WO2022045854A1 - Cell culture apparatus for producing cultured meat - Google Patents

Cell culture apparatus for producing cultured meat Download PDF

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Publication number
WO2022045854A1
WO2022045854A1 PCT/KR2021/011613 KR2021011613W WO2022045854A1 WO 2022045854 A1 WO2022045854 A1 WO 2022045854A1 KR 2021011613 W KR2021011613 W KR 2021011613W WO 2022045854 A1 WO2022045854 A1 WO 2022045854A1
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canister
cell
cell culture
culture apparatus
differentiation
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PCT/KR2021/011613
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French (fr)
Korean (ko)
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이희재
금준호
손예빈
조성천
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주식회사 씨위드
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Publication of WO2022045854A1 publication Critical patent/WO2022045854A1/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M21/00—Bioreactors or fermenters specially adapted for specific uses
    • C12M21/08—Bioreactors or fermenters specially adapted for specific uses for producing artificial tissue or for ex-vivo cultivation of tissue
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
    • A23L13/00—Meat products; Meat meal; Preparation or treatment thereof
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/12—Apparatus for enzymology or microbiology with sterilisation, filtration or dialysis means
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/34—Measuring or testing with condition measuring or sensing means, e.g. colony counters
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/42—Apparatus for the treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/02—Form or structure of the vessel
    • C12M23/06—Tubular
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/26—Constructional details, e.g. recesses, hinges flexible
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
    • C12M29/18—External loop; Means for reintroduction of fermented biomass or liquid percolate
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M3/00—Tissue, human, animal or plant cell, or virus culture apparatus
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
    • C12M35/06—Magnetic means
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
    • C12M35/08—Chemical, biochemical or biological means, e.g. plasma jet, co-culture
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/40—Means for regulation, monitoring, measurement or control, e.g. flow regulation of pressure
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N13/00—Treatment of microorganisms or enzymes with electrical or wave energy, e.g. magnetism, sonic waves
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
    • C12N5/06—Animal cells or tissues; Human cells or tissues
    • C12N5/0602—Vertebrate cells
    • C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
    • C12N5/0658—Skeletal muscle cells, e.g. myocytes, myotubes, myoblasts

Definitions

  • the present invention relates to a cell culture apparatus for producing cultured meat and a cultured meat production system including the apparatus.
  • Substitute meat (replacement meat, artificial meat) can be divided into plant meat made with plant materials such as soybeans, wheat, and rice to taste meat, and cultured meat (cultured meat) made by culturing animal cells.
  • Cultured meat refers to edible meat obtained by harvesting cells from living animals and then proliferating the cells using cell engineering technology, a field of cellular agriculture that obtains meat without raising livestock it can be said
  • Cultured meat production is a complex process that requires consideration of various engineering considerations, such as cells, scaffolds, media and growth factors, and bioreactors.
  • the dual bioreactor supplies oxygen and medium so that the cells attached to the scaffold can grow into cultured meat, the final product, and removes wastes.
  • a rotating bioreactor is often used, but various bioreactors may be used depending on the type of cultured meat to be produced, such as a tank stirring type or a stationary culture type.
  • the technology for producing cultured meat having the taste and texture of general meat is still lacking.
  • An object of the present invention is to provide an ingestible seaweed-derived cell culture support sheet, a cell culture apparatus using a plurality of cell culture support fibers, and a cultured meat production system using the cell culture apparatus.
  • an aspect of the present invention provides a cell culture device including a cell division canister, a cell differentiation canister, a canister fixed end, and a housing.
  • the cell division canister includes a multi-layered cell culture support sheet, an outer shell surrounding the support sheet to form a space separated from the outside, and a multi-channel tube connected to the outer shell.
  • the cell differentiation canister includes a plurality of cell culture support fibers aligned in one direction, an outer shell surrounding the support fibers to form a space separated from the outside, and a multi-channel tube connected to the outer shell.
  • the fixed end of the canister is connected to both ends of the cell differentiation canister to fix the cell differentiation canister, and can move up and down along the moving part of the fixed end of the canister.
  • the housing can be opened and closed, and a cell division canister and a cell division canister are positioned therein, and fixed end moving parts of the canister are installed on both walls.
  • the cell culture apparatus cultured cells selected from the group consisting of embryonic stem cells, muscle stem cells, mesenchymal stem cells, induced pluripotent stem cells, myoblasts and fibroblasts, and finally can produce muscle tissue.
  • the multi-channel tube may include a medium supply tube, a medium recovery tube, and a gas supply tube.
  • the cell differentiation canister may further include a coil for applying an electric field surrounding the canister, and the coil for applying the electric field may periodically apply an electric field of 30 to 90 mV simulating an action potential of a muscle.
  • the cell culture support sheet and the cell culture support fiber have a hydrogel structure containing alginate and cellulose left after removing the cell components from the medulla of seaweed, and the seaweed may include at least one of seaweed, kelp, and shiitake. there is.
  • the cell division canister and the cell differentiation canister may further include sensors for measuring the glucose concentration and pH of the medium.
  • the cell culture apparatus may further include a controller that adjusts the temperature and carbon dioxide concentration in the housing and/or an actuator that moves the fixed end of the canister.
  • the cell differentiation canister contracts and relaxes by the movement of the fixed end of the canister, and the outer shells of the two canisters are made of a polymer material that can be contracted and relaxed so as not to damage the outer shell of the canister in the process, and the tensile strength is 700 % or more, and the elongation may be 20 kN/m or more.
  • Another aspect of the present invention provides a cultured meat production system comprising the following configuration:
  • At least one cell medium supply unit connected to the multi-channel tube of the cell culture device;
  • a cell medium recovery unit connected to the multi-channel tube of the cell culture device
  • a gas mixing unit connected to the multi-channel tube of the cell culture apparatus and including a gas tank and a mixer;
  • one or more peristaltic pumps connected to the cell medium supply unit and the cell medium recovery unit;
  • a microcontroller that controls the operation of the cell culture device, peristaltic pump, and gas mixing section.
  • the cell medium supply unit may include a division medium supply unit and a differentiation medium supply unit.
  • the cell culture apparatus of the present invention uses a cell culture support derived from seaweed that can be ingested, and has the advantage of being able to apply physical and electrical stimulation necessary for differentiation into muscle tissue to cells to be cultured.
  • FIG. 1 schematically shows the structure of a cell culture apparatus including a cell division canister, a cell differentiation canister, a canister fixed end, and a housing.
  • FIG. 2 shows a state in which a cell culture support sheet is formed in multiple layers in a cell division canister (A) and the overall structure of the canister (B).
  • FIG 3 shows a state in which a plurality of cell culture support fibers are aligned in one direction in a cell differentiation canister (A) and the overall structure of the canister (B).
  • FIG. 4 shows a view of a cell differentiation canister connected to a fixed end of the canister.
  • FIG. 5 shows a process of seeding cells in a cell differentiation canister and then differentiating them into muscle tissue through physical and electrical stimulation.
  • FIG. 6 shows a cultured meat production system according to an example of the present invention.
  • the embodiments of the present invention specifically represent ideal embodiments of the present invention. As a result, various modifications of the drawings are expected. Accordingly, the embodiment is not limited to a specific shape of the illustrated area, and includes, for example, a shape modification by manufacturing.
  • the cell culture apparatus 100 of the present invention includes a cell division canister 110 , a cell differentiation canister 120 , a canister fixing end 130 , and a housing 140 .
  • the cell culture apparatus 100 may be used for culturing cells selected from the group consisting of embryonic stem cells, muscle stem cells, mesenchymal stem cells, induced pluripotent stem cells, myoblasts and fibroblasts, and finally the cells After culturing, it can be differentiated to form muscle tissue (cultured meat).
  • the cell division canister 110 will be described with reference to FIGS. 2 and 6 .
  • the cell division canister 110 includes a multi-layered cell culture support sheet 111 , a shell 112 surrounding the support sheet to form a space separated from the outside, and a multi-channel tube 113 connected to the shell.
  • a plurality of cell division canisters 110 may be included in the housing.
  • the cell division canister 110 performs a function of proliferating the seeded cells.
  • Cell division medium is supplied using the medium supply tube 113-1, which is one of the multi-channel tubes 113, and the cells are seeded on the multi-layered cell culture support sheet 111, followed by general cell culture conditions (37° C., Cells can be cultured in 5% CO 2 ).
  • general cell culture conditions 37° C., Cells can be cultured in 5% CO 2 .
  • FIG. 2 it can be seen that the cell culture support sheet of the cell division canister is multi-layered (A) and the overall structure (B).
  • the multi-layered cell culture support sheet 111 may be used as at least one layer or more, and may include clip-type fixing devices 111-1 at both ends of the sheet so that the multi-layer structure can be maintained.
  • the cell division canister may further include a sensor 114 for measuring the glucose concentration and pH of the medium, and when the sensor detects a decrease in the glucose concentration and pH of the medium, one of the multi-channel tubes 113
  • the old medium may be removed through the phosphorus medium recovery tube 113-2, and the fresh cell division medium may be supplied through the medium supply tube 113-1.
  • the cell division canister 110 may use one of the multi-channel tubes 113 as the gas supply tube 113-3 to receive nitrogen, carbon dioxide, and oxygen necessary for cell culture.
  • the multi-channel tube 113 of the cell division canister 110 includes a medium supply tube 113-1, a medium recovery tube 113-2, and a gas supply tube 113-3.
  • Each of the medium supply tube 113 - 1 , the medium recovery tube 113 - 2 , and the gas supply tube 113 - 3 is connected to the division medium supply unit, the medium recovery unit, and the gas mixing unit outside the housing.
  • the cells proliferated in the cell division canister 110 are recovered and re-seeded in the cell division canister 120 .
  • the cell differentiation canister 120 will be described with reference to FIGS. 3 and 6 .
  • the cell differentiation canister 120 includes a plurality of cell culture support fibers 121 aligned in one direction, an outer shell 122 surrounding the support fibers to form a space separated from the outside, and a multi-channel tube 123 connected to the outer shell. includes A plurality of cell differentiation canisters 120 may be included in the housing.
  • FIG. 3 it can be seen that a plurality of cell culture support fibers are aligned in one direction in the cell differentiation canister (A) and the overall structure of the canister (B).
  • the cell differentiation canister 120 performs a function of differentiating the seeded cells into muscle tissue. After supplying the cell differentiation medium to the medium supply tube 123-1, which is one of the multi-channel tubes 123, and seeding the cells on a plurality of cell culture support fibers 121, general cell culture conditions (37° C., 5% The cells can be cultured in CO 2 ). At this time, differentiation promoting factors such as insulin and dexamethasone may be added to the cell differentiation medium for efficient cell differentiation.
  • the cell differentiation canister 120 further includes a fixing device 125, for example, a fixing device such as a cap or a clip, that collects a plurality of cell culture support fibers 121 existing inside the outer shell so as not to separate them. can do.
  • a fixing device for example, a fixing device such as a cap or a clip
  • the cylindrical cell differentiation canister 120 can be connected to the multi-channel tube while fixing the cell culture support fiber 121 through the fixing device connected to the outer skin.
  • the fixing device may be in the form of a cap, for example, a structure that is detachable, such as a bottle cap.
  • the cell division canister 120 may further include a sensor 124 for measuring the glucose concentration and pH of the medium, and when the sensor detects a decrease in the glucose concentration and pH of the medium, the multi-channel tube 123 ), the old medium may be removed through the medium recovery tube 123-2, and a fresh cell differentiation medium may be supplied through the medium supply tube 123-1.
  • the cell division canister may use one of the multi-channel tubes 123 as the gas supply tube 123-3 to receive nitrogen, carbon dioxide, and oxygen necessary for cell culture.
  • the multi-channel tube 123 of the cell differentiation canister 120 includes a medium supply tube 123-1, a medium recovery tube 123-2, and a gas supply tube 123-3.
  • the medium supply tube 123-1 and the gas supply tube 123-3 may be connected to the fixing device 125 at the top of the cell differentiation canister 120, and the medium recovery tube 123- 2) may be connected to the fixing device 125 at the bottom of the cell differentiation canister 120 .
  • the cell differentiation canister 120 may further include a coil 124 for applying an electric field surrounding the canister, and the coil 124 for applying the electric field generates an electric field of 30 to 90 mV simulating an action potential of a muscle. It can be applied periodically to the cell differentiation canister. Periodically applying the electric field means applying the electric field of the above intensity for a predetermined time at regular time intervals.
  • the electric field application coil 124 is controlled by a microcontroller (MCU) of the cultured meat production system, and receives power.
  • MCU microcontroller
  • the cell differentiation canister 120 may contract and relax to promote cell differentiation, and for this, the cell culture apparatus includes a canister fixing end 130 .
  • An actuator 150 may be additionally included in order to move the fixed end of the canister.
  • the canister fixed end 130 is connected to both ends of the cell differentiation canister or the fixing device 125 to fix the canister, and can move up and down along the canister fixed end moving part 141 .
  • FIG. 5 shows the process of seeding cells in the cell differentiation canister and then differentiating them into muscle tissue through physical and electrical stimulation. indicates.
  • the cell culture support has a hydrogel structure including alginate and cellulose left after removing the cellular components from the medullary part of the seaweed, and the seaweed is seaweed, kelp, and seaweed.
  • the seaweed is seaweed, kelp, and seaweed.
  • the method for preparing the cell culture support includes a seaweed pretreatment step, an SDS treatment step, a cortex separation step, a PBS washing step, a gelation step, a freeze drying step, a sterilization step, and a manufacturing step.
  • the seaweed pretreatment step includes an ozone treatment step and a cutting step as a pretreatment step for inducing a decellularization process of seaweed individuals to be treated.
  • the pretreatment solution is a solution to properly swell the tissue of the seaweed, and it is produced by dissolving 1-3% (w/v) sodium chloride (NaCl) in distilled water, and preferably the concentration of sodium chloride is 1-2% (w/v). ) to make and use a pretreatment solution.
  • the ozone treatment step is a process of immersing a seaweed object in a pretreatment solution and performing ozone treatment to sterilize microorganisms present on the surface of the seaweed object. At this time, ozone is irradiated for about 1 minute to 1 minute and 30 seconds.
  • the cutting step is a process of cutting the seaweed individual into a desired size, and the shape and size of the cutting may vary depending on the purpose of the finally manufactured cell culture support.
  • the step of treating SDS is a process of weakening the cortex tissue of seaweed by immersing the cut seaweed in an aqueous SDS solution.
  • concentration of the SDS aqueous solution used is usually maintained in the range of 1 to 5% (w/v), and when immersing seaweed individuals, an SDS aqueous solution having a concentration of preferably 1 to 2% (w/v) is used. .
  • the cortex separation step is a process for separating the cortex layer of seaweed after treatment with an anionic detergent, and is carried out while slowly shaking the SDS solution in which the seaweed is immersed for 15 to 30 minutes. Through this process, the densely arranged cortical part and the medullar layer of the loose cellulose structure of the seaweed individual are separated from each other.
  • the phosphate buffered saline (PBS) washing step is a process of removing the cortex in the cortical part separation step and washing the remaining medulla with PBS
  • the gelation step is a process of gelling the remaining medulla after washing with PBS.
  • the immersion step is a process of immersing the water quality part in an aqueous solution of calcium chloride (CaCl2) in a concentration range of 0.5 to 10% (w/v) for about 1 to 10 minutes.
  • the treatment concentration and treatment time depend on the type of cell culture support and the type of cells to be cultured. can be changed. Through this process, Ca 2+ ions in aqueous solution are used for cross-linking of alginate, and the stiffness of the gel varies according to the degree of cross-linking.
  • the washing step is a process of taking out the water quality part after immersion and washing with PBS.
  • the water quality part becomes a transparent gel state. Since this gel has a unique network structure of seaweed, it is possible to make a sponge-type cell culture support sheet by freeze-drying the gel.
  • Cell culture support fibers can be prepared by cutting a freeze-dried support sheet to a predetermined length.
  • each shell is made of a polymer, and is a material that can contract and relax.
  • a polymer having a tensile strength of 700% or more and an elongation of 20 kN/m or more may be used. there is.
  • a medical polymer having excellent contractility and elongation ability may be used, and examples of the medical polymer are described in Table 1 below.
  • the housing 140 has a cell division canister and a cell differentiation canister positioned therein, and canister fixed end moving parts 141 are installed on both walls.
  • the housing 140 since the housing 140 has to maintain the inside of the housing in a constant environment for cell culture and differentiation, it is possible to create a closed environment with the outside through an openable and openable door.
  • the cell culture apparatus 100 may further include a controller 160 for controlling the temperature and carbon dioxide concentration inside the housing.
  • the cultured meat production system of the present invention includes a cell culture apparatus 100; At least one cell medium supply unit 200 connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture apparatus; a cell medium recovery unit 300 connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture apparatus; a gas mixing unit 400 connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture apparatus, and including a gas tank and a mixer; One or more peristaltic pumps 500 connected to the cell medium supply unit and the cell medium recovery unit; and a microcontroller 600 for controlling the operation of the cell culture device, the peristaltic pump, and the gas mixing unit.
  • the cell medium supply unit 200 is connected to the cell division canister and the multi-channel tubes 113 and 123 of the cell differentiation canister in the cell culture apparatus, and specifically is connected to the medium supply tubes 113-1 and 123-1. Meanwhile, the cell medium supply unit may include a division medium supply unit 210 and a differentiation medium supply unit 220 .
  • the cell medium recovery unit 300 is connected to the cell division canister and the multi-channel tubes 113 and 123 of the cell differentiation canister in the cell culture apparatus, and specifically is connected to the medium recovery tubes 113-2 and 123-2. .
  • the gas mixing unit 400 is connected to the cell division canister and the multi-channel tubes 113 and 123 of the cell differentiation canister in the cell culture apparatus, and specifically is connected to the gas supply tubes 113-3 and 123-3. do.
  • the gas mixing unit 400 includes a gas tank and a mixer.
  • a medium is supplied to the canister for cell division, and animal stem cells (eg, muscle stem cells) derived from mammals are seeded on the cell culture support. After the seeded cells are proliferated for a certain period of time to the target number of cells, the support itself after cell culture is completed is taken out of the canister.
  • animal stem cells eg, muscle stem cells
  • the cells proliferating on the cell culture support are manually separated.
  • the alginate component of the cell culture support is dissolved, and the cells are separated by treatment with trypsin.
  • the separated cells are seeded again in a canister for cell differentiation, and differentiation is induced for a certain period of time by applying physical, electrical, and chemical stimuli.
  • the differentiated cultured meat (cell culture support + cultured muscle tissue) deposited on the cell culture support by deposition from the cell differentiation canister is taken out, processed/cooked, and commercialized into cultured meat.

Abstract

The present invention relates to a cell culture apparatus for producing cultured meat and a cultured meat production system comprising the apparatus, wherein the cell culture apparatus uses an edible cell culture support derived from seaweed, and has the advantage of being able to apply physical and electrical stimulations required to differentiate cultured cells into muscle tissue.

Description

배양육 제조를 위한 세포 배양 장치Cell culture device for cultured meat production
본 발명은 배양육 제조를 위한 세포 배양 장치 및 상기 장치를 포함하는 배양육 제조 시스템에 관한 것이다.The present invention relates to a cell culture apparatus for producing cultured meat and a cultured meat production system including the apparatus.
2021년 6월 기준으로 세계 인구는 약 78억 7,500만 명으로 2030년에는 약 85억, 2050년에는 약 90억으로 예측되며, 세계 인구 증가에 따라 매년 2억톤 이상의 육류 단백질의 추가 공급이 필요해지고 있다. 이러한 상황에서 현재의 공장식 축산 방식은 증가하는 육류 단백질 수요를 충족시킬 수 없고, 공장식 축산 방식은 환경 오염과 동물 학대 논란에서 자유로울 수 없다. 이에 대체 고기에 대한 사람들의 관심이 높아지고 있다.As of June 2021, the world population is about 7.875 billion, which is expected to reach about 8.5 billion in 2030 and about 9 billion in 2050. there is. In this situation, the current factory farming method cannot meet the increasing demand for meat protein, and the factory farming method cannot be free from environmental pollution and controversy over animal cruelty. As a result, people's interest in alternative meat is increasing.
대체 고기(대체육, 인공육)는 콩, 밀, 쌀 등 식물성 재료로 고기의 맛이 나도록 만든 식물 고기와 동물의 세포를 배양해서 만든 배양 고기(배양육)로 나눌 수 있다. 배양육 (cultured meat) 은 살아있는 동물의 세포를 채취한 뒤 세포 공학 기술로 세포를 증식시켜 얻는 식용 고기를 의미하며, 가축을 사육하는 과정을 거치지 않고 고기를 얻는 세포농업 (cellular agriculture)의 한 분야라고 할 수 있다.Substitute meat (replacement meat, artificial meat) can be divided into plant meat made with plant materials such as soybeans, wheat, and rice to taste meat, and cultured meat (cultured meat) made by culturing animal cells. Cultured meat refers to edible meat obtained by harvesting cells from living animals and then proliferating the cells using cell engineering technology, a field of cellular agriculture that obtains meat without raising livestock it can be said
배양육 제조는 세포, 스캐폴드(scaffold), 배지와 성장인자, 생물반응기 (bioreactor) 등 다양한 공학적 사항을 고려해야 하는 복잡한 과정이다. 이중 생물반응기는 스캐폴드에 부착된 세포가 최종 결과물인 배양육으로 생장할 수 있도록 산소 및 배지를 공급하고, 노폐물을 제거한다. 일반적으로 회전생물반응기 (rotating bioreactor)가 많이 사용되나 생산하고자 하는 배양육의 종류에 따라 탱크 교반형, 정치 배양형 등 따라 다양한 생물반응기가 사용될 수 있다. 그러나 일반적인 고기의 맛과 질감을 갖는 배양육을 제조하는 기술은 아직 부족한 상황이다.Cultured meat production is a complex process that requires consideration of various engineering considerations, such as cells, scaffolds, media and growth factors, and bioreactors. The dual bioreactor supplies oxygen and medium so that the cells attached to the scaffold can grow into cultured meat, the final product, and removes wastes. In general, a rotating bioreactor is often used, but various bioreactors may be used depending on the type of cultured meat to be produced, such as a tank stirring type or a stationary culture type. However, the technology for producing cultured meat having the taste and texture of general meat is still lacking.
본 발명의 목적은 취식이 가능한 해조류 유래 세포배양 지지체 시트, 복수개의 세포배양 지지체 섬유를 이용한 세포 배양 장치, 세포 배양 장치를 이용한 배양육 제조 시스템을 제공하는 것이다.An object of the present invention is to provide an ingestible seaweed-derived cell culture support sheet, a cell culture apparatus using a plurality of cell culture support fibers, and a cultured meat production system using the cell culture apparatus.
상기 목적을 달성하기 위하여 본 발명의 일 양상은 세포 분열 캐니스터, 세포 분화 캐니스터, 캐니스터 고정단 및 하우징을 포함하는 세포 배양 장치를 제공한다.In order to achieve the above object, an aspect of the present invention provides a cell culture device including a cell division canister, a cell differentiation canister, a canister fixed end, and a housing.
상기 세포 분열 캐니스터는 다층의 세포배양 지지체 시트, 상기 지지체 시트를 감싸 외부와 분리된 공간을 형성하는 외피, 외피와 연결되는 다채널 튜브를 포함한다.The cell division canister includes a multi-layered cell culture support sheet, an outer shell surrounding the support sheet to form a space separated from the outside, and a multi-channel tube connected to the outer shell.
상기 세포 분화 캐니스터는 일 방향으로 정렬된 복수개의 세포배양 지지체 섬유, 상기 지지체 섬유를 감싸 외부와 분리된 공간을 형성하는 외피, 외피와 연결되는 다채널 튜브를 포함한다.The cell differentiation canister includes a plurality of cell culture support fibers aligned in one direction, an outer shell surrounding the support fibers to form a space separated from the outside, and a multi-channel tube connected to the outer shell.
또한, 상기 캐니스터 고정단은 세포 분화 캐니스터의 양 말단과 연결되어 세포 분화 캐니스터를 고정시키며, 캐니스터 고정단 이동부를 따라 상하로 이동할 수 있다.In addition, the fixed end of the canister is connected to both ends of the cell differentiation canister to fix the cell differentiation canister, and can move up and down along the moving part of the fixed end of the canister.
상기 하우징은 개폐가 가능하고, 내부에 세포 분열 캐니스터 및 세포 분화 캐니스터가 위치하며, 양 벽면에는 캐니스터 고정단 이동부가 설치된다.The housing can be opened and closed, and a cell division canister and a cell division canister are positioned therein, and fixed end moving parts of the canister are installed on both walls.
상기 세포 배양 장치는 배아줄기세포, 근육줄기세포, 중간엽 줄기세포, 유도만능 줄기세포, 근아세포 및 섬유아세포로 이루어진 군에서 선택되는 세포를 배양하며, 최종적으로 근육조직을 생성할 수 있다.The cell culture apparatus cultured cells selected from the group consisting of embryonic stem cells, muscle stem cells, mesenchymal stem cells, induced pluripotent stem cells, myoblasts and fibroblasts, and finally can produce muscle tissue.
상기 다채널 튜브는 배지 공급 튜브, 배지 회수 튜브 및 가스 공급 튜브를 포함할 수 있다.The multi-channel tube may include a medium supply tube, a medium recovery tube, and a gas supply tube.
상기 세포 분화 캐니스터는 캐니스터를 둘러싸고 있는 전기장 인가용 코일을 추가로 포함하며, 상기 전기장 인가용 코일은 근육의 활성 전위를 모사한 30 내지 90 mV의 전기장을 주기적으로 인가할 수 있다.The cell differentiation canister may further include a coil for applying an electric field surrounding the canister, and the coil for applying the electric field may periodically apply an electric field of 30 to 90 mV simulating an action potential of a muscle.
또한, 세포배양 지지체 시트 및 세포배양 지지체 섬유는 해조류 수질부에서 세포성분을 제거하고 남겨진 알지네이트와 셀룰로오스를 포함하는 하이드로젤 구조를 가지며, 상기 해조류는 미역, 다시마, 톳 중 적어도 어느 하나를 포함할 수 있다.In addition, the cell culture support sheet and the cell culture support fiber have a hydrogel structure containing alginate and cellulose left after removing the cell components from the medulla of seaweed, and the seaweed may include at least one of seaweed, kelp, and shiitake. there is.
한편, 세포 분열 캐니스터 및 세포 분화 캐니스터는 배지의 글루코스 농도 및 pH를 측정하는 센서를 추가로 포함할 수 있다.Meanwhile, the cell division canister and the cell differentiation canister may further include sensors for measuring the glucose concentration and pH of the medium.
상기 세포 배양 장치는 하우징 내 온도 및 이산화탄소 농도를 조절하는 컨트롤러 및/또는 캐니스터 고정단을 움직이는 액추에이터(actuator)를 추가로 포함할 수 있다. 이때, 캐니스터 고정단의 움직임에 의해 세포 분화 캐니스터가 수축 및 이완하며, 이 과정에서 캐니스터의 외피가 손상되지 않도록 상기 두 캐니스터의 외피는 수축 및 이완이 가능한 폴리머 재질로 인장 강도(tensile strength)는 700% 이상이고, 신장률(elongation)은 20 kN/m 이상일 수 있다.The cell culture apparatus may further include a controller that adjusts the temperature and carbon dioxide concentration in the housing and/or an actuator that moves the fixed end of the canister. At this time, the cell differentiation canister contracts and relaxes by the movement of the fixed end of the canister, and the outer shells of the two canisters are made of a polymer material that can be contracted and relaxed so as not to damage the outer shell of the canister in the process, and the tensile strength is 700 % or more, and the elongation may be 20 kN/m or more.
한편, 본 발명의 다른 양상은 하기 구성을 포함하는 배양육 제조 시스템을 제공한다:On the other hand, another aspect of the present invention provides a cultured meat production system comprising the following configuration:
상기 세포 배양 장치;the cell culture device;
세포 배양 장치의 다채널 튜브와 연결되는 하나 이상의 세포 배지 공급부;At least one cell medium supply unit connected to the multi-channel tube of the cell culture device;
상기 세포 배양 장치의 다채널 튜브와 연결되는 세포 배지 회수부;a cell medium recovery unit connected to the multi-channel tube of the cell culture device;
상기 세포 배양 장치의 다채널 튜브와 연결되고, 가스 탱크 및 믹서를 포함하는 가스 혼합부;a gas mixing unit connected to the multi-channel tube of the cell culture apparatus and including a gas tank and a mixer;
세포 배지 공급부 및 세포 배지 회수부와 연결되는 하나 이상의 연동 펌; 및one or more peristaltic pumps connected to the cell medium supply unit and the cell medium recovery unit; and
세포 배양 장치, 연동 펌프, 가스 혼합부의 작동을 조절하는 마이크로컨트롤러.A microcontroller that controls the operation of the cell culture device, peristaltic pump, and gas mixing section.
한편, 상기 세포 배지 공급부는 분열 배지 공급부 및 분화 배지 공급부를 포함할 수 있다.Meanwhile, the cell medium supply unit may include a division medium supply unit and a differentiation medium supply unit.
본 발명의 세포 배양 장치는 해조류에서 유래하여 취식이 가능한 세포배양 지지체를 사용하며, 배양하는 세포에 근육조직으로의 분화에 필요한 물리적 및 전기적 자극을 가할 수 있는 장점이 있다.The cell culture apparatus of the present invention uses a cell culture support derived from seaweed that can be ingested, and has the advantage of being able to apply physical and electrical stimulation necessary for differentiation into muscle tissue to cells to be cultured.
도 1은 세포 분열 캐니스터, 세포 분화 캐니스터, 캐니스터 고정단 및 하우징을 포함하는 세포 배양 장치의 구조를 개략적으로 나타낸다.1 schematically shows the structure of a cell culture apparatus including a cell division canister, a cell differentiation canister, a canister fixed end, and a housing.
도 2는 세포 분열 캐니스터에서 세포배양 지지체 시트가 다층을 이루고 있는 모습(A) 및 캐니스터의 전체적인 구조(B)를 나타낸다.2 shows a state in which a cell culture support sheet is formed in multiple layers in a cell division canister (A) and the overall structure of the canister (B).
도 3은 세포 분화 캐니스터에서 복수개의 세포배양 지지체 섬유가 일방향으로 정렬된 모습(A) 및 캐니스터의 전체적인 구조(B)를 나타낸다.3 shows a state in which a plurality of cell culture support fibers are aligned in one direction in a cell differentiation canister (A) and the overall structure of the canister (B).
도 4는 캐니스터 고정단에 연결된 세포 분화 캐니스터의 모습을 나타낸다.4 shows a view of a cell differentiation canister connected to a fixed end of the canister.
도 5는 세포 분화 캐니스터에 세포를 시딩한 후 물리적 및 전기적 자극을 통해 근육 조직으로 분화시키는 과정을 나타낸다.5 shows a process of seeding cells in a cell differentiation canister and then differentiating them into muscle tissue through physical and electrical stimulation.
도 6은 본 발명의 일 예에 따른 배양육 제조 시스템을 나타낸다.6 shows a cultured meat production system according to an example of the present invention.
이하, 첨부된 도면을 참고로 하여 본 발명의 실시예들에 대하여 본 발명이 속하는 기술분야에서 통상의 지식을 가진 자가 용이하게 실시할 수 있도록 상세히 설명한다. 본 발명은 여러 가지 상이한 형태로 구현될 수 있으며 여기에서 설명하는 실시예들에 한정되지 않는다.Hereinafter, with reference to the accompanying drawings, embodiments of the present invention will be described in detail so that those of ordinary skill in the art to which the present invention pertains can easily implement them. The present invention may be embodied in several different forms and is not limited to the embodiments described herein.
도면들은 개략적이고 축적에 맞게 도시되지 않았다는 것을 일러둔다. 도면에 있는 부분들의 상대적인 치수 및 비율은 도면에서의 명확성 및 편의를 위해 그 크기에 있어 과장되거나 감소되어 도시되었으며 임의의 치수는 단지 예시적인 것이지 한정적인 것은 아니다. 그리고 둘 이상의 도면에 나타나는 동일한 구조물, 요소 또는 부품에는 동일한 참조 부호가 유사한 특징을 나타내기 위해 사용된다.It is noted that the drawings are schematic and not drawn to scale. Relative dimensions and proportions of parts in the drawings are shown exaggerated or reduced in size for clarity and convenience in the drawings, and any dimensions are illustrative only and not limiting. And the same reference numerals are used to denote like features to the same structure, element, or part appearing in two or more drawings.
본 발명의 실시예는 본 발명의 이상적인 실시예들을 구체적으로 나타낸다. 그 결과, 도면의 다양한 변형이 예상된다. 따라서 실시예는 도시한 영역의 특정 형태에 국한되지 않으며, 예를 들면 제조에 의한 형태의 변형도 포함한다.The embodiments of the present invention specifically represent ideal embodiments of the present invention. As a result, various modifications of the drawings are expected. Accordingly, the embodiment is not limited to a specific shape of the illustrated area, and includes, for example, a shape modification by manufacturing.
이하, 첨부된 도면을 참조하여 본 발명의 일 실시예에 따른 세포 배양 장치(100)를 설명한다.Hereinafter, a cell culture apparatus 100 according to an embodiment of the present invention will be described with reference to the accompanying drawings.
먼저, 도 1을 참조하면 본 발명의 세포 배양 장치(100)는 세포 분열 캐니스터(110), 세포 분화 캐니스터(120), 캐니스터 고정단(130) 및 하우징(140)을 포함한다.First, referring to FIG. 1 , the cell culture apparatus 100 of the present invention includes a cell division canister 110 , a cell differentiation canister 120 , a canister fixing end 130 , and a housing 140 .
상기 세포 배양 장치(100)는 배아줄기세포, 근육줄기세포, 중간엽 줄기세포, 유도만능 줄기세포, 근아세포 및 섬유아세포로 이루어진 군에서 선택되는 세포의 배양에 사용될 수 있으며, 최종적으로는 상기 세포를 배양한 후 분화시켜 근육조직(배양육)을 형성할 수 있다.The cell culture apparatus 100 may be used for culturing cells selected from the group consisting of embryonic stem cells, muscle stem cells, mesenchymal stem cells, induced pluripotent stem cells, myoblasts and fibroblasts, and finally the cells After culturing, it can be differentiated to form muscle tissue (cultured meat).
도 2 및 도 6을 참고하여 세포 분열 캐니스터(110)를 설명한다.The cell division canister 110 will be described with reference to FIGS. 2 and 6 .
세포 분열 캐니스터(110)는 다층의 세포배양 지지체 시트(111), 상기 지지체 시트를 감싸 외부와 분리된 공간을 형성하는 외피(112), 외피와 연결되는 다채널 튜브(113)를 포함한다. 하우징 내에 세포 분열 캐니스터(110)는 복수개가 포함될 수 있다.The cell division canister 110 includes a multi-layered cell culture support sheet 111 , a shell 112 surrounding the support sheet to form a space separated from the outside, and a multi-channel tube 113 connected to the shell. A plurality of cell division canisters 110 may be included in the housing.
세포 분열 캐니스터(110)는 시딩(seeding)된 세포를 증식(proliferation)시키는 기능을 수행한다. 다채널 튜브(113)의 하나인 배지 공급 튜브(113-1)를 이용하여 세포 분열 배지를 공급하고, 다층의 세포배양 지지체 시트(111)에 세포를 시딩한 후 일반적인 세포 배양 조건(37℃, 5% CO2)에서 세포를 배양할 수 있다. 도 2에서 세포 분열 캐니스터의 세포배양 지지체 시트가 다층을 이루고 있는 모습(A) 및 전체적인 구조(B)를 확인할 수 있다.The cell division canister 110 performs a function of proliferating the seeded cells. Cell division medium is supplied using the medium supply tube 113-1, which is one of the multi-channel tubes 113, and the cells are seeded on the multi-layered cell culture support sheet 111, followed by general cell culture conditions (37° C., Cells can be cultured in 5% CO 2 ). In FIG. 2 , it can be seen that the cell culture support sheet of the cell division canister is multi-layered (A) and the overall structure (B).
한편, 상기 다층의 세포배양 지지체 시트(111)는 최소 1층 이상으로 사용될 수 있으며, 다층 구조가 유지될 수 있도록 시트의 양 말단에 클립형 고정 장치(111-1)를 포함할 수 있다.On the other hand, the multi-layered cell culture support sheet 111 may be used as at least one layer or more, and may include clip-type fixing devices 111-1 at both ends of the sheet so that the multi-layer structure can be maintained.
안정적인 세포 배양을 위해 세포 분열 캐니스터는 배지의 글루코스 농도 및 pH를 측정하는 센서(114)를 추가로 포함할 수 있으며, 센서에 배지의 글루코스 농도 및 pH 감소가 감지되면 다채널 튜브(113)의 하나인 배지 회수 튜브(113-2)로 오래된 배지를 제거하고, 배지 공급 튜브(113-1)로 신선한 세포 분열 배지를 공급할 수 있다.For stable cell culture, the cell division canister may further include a sensor 114 for measuring the glucose concentration and pH of the medium, and when the sensor detects a decrease in the glucose concentration and pH of the medium, one of the multi-channel tubes 113 The old medium may be removed through the phosphorus medium recovery tube 113-2, and the fresh cell division medium may be supplied through the medium supply tube 113-1.
또한, 세포 분열 캐니스터(110)는 세포 배양에 필요한 질소, 이산화탄소 및 산소를 공급 받을 수 있도록 다채널 튜브(113)의 하나를 가스 공급 튜브(113-3)으로 이용할 수 있다.In addition, the cell division canister 110 may use one of the multi-channel tubes 113 as the gas supply tube 113-3 to receive nitrogen, carbon dioxide, and oxygen necessary for cell culture.
따라서, 세포 분열 캐니스터(110)의 다채널 튜브(113)는 배지 공급 튜브(113-1), 배지 회수 튜브(113-2) 및 가스 공급 튜브(113-3)를 포함한다. 배지 공급 튜브(113-1), 배지 회수 튜브(113-2) 및 가스 공급 튜브(113-3) 각각은 하우징 외부의 분열 배지 공급부, 배지 회수부 및 가스 혼합부와 연결된다.Accordingly, the multi-channel tube 113 of the cell division canister 110 includes a medium supply tube 113-1, a medium recovery tube 113-2, and a gas supply tube 113-3. Each of the medium supply tube 113 - 1 , the medium recovery tube 113 - 2 , and the gas supply tube 113 - 3 is connected to the division medium supply unit, the medium recovery unit, and the gas mixing unit outside the housing.
세포 분열 캐니스터(110)에서 증식된 세포는 회수되어 세포 분화 캐니스터(120)에 다시 시딩된다.The cells proliferated in the cell division canister 110 are recovered and re-seeded in the cell division canister 120 .
도 3 및 도 6을 참고하여 세포 분화 캐니스터(120)를 설명한다.The cell differentiation canister 120 will be described with reference to FIGS. 3 and 6 .
세포 분화 캐니스터(120)는 일 방향으로 정렬된 복수개의 세포배양 지지체 섬유(121), 상기 지지체 섬유를 감싸 외부와 분리된 공간을 형성하는 외피(122), 외피와 연결되는 다채널 튜브(123)를 포함한다. 하우징 내에 세포 분화 캐니스터(120)는 복수개가 포함될 수 있다.The cell differentiation canister 120 includes a plurality of cell culture support fibers 121 aligned in one direction, an outer shell 122 surrounding the support fibers to form a space separated from the outside, and a multi-channel tube 123 connected to the outer shell. includes A plurality of cell differentiation canisters 120 may be included in the housing.
도 3에서 세포 분화 캐니스터에서 복수개의 세포배양 지지체 섬유가 일방향으로 정렬된 모습(A) 및 캐니스터의 전체적인 구조(B)를 확인할 수 있다.In FIG. 3 , it can be seen that a plurality of cell culture support fibers are aligned in one direction in the cell differentiation canister (A) and the overall structure of the canister (B).
세포 분화 캐니스터(120)는 시딩된 세포를 근육조직으로 분화 (differentiation)시키는 기능을 수행한다. 다채널 튜브(123)의 하나인 배지 공급 튜브(123-1)로 세포 분화 배지를 공급하고, 복수개의 세포배양 지지체 섬유(121)에 세포를 시딩한 후 일반적인 세포 배양 조건(37℃, 5% CO2)에서 세포를 배양할 수 있다. 이때 효율적인 세포 분화를 위해 세포 분화 배지에 인슐린, 덱사메타손(dexamethasone)과 같은 분화 촉진 인자를 첨가할 수 있다.The cell differentiation canister 120 performs a function of differentiating the seeded cells into muscle tissue. After supplying the cell differentiation medium to the medium supply tube 123-1, which is one of the multi-channel tubes 123, and seeding the cells on a plurality of cell culture support fibers 121, general cell culture conditions (37° C., 5% The cells can be cultured in CO 2 ). At this time, differentiation promoting factors such as insulin and dexamethasone may be added to the cell differentiation medium for efficient cell differentiation.
한편, 세포 분화 캐니스터(120)는 외피 내부에 존재하는 복수개의 세포배양 지지체 섬유(121) 가서로 분리되지 않도록 모아주는 고정 장치(125), 예를 들어 캡 또는 클립과 같은 고정 장치를 추가로 포함할 수 있다. 본 발명의 일 예에 따르면, 원통형의 세포 분화 캐니스터(120)는 외피와 연결된 상기 고정 장치를 통해 세포배양 지지체 섬유(121)를 고정시키면서 동시에 다채널 튜브와 연결될 수 있다. 상기 고정 장치는 캡(cap) 형태, 예를 들어 병 뚜껑과 같이 탈착이 가능한 구조일 수 있다.On the other hand, the cell differentiation canister 120 further includes a fixing device 125, for example, a fixing device such as a cap or a clip, that collects a plurality of cell culture support fibers 121 existing inside the outer shell so as not to separate them. can do. According to an example of the present invention, the cylindrical cell differentiation canister 120 can be connected to the multi-channel tube while fixing the cell culture support fiber 121 through the fixing device connected to the outer skin. The fixing device may be in the form of a cap, for example, a structure that is detachable, such as a bottle cap.
안정적인 세포 배양을 위해 세포 분열 캐니스터(120)는 배지의 글루코스 농도 및 pH를 측정하는 센서(124)를 추가로 포함할 수 있으며, 센서에 배지의 글루코스 농도 및 pH 감소가 감지되면 다채널 튜브(123)의 하나인 배지 회수 튜브(123-2)로 오래된 배지를 제거하고, 배지 공급 튜브(123-1)로 신선한 세포 분화 배지를 공급할 수 있다.For stable cell culture, the cell division canister 120 may further include a sensor 124 for measuring the glucose concentration and pH of the medium, and when the sensor detects a decrease in the glucose concentration and pH of the medium, the multi-channel tube 123 ), the old medium may be removed through the medium recovery tube 123-2, and a fresh cell differentiation medium may be supplied through the medium supply tube 123-1.
또한, 세포 분열 캐니스터는 세포 배양에 필요한 질소, 이산화탄소 및 산소를 공급 받을 수 있도록 다채널 튜브(123)의 하나를 가스 공급 튜브(123-3)으로 이용할 수 있다.In addition, the cell division canister may use one of the multi-channel tubes 123 as the gas supply tube 123-3 to receive nitrogen, carbon dioxide, and oxygen necessary for cell culture.
따라서, 세포 분화 캐니스터(120)의 다채널 튜브(123)는 배지 공급 튜브(123-1), 배지 회수 튜브(123-2) 및 가스 공급 튜브(123-3)를 포함한다. 본 발명의 일 예에 따르면, 배지 공급 튜브(123-1)와 가스 공급 튜브(123-3)는 세포 분화 캐니스터(120) 상단의 고정장치(125)와 연결될 수 있고, 배지 회수 튜브(123-2)는 세포 분화 캐니스터(120) 하단의 고정장치(125)와 연결될 수 있다.Accordingly, the multi-channel tube 123 of the cell differentiation canister 120 includes a medium supply tube 123-1, a medium recovery tube 123-2, and a gas supply tube 123-3. According to an example of the present invention, the medium supply tube 123-1 and the gas supply tube 123-3 may be connected to the fixing device 125 at the top of the cell differentiation canister 120, and the medium recovery tube 123- 2) may be connected to the fixing device 125 at the bottom of the cell differentiation canister 120 .
또한, 세포 분화 캐니스터(120)는 캐니스터를 둘러싸고 있는 전기장 인가용 코일(124)을 추가로 포함할 수 있으며, 전기장 인가용 코일(124)은 근육의 활성 전위를 모사한 30 내지 90 mV의 전기장을 세포 분화 캐니스터에 주기적으로 인가할 수 있다. 전기장을 주기적으로 인가한다는 것은 일정 시간 간격으로 일정 시간 동안 상기 세기의 전기장을 인가하는 것을 의미한다.In addition, the cell differentiation canister 120 may further include a coil 124 for applying an electric field surrounding the canister, and the coil 124 for applying the electric field generates an electric field of 30 to 90 mV simulating an action potential of a muscle. It can be applied periodically to the cell differentiation canister. Periodically applying the electric field means applying the electric field of the above intensity for a predetermined time at regular time intervals.
전기장 인가용 코일(124)은 배양육 제조 시스템의 마이크로컨트롤러(MCU)에 의해 제어되고, 전원을 공급 받는다.The electric field application coil 124 is controlled by a microcontroller (MCU) of the cultured meat production system, and receives power.
한편, 세포 분화 캐니스터(120)는 세포 분화를 촉진하기 위해 수축 및 이완될 수 있으며, 이를 위해 세포 배양 장치는 캐니스터 고정단(130)를 포함한다. 캐니스터 고정단의 이동을 위해서는 액추에이터(150)를 추가로 포함할 수 있다.Meanwhile, the cell differentiation canister 120 may contract and relax to promote cell differentiation, and for this, the cell culture apparatus includes a canister fixing end 130 . An actuator 150 may be additionally included in order to move the fixed end of the canister.
캐니스터 고정단(130)은 세포 분화 캐니스터의 양 말단 또는 고정장치(125)와 연결되어 캐니스터를 고정시키며, 캐니스터 고정단 이동부(141)를 따라 상하로 이동할 수 있다.The canister fixed end 130 is connected to both ends of the cell differentiation canister or the fixing device 125 to fix the canister, and can move up and down along the canister fixed end moving part 141 .
도 1 및 도 4에서 캐니스터 고정단(130)에 연결된 세포 분화 캐니스터(120)를 확인할 수 있으며, 도 5는 세포 분화 캐니스터에 세포를 시딩한 후 물리적 및 전기적 자극을 통해 근육 조직으로 분화시키는 과정을 나타낸다.1 and 4, the cell differentiation canister 120 connected to the canister fixing end 130 can be seen, and FIG. 5 shows the process of seeding cells in the cell differentiation canister and then differentiating them into muscle tissue through physical and electrical stimulation. indicates.
한편, 세포 배양 캐니스터(110) 및 세포 분화 캐니스터(120)에서 세포배양 지지체는 해조류 수질부에서 세포성분을 제거하고 남겨진 알지네이트와 셀룰로오스를 포함하는 하이드로젤 구조를 가지고, 상기 해조류는 미역, 다시마, 톳 중 적어도 어느 하나를 포함할 수 있다.On the other hand, in the cell culture canister 110 and the cell differentiation canister 120, the cell culture support has a hydrogel structure including alginate and cellulose left after removing the cellular components from the medullary part of the seaweed, and the seaweed is seaweed, kelp, and seaweed. may include at least one of
구체적으로 세포 배양 지지체의 제조 방법은 해조류 전처리 단계, SDS 처리 단계, 피질부 분리 단계, PBS 워싱 단계, 젤화 단계, 동결 건조 단계, 멸균 단계, 제작 단계를 포함한다.Specifically, the method for preparing the cell culture support includes a seaweed pretreatment step, an SDS treatment step, a cortex separation step, a PBS washing step, a gelation step, a freeze drying step, a sterilization step, and a manufacturing step.
해조류 전처리 단계는 처리 대상인 미역 개체의 탈세포화 공정을 유도하기 위한 전처리 공정으로서 오존 처리 단계 및 절단 단계를 포함한다. 전처리 용액은 해조류 개체의 조직을 적절히 팽윤시키기 위한 용액으로 1~3%(w/v)의 염화나트륨(NaCl)을 증류수에 녹여 생성하며, 바람직하게는 염화나트륨의 농도가 1~2%(w/v)인 전처리 용액을 만들어 사용한다. 오존 처리 단계는 해조류 개체를 전처리 용액에 침지하고, 해조류 개체 표면에 존재하는 미생물을 살균하기 위해 오존 처리를 하는 공정이다. 이때, 오존은 1분~1분 30초 가량 조사된다.The seaweed pretreatment step includes an ozone treatment step and a cutting step as a pretreatment step for inducing a decellularization process of seaweed individuals to be treated. The pretreatment solution is a solution to properly swell the tissue of the seaweed, and it is produced by dissolving 1-3% (w/v) sodium chloride (NaCl) in distilled water, and preferably the concentration of sodium chloride is 1-2% (w/v). ) to make and use a pretreatment solution. The ozone treatment step is a process of immersing a seaweed object in a pretreatment solution and performing ozone treatment to sterilize microorganisms present on the surface of the seaweed object. At this time, ozone is irradiated for about 1 minute to 1 minute and 30 seconds.
절단 단계는 해조류 개체를 목적하는 크기로 절단하는 공정이며, 절단하는 모양 및 크기는 최종적으로 제조되는 세포 배양 지지체의 목적에 따라 달라질 수 있다.The cutting step is a process of cutting the seaweed individual into a desired size, and the shape and size of the cutting may vary depending on the purpose of the finally manufactured cell culture support.
음이온성 세정제인 SDS(Sodium Dodecyl Sulfate) 처리 단계는 절단된 해조류 개체를 SDS 수용액에 침지하여 해조류의 피질(cortex) 조직을 약화시키는 공정이다. 사용하는 SDS 수용액의 농도는 통상적으로 1~5%(w/v) 범위를 유지하며, 미역 개체를 침지하는 경우에는 바람직하게는 1~2%(w/v)의 농도인 SDS 수용액을 사용한다. The step of treating SDS (Sodium Dodecyl Sulfate), an anionic detergent, is a process of weakening the cortex tissue of seaweed by immersing the cut seaweed in an aqueous SDS solution. The concentration of the SDS aqueous solution used is usually maintained in the range of 1 to 5% (w/v), and when immersing seaweed individuals, an SDS aqueous solution having a concentration of preferably 1 to 2% (w/v) is used. .
피질부 분리 단계는 음이온성 세정제로 처리 후 미역 개체의 피질부(cortex layer)를 분리하기 위한 공정이며, 해조류 개체가 침지된 SDS 수용액을 15~30분간 천천히 흔들면서 진행된다. 이러한 과정을 통해 해조류 개체에 치밀하게 배열된 피질부와 느슨한 셀룰로오스 구조의 수질부(medullar layer)가 서로 분리된다.The cortex separation step is a process for separating the cortex layer of seaweed after treatment with an anionic detergent, and is carried out while slowly shaking the SDS solution in which the seaweed is immersed for 15 to 30 minutes. Through this process, the densely arranged cortical part and the medullar layer of the loose cellulose structure of the seaweed individual are separated from each other.
PBS(phosphate buffered saline) 세척 단계는 피질부 분리 단계에서 피질부를 제거하고 남은 수질부를 PBS로 세척하는 공정이며, 젤화 단계는 PBS로 세척하고 남은 수질부를 젤화시키는 공정으로 침지 단계 및 세척 단계를 포함한다.The phosphate buffered saline (PBS) washing step is a process of removing the cortex in the cortical part separation step and washing the remaining medulla with PBS, and the gelation step is a process of gelling the remaining medulla after washing with PBS. .
침지 단계는 수질부를 0.5~10%(w/v) 농도 범위의 염화칼슘(CaCl2) 수용액에 1~10분 정도 침지하는 공정으로 세포 배양 지지체의 종류 및 배양되는 세포의 종류에 따라 처리 농도와 처리 시간은 변경할 수 있다. 이 과정을 통해 수용액 상의 Ca2+ 이온이 알지네이트의 가교(cross-linking)에 사용되며, 가교 정도에 따라 젤의 강성도(stiffness)가 달라지게 된다. 이어서, 세척 단계는 침지 후 수질부를 꺼내어 PBS로 세척하는 공정이다.The immersion step is a process of immersing the water quality part in an aqueous solution of calcium chloride (CaCl2) in a concentration range of 0.5 to 10% (w/v) for about 1 to 10 minutes. The treatment concentration and treatment time depend on the type of cell culture support and the type of cells to be cultured. can be changed. Through this process, Ca 2+ ions in aqueous solution are used for cross-linking of alginate, and the stiffness of the gel varies according to the degree of cross-linking. Then, the washing step is a process of taking out the water quality part after immersion and washing with PBS.
상기 침지 단계와 세척 단계 이후 수질부는 투명한 젤 상태가 된다. 이 젤은 해조류 고유의 망 구조를 갖기 때문에 이 젤을 동결건조시키면 스펀지 형태의 세포배양 지지체 시트를 만들 수 있다. 세포배양 지지체 섬유는 동결건조시킨 지지체 시트를 일정 길이로 절단하여 제조할 수 있다.After the immersion step and the washing step, the water quality part becomes a transparent gel state. Since this gel has a unique network structure of seaweed, it is possible to make a sponge-type cell culture support sheet by freeze-drying the gel. Cell culture support fibers can be prepared by cutting a freeze-dried support sheet to a predetermined length.
상기 세포 분열 및 세포 분화 캐니스터에서 각각의 외피는 폴리머로 이루어지며, 수축 및 이완이 가능한 재질로 인장 강도(tensile strength)는 700% 이상이고, 신장률(elongation)은 20 kN/m 이상인 폴리머가 사용될 수 있다.In the cell division and cell differentiation canister, each shell is made of a polymer, and is a material that can contract and relax. A polymer having a tensile strength of 700% or more and an elongation of 20 kN/m or more may be used. there is.
구체적으로는 수축 및 신장 능력이 우수한 의료용 폴리머가 사용될 수 있으며, 아래 표 1에 의료용 폴리머의 예시를 기재하였다.Specifically, a medical polymer having excellent contractility and elongation ability may be used, and examples of the medical polymer are described in Table 1 below.
Avantor (제조사)Avantor (manufacturer) MED-6400MED-6400 MED-6600MED-6600
Tensile StrengthTensile Strength 850%850% 800%800%
Elongationelongation 160 ppi (28.2 kN/m)160 ppi (28.2 kN/m) 160 ppi (28.2 kN/m)160 ppi (28.2 kN/m)
Tissue Culture (Cytotoxicity Testing)Tissue Culture (Cytotoxicity Testing) USP ISO 10993-5USP ISO 10993-5
Elemental Analysis of Trace MetalsElemental Analysis of Trace Metals ASTM E305ASTM E305
하우징(140)은 내부에 세포 분열 캐니스터 및 세포 분화 캐니스터가 위치하고, 양 벽면에는 캐니스터 고정단 이동부(141)가 설치된다. 또한, 하우징 (140)은 세포 배양 및 분화를 위해 하우징 내부를 일정한 환경으로 유지해야 하므로 개폐가 가능한 문을 통해 외부와 밀폐된 환경을 조성할 수 있다.The housing 140 has a cell division canister and a cell differentiation canister positioned therein, and canister fixed end moving parts 141 are installed on both walls. In addition, since the housing 140 has to maintain the inside of the housing in a constant environment for cell culture and differentiation, it is possible to create a closed environment with the outside through an openable and openable door.
한편, 하우징 내부의 환경 조절을 위해 세포 배양 장치(100)는 하우징 내부의 온도 및 이산화탄소 농도를 조절하는 컨트롤러(160)를 추가로 포함할 수 있다.On the other hand, to control the environment inside the housing, the cell culture apparatus 100 may further include a controller 160 for controlling the temperature and carbon dioxide concentration inside the housing.
하우징(140) 내부의 양 벽면에는 벽면에서 돌출된 캐니스터 고정단 이동부(141)가 설치되어 캐니스터 고정단이 상하로 이동할 수 있다. 도 1에서 하우징 내부의 양쪽 벽면에 형성된 캐니스터 고정단 이동부(141)를 확인할 수 있다.On both walls of the housing 140 , fixed end moving parts 141 protruding from the wall are installed, so that the fixed end of the canister can move up and down. In FIG. 1 , it can be seen that the canister fixed end moving parts 141 formed on both wall surfaces of the inside of the housing.
한편, 배양육 제조를 위해서는 상기 세포 배양 장치 이외에 추가 장치들이 필요하다. 이하, 도 6을 참조하여 배양육 제조 시스템을 설명한다.On the other hand, additional devices are required in addition to the cell culture device for the production of cultured meat. Hereinafter, a cultured meat production system will be described with reference to FIG. 6 .
본 발명의 배양육 제조 시스템은 세포 배양 장치(100); 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브와 연결되는 하나 이상의 세포 배지 공급부(200); 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브와 연결되는 세포 배지 회수부(300); 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브와 연결되고, 가스 탱크 및 믹서를 포함하는 가스 혼합부(400); 세포 배지 공급부 및 세포 배지 회수부와 연결되는 하나 이상의 연동 펌프(500); 및 세포 배양 장치, 연동 펌프, 가스 혼합부의 작동을 조절하는 마이크로컨트롤러(600)를 포함한다.The cultured meat production system of the present invention includes a cell culture apparatus 100; At least one cell medium supply unit 200 connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture apparatus; a cell medium recovery unit 300 connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture apparatus; a gas mixing unit 400 connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture apparatus, and including a gas tank and a mixer; One or more peristaltic pumps 500 connected to the cell medium supply unit and the cell medium recovery unit; and a microcontroller 600 for controlling the operation of the cell culture device, the peristaltic pump, and the gas mixing unit.
세포 배지 공급부(200)는 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브(113 및 123)와 연결되며, 구체적으로는 배지 공급 튜브(113-1 및 123-1)와 연결된다. 한편, 상기 세포 배지 공급부는 분열 배지 공급부(210) 및 분화 배지 공급부(220)를 포함할 수 있다.The cell medium supply unit 200 is connected to the cell division canister and the multi-channel tubes 113 and 123 of the cell differentiation canister in the cell culture apparatus, and specifically is connected to the medium supply tubes 113-1 and 123-1. Meanwhile, the cell medium supply unit may include a division medium supply unit 210 and a differentiation medium supply unit 220 .
세포 배지 회수부(300)는 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브(113 및 123)와 연결되며, 구체적으로는 배지 회수 튜브(113-2 및 123-2)와 연결된다.The cell medium recovery unit 300 is connected to the cell division canister and the multi-channel tubes 113 and 123 of the cell differentiation canister in the cell culture apparatus, and specifically is connected to the medium recovery tubes 113-2 and 123-2. .
또한, 가스 혼합부(400)는 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브(113 및 123)와 연결되며, 구체적으로는 가스 공급 튜브(113-3 및 123-3)와 연결된다. 또한, 가스 혼합부(400)는 가스 탱크 및 믹서를 포함한다.In addition, the gas mixing unit 400 is connected to the cell division canister and the multi-channel tubes 113 and 123 of the cell differentiation canister in the cell culture apparatus, and specifically is connected to the gas supply tubes 113-3 and 123-3. do. Also, the gas mixing unit 400 includes a gas tank and a mixer.
본 발명의 배양육 제조 시스템으로 배양육을 제조하는 과정을 간략하게 설명한다.A process for producing cultured meat with the cultured meat production system of the present invention will be briefly described.
세포 분열용 캐니스터에 배지를 공급하고, 포유류로부터 유래된 동물 줄기세포 (예를 들어, 근육 줄기세포)를 세포배양 지지체에 시딩한다. 시딩한 세포를 목표로 하는 세포 수까지 일정 기간 증식시킨 후 세포 배양이 완료된 지지체 자체를 캐니스터에서 꺼낸다.A medium is supplied to the canister for cell division, and animal stem cells (eg, muscle stem cells) derived from mammals are seeded on the cell culture support. After the seeded cells are proliferated for a certain period of time to the target number of cells, the support itself after cell culture is completed is taken out of the canister.
이후 세포배양 지지체에 증식하고 있는 세포들을 매뉴얼로 분리한다. 예를 들어, 세포배양 지지체의 알지네이트 성분을 녹이고, 트립신을 처리하여 세포를 분리한다. 분리한 세포를 세포 분화용 캐니스터에 다시 시딩하고, 물리적, 전기적 및 화학적 자극을 인가하여 일정 기간 동안 분화를 유도한다. 시간이 흐른 후 세포 분화용 캐니스터로부터 세포 배양 지지체에 증착하여 분화한 배양육 분화체(세포 배양 지지체+배양된 근육조직)를 꺼내고, 가공/조리하여 배양육으로 제품화시킨다.Thereafter, the cells proliferating on the cell culture support are manually separated. For example, the alginate component of the cell culture support is dissolved, and the cells are separated by treatment with trypsin. The separated cells are seeded again in a canister for cell differentiation, and differentiation is induced for a certain period of time by applying physical, electrical, and chemical stimuli. After time has elapsed, the differentiated cultured meat (cell culture support + cultured muscle tissue) deposited on the cell culture support by deposition from the cell differentiation canister is taken out, processed/cooked, and commercialized into cultured meat.

Claims (12)

  1. 세포 분열 캐니스터, 세포 분화 캐니스터, 캐니스터 고정단 및 하우징을 포함하는 세포 배양 장치로서,A cell culture device comprising a cell division canister, a cell differentiation canister, a canister fixed end, and a housing, comprising:
    상기 세포 분열 캐니스터는 다층의 세포배양 지지체 시트, 상기 지지체 시트를 감싸 외부와 분리된 공간을 형성하는 외피, 외피와 연결되는 다채널 튜브를 포함하고,The cell division canister includes a multi-layered cell culture support sheet, an outer shell surrounding the support sheet to form a space separated from the outside, and a multi-channel tube connected to the outer shell,
    상기 세포 분화 캐니스터는 일 방향으로 정렬된 복수개의 세포배양 지지체 섬유, 상기 지지체 섬유를 감싸 외부와 분리된 공간을 형성하는 외피, 외피와 연결되는 다채널 튜브를 포함하며,The cell differentiation canister includes a plurality of cell culture support fibers aligned in one direction, an outer shell surrounding the support fibers to form a space separated from the outside, and a multi-channel tube connected to the outer shell,
    상기 캐니스터 고정단은 세포 분화 캐니스터의 양 말단과 연결되어 세포 분화 캐니스터를 고정시키며, 캐니스터 고정단 이동부를 따라 상하로 이동가능하고,The fixed end of the canister is connected to both ends of the cell differentiation canister to fix the cell differentiation canister, and is movable up and down along the moving part of the fixed end of the canister;
    상기 하우징은 개폐가 가능하고, 내부에 세포 분열 캐니스터 및 세포 분화 캐니스터가 위치하며, 내부의 양 벽면에는 캐니스터 고정단 이동부가 설치되는, 세포 배양 장치.The housing can be opened and closed, a cell division canister and a cell differentiation canister are positioned therein, and a canister fixed end moving part is installed on both inner walls.
  2. 제1항에 있어서, 상기 세포 배양 장치는 배아줄기세포, 근육줄기세포, 중간엽 줄기세포, 유도만능줄기세포, 근아세포 및 섬유아세포로 이루어진 군에서 선택되는 세포를 배양하는 것인, 세포 배양 장치.The cell culture apparatus of claim 1, wherein the cell culture apparatus is to culture cells selected from the group consisting of embryonic stem cells, muscle stem cells, mesenchymal stem cells, induced pluripotent stem cells, myoblasts and fibroblasts. .
  3. 제1항에 있어서, 상기 세포 분화 캐니스터는 캐니스터를 둘러싸고 있는 전기장 인가용 코일을 추가로 포함하는, 세포 배양 장치.The cell culture apparatus of claim 1 , wherein the cell differentiation canister further comprises a coil for applying an electric field surrounding the canister.
  4. 제3항에 있어서, 상기 전기장 인가용 코일은 30 내지 90 mV의 전기장을 주기적으로 인가하는, 세포 배양 장치.The cell culture apparatus of claim 3, wherein the electric field application coil periodically applies an electric field of 30 to 90 mV.
  5. 제1항에 있어서, 상기 세포배양 지지체는 해조류 수질부에서 세포성분을 제거하고 남겨진 알지네이트와 셀룰로오스를 포함하는 하이드로젤 구조를 가지며, 상기 해조류는 미역, 다시마, 톳 중 적어도 어느 하나를 포함하는, 세포 배양 장치.The cell according to claim 1, wherein the cell culture support has a hydrogel structure including alginate and cellulose left after removing the cell components from the medulla of seaweed, and the seaweed contains at least one of seaweed, kelp, and shiitake. incubation device.
  6. 제1항에 있어서, 상기 외피는 인장 강도(tensile strength)는 700% 이상이고, 신장률(elongation)은 20 kN/m 이상인 폴리머인, 세포 배양 장치.The cell culture apparatus according to claim 1, wherein the sheath is a polymer having a tensile strength of 700% or more and an elongation of 20 kN/m or more.
  7. 제1항에 있어서, 상기 다채널 튜브는 배지 공급 튜브, 배지 회수 튜브 및 가스 공급 튜브를 포함하는 것인, 세포 배양 장치.The cell culture apparatus according to claim 1, wherein the multi-channel tube includes a medium supply tube, a medium recovery tube and a gas supply tube.
  8. 제1항에 있어서, 상기 세포 분열 캐니스터 및 세포 분화 캐니스터는 배지의 글루코스 농도 및 pH를 측정하는 센서를 추가로 포함하는 것인, 세포 배양 장치.The cell culture apparatus according to claim 1, wherein the cell division canister and the cell differentiation canister further include a sensor for measuring the glucose concentration and pH of the medium.
  9. 제1항에 있어서, 상기 세포 배양 장치는 하우징 내 온도 및 이산화탄소 농도를 조절하는 컨트롤러를 추가로 포함하는 것인, 세포 배양 장치.The cell culture apparatus according to claim 1, wherein the cell culture apparatus further comprises a controller for controlling the temperature and carbon dioxide concentration in the housing.
  10. 제1항에 있어서, 상기 세포 배양 장치는 캐니스터 고정단을 움직이는 액추에이터(actuator)를 추가로 포함하는, 세포 배양 장치.According to claim 1, wherein the cell culture apparatus further comprises an actuator (actuator) for moving the canister fixed end, the cell culture apparatus.
  11. 제1항의 세포 배양 장치;The cell culture apparatus of claim 1;
    상기 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브와 연결되는 하나 이상의 세포 배지 공급부;At least one cell medium supply unit connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture device;
    상기 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브와 연결되는 세포 배지 회수부;a cell medium recovery unit connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture device;
    상기 세포 배양 장치에서 세포 분열 캐니스터 및 세포 분화 캐니스터의 다채널 튜브와 연결되고, 가스 탱크 및 믹서를 포함하는 가스 혼합부;a gas mixing unit connected to the cell division canister and the multi-channel tube of the cell differentiation canister in the cell culture device, and including a gas tank and a mixer;
    세포 배지 공급부 및 세포 배지 회수부와 연결되는 하나 이상의 연동 펌프; 및one or more peristaltic pumps connected to the cell medium supply unit and the cell medium recovery unit; and
    세포 배양 장치, 연동 펌프, 가스 혼합부의 작동을 조절하는 마이크로컨트롤러를 포함하는, 배양육 제조 시스템.A cultured meat production system comprising a cell culture device, a peristaltic pump, and a microcontroller for controlling the operation of the gas mixing unit.
  12. 제10항에 있어서, 상기 세포 배지 공급부는 분열 배지 공급부 및 분화 배지 공급부를 포함하는 것인, 배양육 제조 시스템.The system according to claim 10, wherein the cell medium supply unit includes a division medium supply unit and a differentiation medium supply unit.
PCT/KR2021/011613 2020-08-28 2021-08-30 Cell culture apparatus for producing cultured meat WO2022045854A1 (en)

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