

[0109] In addition, the lifting unit 600 may include: a coupling plate 610 coupled to the cover 210 so as to be liftable integrally therewith; and a lifting cylinder 620 coupled to the lower end of the coupling plate 610 and having a lifting rod 621 for lifting the coupling plate 620.

[0110] The lifting cylinder 620 may be installed and supported on the upper end surface of the base 10 relative to the base 10 in the vertical direction ( Figure 2 the Y direction in

[0111] An air supply pipe 623 for supplying air into the lifting cylinder 620 may be connected to the lifting cylinder 620, and a regulator 625 for adjusting the air pressure supplied to the air supply pipe 623 may be installed on the air supply pipe 623.

[0112] In addition, a second support frame 30 may be installed on the base 10. The second support frame 30 is arranged to be parallel to the first support frame 20 and is supported on the base.

[0113] A guide rod 630 may be coupled to the second support frame 30. The guide rod 630 is connected to the second support frame 30 through a connection frame 31 and is used to guide the lifting of the lifting rod 621.

[0114] Next, with reference to Figures 1 to 3 , the operation of the decellularization reaction device according to the first embodiment of the present invention will be described.

[0115] First, prepare a decellularization target xenogeneic organ tissue and a decellularization solution for performing a decellularization reaction with the xenogeneic organ tissue.

[0116] To inject the decellularization target xenogeneic organ tissue and the decellularization solution into the grid chamber 100 and the reaction vessel 200 respectively, as Figure 1 shown, the lid 110 is set to an open state from the grid chamber 100, and the cover 210 is set to an open state from the reaction vessel 200.

[0117] In this state, the decellularization target xenogeneic organ tissue is placed inside the grid chamber 100, and the upper end of the grid chamber is covered with the lid 110. Then, the lid 110 is coupled to the grid chamber 100 using the buckle 111.

[0118] In addition, when the lifting rod 621 of the lifting cylinder 620 of the lifting unit 600 is contracted, the lifting rod 621 descends by a set length. Subsequently, the cover 210 covers the upper part of the reaction vessel 200, thereby sealing the reaction vessel 200.

[0119] In this way, as the cover 210 seals the reaction vessel 200, the grid chamber 100 is inserted into the interior of the reaction vessel 200.

[0120] At this time, the decellularization solution is injected into the reaction vessel 200 through the injection port 211 installed on the cover 210 and connected to the solution injection tube 220.

[0121] In addition, after the injection of the decellularization solution into the reaction vessel 200 is completed, the drive motor 320 of the drive unit 300 is started to rotate the rotary shaft 310 at a set speed in the clockwise or counterclockwise direction, so that the grid chamber 100 rotates and stirs.

[0122] As the grid chamber 100 rotates and stirs in this way, the xenogeneic organ tissue inside the grid chamber 100 is stirred while undergoing a decellularization reaction with the decellularization solution inside the reaction vessel 200.

[0123] At this time, since the xenogeneic organ tissue is stirred under the rotation of the grid chamber 100, and the decellularized solution moves vertically from the cover 210 to the inside of the reaction vessel 200 through the circulation pipe 410, the xenogeneic organ tissue and the decellularized solution are sufficiently stirred, so that a decellularized solution with a uniform concentration can be obtained in real time.

[0124] In this state, the delivery pump 420 is started to circulate the decellularized solution undergoing the decellularization reaction in the reaction vessel 200 through the circulation pipe 410 at a set speed.

[0125] For the physical properties of the decellularized solution circulated from the solution outlet 207 of the reaction vessel 200 to the cover 210 through the circulation pipe 410, real-time measurement can be performed by the measurement unit 500 installed on the circulation pipe 410.

[0126] That is, the absorbance (UV-Vis) of the decellularized solution is measured and analyzed by the absorbance analyzer 510, the conductivity of the decellularized solution is measured by the conductivity meter 520, the turbidity of the decellularized solution is measured by the turbidity meter 530, and the temperature of the decellularized solution is measured by the thermometer 540.

[0127] In addition, by measuring the decellularized solution with a uniform concentration, it is easy to analyze various physical properties of the decellularized solution (such as absorbance (UV-Vis), conductivity, turbidity, etc.), so that the progress of the decellularization process can be accurately grasped in real time.

[0128] (Example)

[0129] Next, a decellularization experiment on a porcine placenta, which is an example of a xenogeneic organ to be decellularized, will be described.

[0130] First, after putting 500 g of a porcine placenta that has undergone pretreatment such as impurity removal into the grid chamber 100 in the decellularization reaction vessel 200, the lid 110 of the grid chamber 100 is closed. Since the porcine placenta can be decellularized at room temperature of about 20°C, there is no need to specifically set the temperature.

[0131] In addition, by pressurizing the lifting cylinder 620 with gas (air), the reaction vessel 200 is covered with the cover 210 for sealing.

[0132] After injecting the decellularized solution into the reaction vessel 200, the speed of the rotating shaft 310 driven by the driving motor 320 is set to 80 rpm, and the grid chamber 100 is rotated for stirring, for example, in the clockwise direction for 10 seconds, with a pause time of 8 seconds, and in the counterclockwise direction for 10 seconds.

[0133] Meanwhile, the decellularized solution inside the reaction vessel 200 is circulated through the circulation pipe 410 by using the metering pump 420. At this time, the circulation speed of the decellularized solution is set to 1,000 ml / minute.

[0134] For the decellularized solution in the reaction vessel 200, a uniform concentration can be obtained by the circulation using the delivery pump 420 and the stirring performed by the rotation of the grid chamber 100.

[0135] Then, in the case of the reaction vessel 100 of the first embodiment, various physical property data of the decellularized solution in the decellularization process can be analyzed in real time by using the absorbance (UV-Vis) analyzer 510, the conductivity meter 520, and the turbidity meter 530 installed on the circulation pipe 410 (refer to Figure 1 ).

[0136] In addition, in the case of the reaction vessel 100A of the second embodiment, various physical properties of the decellularized solution can be analyzed by sampling the decellularized solution through the solution sampling pipe 550 installed on the circulation pipe 410 (refer to Figure 4 ).

[0137] Figure 5 is a chart showing the analysis results of the conductivity of the decellularized solution measured during the porcine placenta decellularization process, Figure 6 is a chart showing the analysis results of the turbidity of the decellularized solution measured during the porcine placenta decellularization process.

[0138] Figure 5 and Figure 6 are the results of analyzing the decellularized solution while performing the three ultra-pure water processes. Among them, ① refers to the case of performing the first ultra-pure water process, ② refers to the case of performing the second ultra-pure water process, and ③ refers to the case of performing the third ultra-pure water process.

[0139] Figure 7 is a chart showing the analysis results of the absorbance (UV-Vis) of the decellularized solution measured during the porcine placenta decellularization process.

[0140] In Figure 7 , ①, ②, ③, ④, ⑤, and ⑥ are the absorbance data over time during the first ultra-pure water process.

[0141] Among them, ① is the data measured after 10 minutes, ② is the data measured after 20 minutes, ③ is the data measured after 30 minutes, ④ is the data measured after 40 minutes, ⑤ is the data measured after 50 minutes, and ⑥ is the data measured after 60 minutes.

[0142] In the decellularization process, over time, the numerical values of the physical properties in the reactor show a tendency of initially increasing and then becoming slow. When the decellularization solution is discharged and the same decellularization solution is injected again, it can be confirmed that, as shown in ①-②-③ of Figure 5 and Figure 6 , the change in physical properties is very large.

[0143] Thus, while monitoring various physical property values of the decellularization solution, it is possible to determine whether it is necessary to inject various decellularization solutions again or the process end conditions, and as shown in Figure 8 , this process-guided decellularization can proceed stably as shown in Figure 8 . Therefore, the quality of the finally decellularized placental tissue material is high, and its uniformity is ensured.

[0144] (Test results of performance, etc.)

[0145] When evaluating the quality of bioink, the most basic is the analysis of DNA content and collagen content.

[0146] Table 1 shows the results of comparing the DNA content of porcine placentas before and after using the decellularization reaction device. Each porcine placenta is from a different supplier, so there is a large deviation in the DNA content of each porcine placenta.

[0147] [Table 1] Comparison of DNA content of porcine placenta tissue before and after using the decellularization reaction device

[0148] DNA content Placenta A (Supplier A) Placenta B (Supplier B) Placenta C (Supplier C) Before decellularization 87.2 ng / mg 77.5 ng / mg 244.8 ng / mg After decellularization 0.37 ng / mg 0.48 ng / mg 0.46 ng / mg

[0149] From the above results, it can be seen that the DNA content before using the decellularization reaction device is 87 - 244 ng / mg, while the DNA content after using the decellularization reaction device is 0.37 - 0.48 ng / mg, showing an extremely low content level.

[0150] This not only reduces the DNA content, but also shows high quality and uniform results even when using placentas from pigs raised in different places, which is of great significance.

[0151] [Table 2] Comparison of collagen content of porcine placenta tissue before and after using the decellularization device

[0152] Collagen content Placenta A (Supplier A) Placenta B (Supplier B) Placenta C (Supplier C) Before decellularization 10.1 μg / mg 8.9 μg / mg 6.5 μg / mg After decellularization 22.4 μg / mg 23.2 μg / mg 22.3 μg / mg

[0153] Table 2 shows the results of comparing the collagen content in porcine placentas before and after using the decellularization device. From the above results, it can be seen that after using the decellularization reaction device, the collagen content is still at the level of 22 - 23 μg / mg and is well preserved.

[0154] Therefore, after using the decellularization reaction device, the DNA content in the tissue drops to an extremely low level, while the collagen in the tissue is well preserved, indicating that high-quality bioink materials can be produced.

[0155] Figure 4 It is a schematic side view of the second reactor in the decellularization reaction device of the second embodiment of the present invention.

[0156] Except for the content specifically described below, the second reactor in the decellularization reaction device of the second embodiment of the present invention is the same as the content described for the first reactor in the decellularization reaction device of the first embodiment of the present invention, so its detailed description is omitted.

[0157] Refer to Figure 4 , the decellularization reaction device of the second embodiment of the present invention may include a second reactor for extracting the decellularization solution that is undergoing a decellularization reaction with the decellularization target xenogeneic organ tissue in the reaction vessel 200 to the outside of the reaction vessel 200 and performing ex-situ analysis on various physical properties of the decellularization solution.

[0158] The second reactor may include a solution sampling tube 550 installed on the circulation tube 410 of the solution circulation unit 400 for extracting and sampling the decellularization solution circulated through the circulation tube 410.

[0159] After the decellularization solution sampled by the solution sampling tube 550 can be loaded into a small container (not shown) such as a vial, ex-situ measurement and analysis are performed on various physical properties (absorbance, conductivity, turbidity, etc.) of the decellularization solution loaded into the small container.

[0160] A solution sampling tube control valve 551 for controlling the opening and closing of the solution sampling tube 550 may be installed on the solution sampling tube 550.

[0161] 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 chamber that can accommodate at most less than 25 g of the decellularization target xenogeneic organ. Accordingly, the size of the mesh 101A of the mesh chamber 100A can also be set to be small, that is, set to be less than 1 mm.

[0162] The reaction vessels 200 may be arranged on the base 10 at set intervals and supported by the first support frame 20, and the first support frame 20 is arranged relative to the base 10 along the vertical direction ( Figure 1 the Y direction in

[0163] For easy cleaning, the reaction vessel 200 and the first support frame 20 can be configured to be movable and detachable, and alignment grooves 11 for aligning the positions of the reaction vessel 200 and the cover 210 can be arranged on the upper end surface of the base 10.

[0164] An elastic member 21 can be coupled to the upper end portion of the first support frame 20, and the elastic member 21 is used to buffer the impact generated by the reaction vessel 200 when the cover 210 descends and seals the reaction vessel 200.

[0165] Although the present disclosure has been illustrated by the preferred embodiments as described above, the present invention is not limited thereto, and those skilled in the art to which the present invention pertains should easily understand that various modifications and deformations can be made without departing from the scope of the appended claims.

[0166] Description of Reference Numerals

[0167] 100: Grid chamber

[0168] 200: Reaction vessel

Claims

1. A decellularization reaction device, comprising: A grid chamber for accommodating the tissue of a xenogeneic organ, the decellularization target, inside; At least one reaction vessel for storing a decellularization solution that undergoes a decellularization reaction with the tissue of the xenogeneic organ inserted into the grid chamber inside the reaction vessel; and A first reactor for discharging the decellularization solution that is undergoing a decellularization reaction in the reaction vessel from the reaction vessel and then circulating the decellularization solution back into the interior of the reaction vessel again, and for in-situ analysis of various physical properties of the circulated decellularization solution.

2. The decellularization reaction device according to claim 1, further comprising: A second reactor for extracting the decellularization solution that is undergoing a decellularization reaction in the reaction vessel to the outside of the reaction vessel and for off-site analysis of the decellularization solution.

3. The decellularization reaction device according to claim 2, wherein The first reactor includes: a solution circulation part connected between the reaction vessel and the cover of the reaction vessel and communicating with each other, the solution circulation part for circulating the decellularization solution between the reaction vessel and the cover.

4. The decellularization reaction device according to claim 3, wherein The first reactor includes: a measurement part installed on the solution circulation part for in-situ measurement of various physical properties of the decellularization solution that is circulating in the solution circulation part and undergoing a decellularization reaction.

5. The decellularization reaction device according to claim 4, wherein The grid chamber is combined with a driving part for rotating the grid chamber inside the reaction vessel for stirring.

6. The decellularization reaction device according to claim 5, wherein A lifting part is combined on the cover for lifting the cover vertically relative to the upper end of the reaction vessel.

7. The decellularization reaction device according to claim 6, wherein The grid chamber is configured in a hollow cylindrical shape inside.

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

9. The decellularization reaction device according to claim 7, wherein On the outer peripheral surface of the grid chamber, grids are arranged in a lattice pattern.

10. The decellularization reaction device according to claim 9, wherein The size of the grids is set to be smaller than the size of the tissue of the xenogeneic organ, the decellularization target.

11. The decellularization reaction device according to claim 10, wherein A lid for covering the grid chamber is combined at the upper end of the grid chamber.

12. The decellularization reaction device 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 as to be able to observe the interior.

13. The decellularization reaction device according to claim 12, wherein The reaction vessel is configured in a double-jacket form including an inner tube and an outer tube so as to control the temperature of the decellularization process.

14. The acellular reaction device according to claim 13, wherein the reaction vessel has a solution outlet for flowing out the acellular solution.

15. The acellular reaction device according to claim 14, wherein a solution injection tube for injecting the acellular solution into the interior of the reaction vessel is connected to the cover.

16. The acellular reaction device according to claim 11, wherein the driving part includes: a rotating shaft, which is combined with the lid of the grid chamber; and a driving motor, which is installed on the cover of the reaction vessel and combined with the rotating shaft, and is used to rotate the rotating shaft.

17. The acellular reaction device according to claim 14, wherein the solution circulation part includes: a circulation pipe, which is connected between the cover and the solution outlet, and is used to circulate the acellular solution in the reaction vessel; and a delivery pump, which is installed on the circulation pipe and is used to quantitatively deliver the acellular solution circulated in the circulation pipe.

18. The acellular reaction device according to claim 17, wherein the circulation pipe is composed of a flexible pipe that can be telescoped.

19. The acellular reaction device according to claim 18, wherein the measuring part includes an absorbance analyzer, a conductivity meter, and a turbidity meter for respectively measuring the absorbance, conductivity, and turbidity of the acellular solution circulated in the circulation pipe.

20. The acellular reaction device according to claim 19, wherein the measuring part includes a thermometer for measuring the temperature of the acellular solution circulated in the circulation pipe.

21. The acellular reaction device according to claim 6, wherein the lifting part includes: a connecting plate, which is combined with the cover and can be lifted and lowered; and a lifting cylinder, which is combined with the connecting plate and has a lifting rod for lifting and lowering the connecting plate.

22. The acellular reaction device according to claim 21, wherein 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 is installed on the air supply pipe.

23. The acellular reaction device according to claim 2, wherein the second reactor includes: a solution sampling pipe, which is installed on the circulation pipe and is used to extract and sample the acellular solution circulated through the circulation pipe.

24. The acellular reaction device according to claim 23, wherein a solution sampling pipe control valve for controlling the opening and closing of the solution sampling pipe is installed on the solution sampling pipe.

25. An acellular reaction device, comprising: a grid chamber for accommodating the tissue of a xenogeneic organ to be acellularized therein; at least one reaction vessel for storing an acellular solution for performing an acellular reaction with the tissue of the xenogeneic organ inserted into the grid chamber inside the reaction vessel; and a second reactor for extracting the acellular solution undergoing an acellular reaction in the reaction vessel to the outside of the reaction vessel and performing an ex situ analysis of the physical properties of the acellular solution.

26. The acellular reaction device according to claim 25, comprising: The solution circulation part is connected between the reaction vessel and the cover of the reaction vessel and communicates with each other, and is used to circulate the decellularized solution between the reaction vessel and the cover.

27. The decellularization reaction device according to claim 26, wherein, The grid chamber is combined with a driving part, and the driving part is used to rotate the grid chamber in the reaction vessel for stirring.

28. The decellularization reaction device according to claim 27, wherein, A lifting part is combined on the cover, and the lifting part is used to lift the cover relative to the upper end of the reaction vessel in the vertical direction.

29. The decellularization reaction device according to claim 25, wherein, Grids are arranged in a lattice form on the outer peripheral surface of the grid chamber.

30. The decellularization reaction device according to claim 29, wherein, The size of the grid is set to be smaller than the size of the tissue of the decellularization target xenogeneic organ.

31. The decellularization reaction device according to any one of claims 25 to 30, wherein, The solution circulation part includes: A circulation pipe, which is connected between the cover and the reaction vessel and is used to circulate the decellularized solution in the reaction vessel; and A delivery pump, which is installed on the circulation pipe and is used to quantitatively deliver the decellularized solution circulating in the circulation pipe.

32. The decellularization reaction device according to claim 31, wherein, The circulation pipe is composed of a flexible pipe that can be stretched.

33. The decellularization reaction device according to claim 32, wherein, The second reactor includes: a solution sampling pipe, which is installed on the circulation pipe and is used to extract and sample the decellularized solution circulating through the circulation pipe.

34. The decellularization reaction device according to claim 31, wherein, The reaction vessels are arranged on the base at a set interval and are supported by a first support frame arranged in the vertical direction relative to the base, An alignment groove for aligning the positions of the reaction vessel and the cover is arranged on the upper end surface of the base.

35. The decellularization reaction device according to claim 34, wherein, An elastic member for buffering the impact of the reaction vessel is combined at the upper end of the first support frame.

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