WO2022009560A1 - 細胞培養システム - Google Patents

細胞培養システム Download PDF

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Publication number
WO2022009560A1
WO2022009560A1 PCT/JP2021/020605 JP2021020605W WO2022009560A1 WO 2022009560 A1 WO2022009560 A1 WO 2022009560A1 JP 2021020605 W JP2021020605 W JP 2021020605W WO 2022009560 A1 WO2022009560 A1 WO 2022009560A1
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WO
WIPO (PCT)
Prior art keywords
cell culture
medium
anaerobic chamber
operation unit
main body
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/JP2021/020605
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English (en)
French (fr)
Japanese (ja)
Inventor
恭子 米田
智樹 大久保
豊之 橋本
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Shimadzu Corp
Original Assignee
Shimadzu Corp
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Shimadzu Corp filed Critical Shimadzu Corp
Priority to JP2022534949A priority Critical patent/JPWO2022009560A1/ja
Priority to US18/014,866 priority patent/US20230287326A1/en
Priority to EP21837468.4A priority patent/EP4180511A4/en
Priority to CN202180048286.9A priority patent/CN115916950A/zh
Publication of WO2022009560A1 publication Critical patent/WO2022009560A1/ja
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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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
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/48—Automatic or computerized control
    • 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
    • C12N1/00—Microorganisms; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • 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
    • C12M23/24—Gas permeable parts
    • 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
    • C12M23/34—Internal compartments or partitions
    • 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
    • C12M25/00—Means for supporting, enclosing or fixing the microorganisms, e.g. immunocoatings
    • C12M25/02—Membranes; Filters
    • C12M25/04—Membranes; Filters in combination with well or multiwell plates, i.e. culture inserts
    • 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
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C12M41/34—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of gas
    • 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
    • C12N2500/00—Specific components of cell culture medium
    • C12N2500/02—Atmosphere, e.g. low oxygen conditions
    • 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
    • C12N2529/00—Culture process characterised by the use of electromagnetic stimulation

Definitions

  • the present invention relates to a cell culture system.
  • Patent Document 1 International Publication No. 2018/07793
  • a device in which intestinal epithelial cells are seeded on a porous membrane is placed in an anaerobic chamber, and the intestinal epithelial cells and bacteria contained in the medium are co-cultured.
  • the system to be used is disclosed.
  • the external device required for these treatments is also anaerobic. It will be placed in the chamber. Since the operation of this external device is performed by inserting a hand into the anaerobic chamber from a glove or sleeve provided on the front surface of the anaerobic chamber, the operability is low.
  • the external device may be separated from the main body located inside the anaerobic chamber and the operation unit arranged outside the anaerobic chamber, and the main body and the operation unit may be connected via a cable. Conceivable. In this case, the operability of the external device is improved, but as a result of providing a through hole for passing the cable in the anaerobic chamber, a separate measure for ensuring the airtightness of the anaerobic chamber around the through hole is required. ..
  • the present invention provides a cell culture system capable of enhancing the operability of an external device without providing a through hole in the anaerobic chamber.
  • the cell culture system of the present invention includes a cell culture container arranged in an anaerobic chamber and an external device.
  • the external device has a main body portion arranged in the anaerobic chamber and an operation unit arranged outside the anaerobic chamber.
  • the main body is configured to perform a predetermined treatment on the cell culture container.
  • the operation unit is configured to transmit or receive a signal related to a predetermined process by wireless communication with the main body unit.
  • the cell culture system of the present invention it is possible to improve the operability of the external device without providing a through hole in the anaerobic chamber.
  • cell culture system 100 Structure of cell culture system according to embodiment
  • cell culture system 100 the configuration of the cell culture system (hereinafter referred to as “cell culture system 100”) according to the embodiment will be described.
  • FIG. 1 is a schematic diagram of the cell culture system 100.
  • the cell culture system 100 includes a cell culture container 10, a tube 20a and a tube 20b, a pump 30, an electric resistance measuring device 40, a dissolved oxygen concentration measuring device 50, and an operation unit 60.
  • the pump 30, the electric resistance measuring device 40, the dissolved oxygen concentration measuring device 50, and the operating unit 60 are external devices of the cell culture system 100.
  • the pump 30, the electric resistance measuring device 40, and the dissolved oxygen concentration measuring device 50 are the main bodies of the external device.
  • the cell culture container 10 has a container body 11, a cell culture insert 12, a lid member 13, and a sealing member 14.
  • the container body 11 has a side wall 11a, a bottom wall 11b, and a partition wall 11c.
  • the bottom wall 11b closes the lower end side of the side wall 11a.
  • the upper end side of the side wall 11a is open. This opening is closed by the lid member 13.
  • the partition wall 11c is connected to the side wall 11a between the upper end of the side wall 11a and the lower end of the side wall 11a.
  • the partition wall 11c extends parallel to, for example, the bottom wall 11b.
  • An opening 11ca is formed in the partition wall 11c. The opening 11ca penetrates the partition wall 11c along the thickness direction.
  • the cell culture insert 12 has a tubular portion 12a, a bottom portion 12b, and a flange portion 12c.
  • the tubular portion 12a has a tubular shape. The upper end of the tubular portion 12a is open. The lower end of the tubular portion 12a is closed by the bottom portion 12b.
  • the bottom portion 12b is a membrane 12e.
  • the membrane 12e is oxygen permeable.
  • the membrane 12e is, for example, a track etch film made of polycarbonate.
  • the membrane 12e has a first main surface 12ea and a second main surface 12eb.
  • the second main surface 12eb is the opposite surface of the first main surface 12ea.
  • the tubular portion 12a is inserted into the opening 11ca so that the bottom portion 12b (membrane 12e) is located between the bottom wall 11b and the partition wall 11c.
  • the space defined by the outer peripheral surface of the side wall 11a between the bottom wall 11b and the partition wall 11b, the bottom wall 11b, the partition wall 11c, the tubular portion 12a below the partition wall 11c, and the membrane 12e is referred to as the first space SP1. do.
  • the internal space of the tubular portion 12a is referred to as a second space SP2.
  • the first main surface 12ea is the main surface of the membrane 12e on the first space SP1 side
  • the second main surface 12eb is the main surface of the membrane 12e on the second space SP2 side. That is, a part of the first space SP1 is drawn by the first main surface 12ea, and a part of the second space SP2 is drawn by the second main surface 12eb.
  • the first medium 15 and the second medium 16 are stored in the first space SP1 and the second space SP2, respectively.
  • the second medium 16 is a medium having a lower dissolved oxygen concentration than the first medium 15.
  • the second medium 16 contains, for example, anaerobic bacteria.
  • Cells are cultured on the second main surface 12eb. These cells are, for example, intestinal epithelial cells that form tight junctions on the second main surface 12eb. Specific examples of these cells include Caco-2 cells. Oxygen in the first medium 15 is supplied to the cells via the membrane 12e.
  • the flange portion 12c projects from the outer peripheral surface of the tubular portion 12a in a plane orthogonal to the extending direction of the tubular portion 12a.
  • the flange portion 12c is located at the upper end of the tubular portion 12a.
  • the flange portion 12c and the partition wall 11c are hermetically sealed by the sealing member 14.
  • the tube 20a penetrates the lid member 13.
  • the tube 20a is connected to the second space SP2 at one end.
  • the tube 20a is connected to the culture medium container 21a at the other end.
  • the second medium 16 is stored in the medium container 21a.
  • the tube 20b penetrates the lid member 13.
  • the tube 20b is connected to the second space SP2 at one end.
  • the tube 20b is connected to the culture medium container 21b at the other end.
  • the medium container 21b stores the second medium 16 used for cell culture in the second space SP2.
  • the pump 30 is, for example, a tube pump. However, the pump 30 is not limited to this.
  • the pump 30 sends the second medium 16 stored in the medium container 21a via the tube 20a to the second space SP2, and the pump 30 sends the second medium stored in the second space SP2 via the tube 20b. 16 is discharged to the medium container 21b. As a result, the second medium 16 is replaced.
  • FIG. 2 is a block diagram of the pump 30.
  • the pump 30 includes a pump head 30a, a drive motor 30b, a controller 30c, and a wireless module 30d.
  • the pump head 30a is attached to the tube 20a and is rotationally driven by the drive motor 30b to perform the above-mentioned liquid feeding operation and the like.
  • the controller 30c is composed of, for example, a microcontroller.
  • the controller 30c controls the rotation speed of the drive motor 30b and the like based on the control signal supplied from the wireless module 30d.
  • the wireless module 30d has a circuit for performing baseband signal processing, a circuit for performing RF signal processing, and an antenna for transmitting and receiving RF signals, and is configured to enable wireless communication compliant with WiFi (registered trademark). It is a module that has been used.
  • the wireless module 30d may be configured to enable wireless communication in accordance with Bluetooth (registered trademark).
  • the wireless module 30d receives the above control signal.
  • FIG. 3 is a block diagram of the electrical resistance measuring device 40.
  • the electric resistance measuring device 40 has an electrode 40a and an electrode 40b, a measuring device main body 40c, and a wireless module 40d.
  • the electrode 40a penetrates the container body 11 and is in contact with the first medium 15 (see FIG. 1).
  • the electrode 40b penetrates the lid member 13 and is in contact with the second medium 16 (see FIG. 1).
  • the measuring device main body 40c is a trans-epithelial electrical resistance (TEER) measuring device.
  • the measuring device main body 40c is connected to the electrodes 40a and 40b.
  • the measuring device main body 40c measures the electric resistance value between the electrodes 40a and 40b, and outputs the electric resistance value to the wireless module 40d.
  • the wireless module 40d has the same configuration as the wireless module 30d.
  • the wireless module 40d transmits the above electric resistance value.
  • the electrical resistance value between the electrodes 40a and 40b varies depending on whether the cells cultured on the second main surface 12eb form a tight junction or not. Therefore, by measuring the above electric resistance value, it is possible to determine whether or not the cells cultured on the second main surface 12eb form a tight junction.
  • FIG. 4 is a block diagram of the dissolved oxygen concentration measuring device 50.
  • the dissolved oxygen concentration measuring device 50 includes an oxygen sensor 50a, a measuring device main body 50b, and a wireless module 50c.
  • the oxygen sensor 50a is arranged, for example, on the path of the tube 20b (see FIG. 1). As a result, the oxygen sensor 50a outputs a signal corresponding to the dissolved oxygen concentration of the second medium 16 discharged from the second space SP2.
  • the measuring device main body 50b is connected to the oxygen sensor 50a.
  • the measuring device main body 50b calculates the dissolved oxygen concentration of the second medium 16 based on the signal from the oxygen sensor 50a, and outputs the dissolved oxygen concentration to the wireless module 50c.
  • the wireless module 50c has the same configuration as the wireless module 30d (wireless module 40d).
  • the wireless module 50c transmits the above dissolved oxygen concentration.
  • the operation unit 60 is composed of, for example, a personal computer.
  • the operation unit 60 may be configured by a tablet terminal or the like.
  • the operation unit 60 has a built-in wireless module similar to the wireless module 30d (wireless module 40d, wireless module 50c).
  • the operation unit 60 receives an operation on the pump 30 via a keyboard, a mouse, a touch panel type liquid crystal display, or the like.
  • the operation unit 60 generates a control signal based on this operation, and transmits the control signal to the wireless module 30d via the wireless module built in the operation unit 60. As a result, the pump 30 is operated.
  • the electric resistance value transmitted from the wireless module 40d and the dissolved oxygen concentration transmitted from the wireless module 50c are received by the wireless module built in the operation unit 60.
  • the received electric resistance value and dissolved oxygen concentration are stored in the storage device of the operation unit 60 or displayed on the liquid crystal display or the like of the operation unit 60.
  • the operation unit 60 is arranged outside the anaerobic chamber 70.
  • the second medium 16 is replaced, but the first medium 15 may be further replaced. Further, in the above example, the dissolved oxygen concentration of the second medium 16 is measured, but the dissolved oxygen concentration of the first medium 15 may be further measured.
  • transmission / reception of a signal related to processing to the cell culture container 10 transmission of a control signal to the pump 30, reception of an electric resistance value from the electric resistance measuring device 40, and dissolved oxygen from the dissolved oxygen concentration measuring device 50). (Reception of density) can be performed by wireless communication with the operation unit 60.
  • the operability of the pump 30, the electric resistance measuring device 40, and the dissolved oxygen concentration measuring device 50 is provided, and the anaerobic chamber 70 is provided with a through hole through which a cable connecting these devices and the operating unit 60 is passed. It can be improved without providing it.
  • the electric resistance measuring device 40, and the dissolved oxygen concentration measuring device 50 are operated by inserting a hand from a glove or sleeve provided on the front surface of the anaerobic chamber 70, pressure fluctuation or vibration occurs when the hand is inserted. However, it may affect the cell culture in the cell culture container 10. In the cell culture system 100, it is not necessary to insert a hand from a glove or a sleeve provided on the front surface of the anaerobic chamber 70 in order to operate these devices. Therefore, according to the cell culture system 100, it is possible to suppress the generation of pressure fluctuations and vibrations associated with the operation of the pump 30, the electric resistance measuring device 40, and the dissolved oxygen concentration measuring device 50, and it is possible to perform stable cell culture.

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Organic Chemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Biotechnology (AREA)
  • Genetics & Genomics (AREA)
  • Biomedical Technology (AREA)
  • Microbiology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • Sustainable Development (AREA)
  • Analytical Chemistry (AREA)
  • Clinical Laboratory Science (AREA)
  • Computer Hardware Design (AREA)
  • Virology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Medicinal Chemistry (AREA)
  • Immunology (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)
PCT/JP2021/020605 2020-07-07 2021-05-31 細胞培養システム Ceased WO2022009560A1 (ja)

Priority Applications (4)

Application Number Priority Date Filing Date Title
JP2022534949A JPWO2022009560A1 (https=) 2020-07-07 2021-05-31
US18/014,866 US20230287326A1 (en) 2020-07-07 2021-05-31 Cell culture system
EP21837468.4A EP4180511A4 (en) 2020-07-07 2021-05-31 CELL CULTURE SYSTEM
CN202180048286.9A CN115916950A (zh) 2020-07-07 2021-05-31 细胞培养系统

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
JP2020116949 2020-07-07
JP2020-116949 2020-07-07

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WO2022009560A1 true WO2022009560A1 (ja) 2022-01-13

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PCT/JP2021/020605 Ceased WO2022009560A1 (ja) 2020-07-07 2021-05-31 細胞培養システム

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US (1) US20230287326A1 (https=)
EP (1) EP4180511A4 (https=)
JP (1) JPWO2022009560A1 (https=)
CN (1) CN115916950A (https=)
WO (1) WO2022009560A1 (https=)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2025225118A1 (ja) * 2024-04-23 2025-10-30 ウシオ電機株式会社 細胞培養デバイス

Citations (3)

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Publication number Priority date Publication date Assignee Title
WO2014002271A1 (ja) * 2012-06-29 2014-01-03 株式会社日立製作所 細胞組織検査システム、細胞培養装置、及び細胞組織検査方法
JP2015128426A (ja) * 2010-01-20 2015-07-16 イー・エム・デイー・ミリポア・コーポレイシヨン 細胞画像取得および遠隔モニタリングシステム
WO2018079793A1 (ja) 2016-10-28 2018-05-03 国立大学法人京都大学 嫌気性細菌などの細菌と上皮細胞との共培養装置及び共培養方法

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GB8916578D0 (en) * 1989-07-20 1989-09-06 Foster William J Anaerobic cabinet
US20140045252A1 (en) * 2011-04-28 2014-02-13 Hitachi, Ltd. Cell cultivation container and cell culturing apparatus
JP2015171344A (ja) * 2014-03-12 2015-10-01 オリンパス株式会社 細胞培養容器、細胞培養装置および細胞培養方法
JP6975373B2 (ja) * 2017-12-27 2021-12-01 澁谷工業株式会社 無菌作業システム
CN109609369A (zh) * 2018-12-24 2019-04-12 山东省食品药品检验研究院 一种具有自动控制监测功能的厌氧微生物培养系统
WO2020206121A2 (en) * 2019-04-03 2020-10-08 Flaskworks, Llc Cell culture systems and uses thereof

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JP2015128426A (ja) * 2010-01-20 2015-07-16 イー・エム・デイー・ミリポア・コーポレイシヨン 細胞画像取得および遠隔モニタリングシステム
WO2014002271A1 (ja) * 2012-06-29 2014-01-03 株式会社日立製作所 細胞組織検査システム、細胞培養装置、及び細胞組織検査方法
WO2018079793A1 (ja) 2016-10-28 2018-05-03 国立大学法人京都大学 嫌気性細菌などの細菌と上皮細胞との共培養装置及び共培養方法

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Title
See also references of EP4180511A4

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2025225118A1 (ja) * 2024-04-23 2025-10-30 ウシオ電機株式会社 細胞培養デバイス

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US20230287326A1 (en) 2023-09-14
CN115916950A (zh) 2023-04-04
EP4180511A4 (en) 2024-09-18
JPWO2022009560A1 (https=) 2022-01-13
EP4180511A1 (en) 2023-05-17

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