JP2007167002A - Cassette for cell culture, tool for attaching and detaching cassette for cell culture and cell culture apparatus - Google Patents

Cassette for cell culture, tool for attaching and detaching cassette for cell culture and cell culture apparatus Download PDF

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JP2007167002A
JP2007167002A JP2005370657A JP2005370657A JP2007167002A JP 2007167002 A JP2007167002 A JP 2007167002A JP 2005370657 A JP2005370657 A JP 2005370657A JP 2005370657 A JP2005370657 A JP 2005370657A JP 2007167002 A JP2007167002 A JP 2007167002A
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cell culture
cylindrical member
cassette
flange
outer cylindrical
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Aya Nitani
綾 二谷
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AGC Techno Glass Co Ltd
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Asahi Techno Glass Corp
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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
    • C12M23/00—Constructional details, e.g. recesses, hinges
    • C12M23/42—Integrated assemblies, e.g. cassettes or cartridges
    • 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

Abstract

<P>PROBLEM TO BE SOLVED: To provide a cassette for cell culture, which attaches and detaches a cell culture carrier without damaging it. <P>SOLUTION: The cell culture apparatus 10 is obtained by holding a cell culture carrier 3 between an outside tubular member 1 and an inside tubular member 2 constituting the cassette 20 for cell culture. Both the outside tubular member 1 and the inside tubular member 2 have cylindrical shapes and are equipped with flange-like projections 11 and 21 projecting inside at a lower opening part. The cell culture carrier 3 to be arranged between them is nipped by the projections 11 and 12 and fixed. Connection screws 13 and 23 to be mutually connected are formed on the inside surface of the outside tubular member 1 and on the outside surface of the inside tubular member 2. Two collar-shaped projections 14 extended outward are formed at mutually facing positions on the upper opening part of the outside tubular member 1 and holes 15 are formed on the collar-shaped projections 14. A fixed number of freestanding members 12 is installed under the projection 11 of he outside tubular member 1. <P>COPYRIGHT: (C)2007,JPO&INPIT

Description

本発明は、生体外で細胞または組織を培養する細胞培養装置に係り、特に、かかる細胞培養中に細胞間相互作用を解析するために有用な細胞培養用カセット、細胞培養用カセット着脱用治具及び細胞培養装置に関するものである。   The present invention relates to a cell culture apparatus for culturing cells or tissues in vitro, and in particular, a cell culture cassette useful for analyzing cell-cell interactions during such cell culture, and a jig for attaching / detaching a cell culture cassette And a cell culture device.

従来から、細胞を培養し、且つ培養後の細胞観察を容易なものとすることを目的として種々の提案がなされている。例えば、特許文献１には、ガラス製の底面部培養板を着脱可能とした培養容器が示されている。この培養容器は、細胞培養後にガラス製の底面部培養板を培養容器から取り外すことによって、培養細胞の観察を容易なものとしたものである。しかしながら、この特許文献１に記載された発明では、その構造上、ガラス製の底面部培養板を無菌的に取り外すことは困難であった。   Conventionally, various proposals have been made for the purpose of culturing cells and facilitating cell observation after culturing. For example, Patent Literature 1 discloses a culture vessel in which a glass bottom culture plate can be attached and detached. This culture vessel facilitates observation of cultured cells by removing the glass bottom culture plate from the culture vessel after cell culture. However, in the invention described in Patent Document 1, it is difficult to remove the glass bottom culture plate aseptically because of its structure.

また、特許文献２には、細胞培養膜として透析膜を利用し、この細胞培養膜を外側容器と内側容器で着脱自在に挟み込むように構成された培養用シャーレが示されている。この培養用シャーレにおいては、透析膜を外側容器と内側容器の側面と底面で強力に挟み込むことで、内側の培地と外側の培地を分離していた。   Patent Document 2 discloses a petri dish for culture configured such that a dialysis membrane is used as a cell culture membrane, and the cell culture membrane is detachably sandwiched between an outer container and an inner container. In this culture petri dish, the inner culture medium and the outer culture medium are separated by strongly sandwiching the dialysis membrane between the side surface and the bottom surface of the outer container and the inner container.

しかしながら、特許文献２に示された培養用シャーレでは、外側容器と内側容器に挟まれる透析膜には、側面方向への引っ張り力が強く加わるため、透析膜等のようにある程度強度を持った膜担体でしか使用することができないという欠点があった。すなわち、比較的強度の弱い素材を挟む場合には、膜担体が破損するという問題点があった。   However, in the petri dish for culture shown in Patent Document 2, a dialysis membrane sandwiched between the outer vessel and the inner vessel is strongly applied with a tensile force in the lateral direction, and thus a membrane having some strength such as a dialysis membrane. There was a drawback that it could only be used with a carrier. That is, when a relatively weak material is sandwiched, the membrane carrier is damaged.

例えば、特許文献３に示されたような、細胞外マトリックス（ＥＣＭ）成分含有溶液をゲル化させ、さらに乾燥させてガラス化し、次いで水和して得られるハイドロゲル薄膜のような比較的強度の弱い細胞培養担体は、それ自体に無理な力が加わると容易に破れてしまうことも少なくないため、特許文献２に示された培養用シャーレに適用することは困難であった。   For example, as shown in Patent Document 3, a solution containing an extracellular matrix (ECM) component is gelled, further dried, vitrified, and then hydrated to obtain a relatively strong strength such as a hydrogel thin film. Since weak cell culture carriers often break easily when an excessive force is applied to them, it has been difficult to apply the weak cell culture carrier to the culture dish shown in Patent Document 2.

さらなる従来技術として、培養容器内で、タンパク透過可能なポアを持ったＰＥＴ（ポリエチレンテレフタレート）膜を介して、上部培地と下部培地を分離することが可能な細胞培養インサートとして、セルカルチャーインサートTM（BDファルコンCode#353090）が市販されている。この培養容器を用いて培養を行うことで、細胞間相互作用、管腔形成、ガン浸潤の研究モデルの構築や、肝臓細胞を用いた薬物代謝試験・毒性試験を行うことが可能である。 As a further conventional technique, as a cell culture insert capable of separating an upper medium and a lower medium through a PET (polyethylene terephthalate) membrane having a pore capable of protein permeation in a culture container, Cell Culture Insert TM ( BD Falcon Code # 353090) is commercially available. By culturing using this culture vessel, it is possible to construct a research model for cell-cell interaction, tube formation, and cancer invasion, and to conduct drug metabolism tests and toxicity tests using liver cells.

しかしながら、この技術では、細胞培養担体部分が培養容器に固着されているため、それ自体を容易に着脱することができず、細胞培養膜を細胞が接着したまま回収するには、培養容器を破壊する必要があった。   However, in this technique, since the cell culture carrier part is fixed to the culture container, it cannot be easily attached and detached. To recover the cell culture membrane with the cells adhered, the culture container is destroyed. There was a need to do.

一方、着脱可能な透過解析装置としては、パームセル（ビードレックスCode#２００１０−０１）が市販されている。この透過解析装置は、腸間膜等を用いて薬物代謝や透過を調査する目的で薬理学の分野で多く用いられている。しかしながら、この透過解析装置は、その構造上、培養継続中の細胞の顕微鏡観察が不可能であった。   On the other hand, as a detachable transmission analyzer, a palm cell (Beadrex Code # 20010-01) is commercially available. This permeation analyzer is often used in the field of pharmacology for the purpose of investigating drug metabolism and permeation using the mesentery and the like. However, this permeation analyzer cannot be used for microscopic observation of cells during continuation of culture due to its structure.

つまり、エンドポイントでの評価しかできないため、事前の詳細な予備試験、もしくは経時的な細胞形態観察には、膨大な試験サンプルを必要としていた。また、培養液注入カラムの容積が大きく、サイトカインや増殖因子等の微量な細胞産生物の影響を調べるのには適していなかった。さらに、装置自体が大きく、細胞培養用インキュベータに入れることが困難であった。また、膜挟み部が開放系であることから、クリーンベンチ内での操作後にインキュベータへ移動した際、外気に接するという問題点もあった。
特開平１１−２０６３６６号公報 実開昭５２−１３５５００号公報 特開平８−２２８７６８号公報 In other words, since evaluation can only be performed at the end point, a large number of test samples are required for detailed preliminary tests in advance or observation of cell morphology over time. Moreover, the volume of the culture solution injection column is large, and it was not suitable for examining the influence of a small amount of cell products such as cytokines and growth factors. Furthermore, the apparatus itself is large and difficult to put in a cell culture incubator. In addition, since the membrane sandwiching portion is an open system, there is a problem in that it contacts outside air when it moves to the incubator after operation in the clean bench.
JP-A-11-206366 Japanese Utility Model Publication No. 52-135500 JP-A-8-228768

上述したような特許文献１及び特許文献２等で細胞培養担体として用いられているガラス板、透析膜及びＰＥＴ膜は、培養容器に挟持されるときに、細胞培養担体に強い引っ張り力が働いても、それに耐えられるだけの強度を備えているものであるが、近年開発が進んでいる特許文献３に開示されているような、ＥＣＭ成分等で構成されるハイドロゲル薄膜等は、細胞培養に適した多くの特徴を有するものの、その強度は、従来用いられていたガラス板、透析膜及びＰＥＴ膜よりも劣っている。従って、特許文献１及び特許文献２等に示された技術では、ハイドロゲル薄膜等の細胞培養担体を破損せずに挟み込むことは非常に困難であった。   The glass plate, dialysis membrane, and PET membrane used as the cell culture carrier in Patent Document 1 and Patent Document 2 as described above have a strong tensile force acting on the cell culture carrier when sandwiched between the culture vessels. However, a hydrogel thin film composed of ECM components, etc., as disclosed in Patent Document 3 which has been developed in recent years, is strong enough to withstand it. Although it has many suitable features, its strength is inferior to that of conventionally used glass plates, dialysis membranes and PET membranes. Therefore, it is very difficult to sandwich the cell culture carrier such as the hydrogel thin film without damaging it with the techniques shown in Patent Document 1 and Patent Document 2.

また、近年、細胞を培養後に、細胞を組織として移動・移植するという方法が再生医療の観点から注目されている。しかしながら、従来、培養後に細胞を回収する際には、トリプシン等の酵素で細胞をばらばらにした状態で回収しなければならなかった。つまり、培養後に細胞、特に組織を移動・回収することが困難であり、その後の細胞観察、固定染色、移植、その他解析操作が困難もしくは不可能であった。そのため、培養後の細胞の観察・移動・分泌物の解析・細胞間相互作用の解析を効率的に行うことができる細胞培養装置の開発が切望されていた。   In recent years, a method of transferring and transplanting cells as tissues after culturing the cells has attracted attention from the viewpoint of regenerative medicine. However, conventionally, when cells were collected after culturing, they had to be collected in a state where the cells were separated with an enzyme such as trypsin. That is, it is difficult to move and collect cells, particularly tissues after culture, and subsequent cell observation, fixed staining, transplantation, and other analysis operations are difficult or impossible. Therefore, the development of a cell culture device that can efficiently perform observation, migration, secretion analysis, and cell-cell interaction after culture has been eagerly desired.

その一方で、上記ハイドロゲル薄膜のようにＥＣＭ成分等で構成された細胞培養担体が市販されている。このハイドロゲル薄膜は、片面もしくは両面に細胞を培養することが可能であり、膜自体にタンパク透過能があり、素材がＥＣＭ等の生体適合性物質であることから、そのまま生体に戻すことができるという特性を有している。   On the other hand, cell culture carriers composed of ECM components and the like like the hydrogel thin film are commercially available. This hydrogel thin film can cultivate cells on one side or both sides, and the membrane itself has protein permeability, and since the material is a biocompatible substance such as ECM, it can be returned to the living body as it is. It has the characteristic.

このようなハイドロゲル薄膜を用いた従来の培養方法は、ディッシュ（通常は３５ｍｍを使用）に貼り付いた状態で培養し、その後、細胞が培養されたハイドロゲル薄膜をディッシュから剥がして移動・移植を行うというものであった。しかしながら、このようなハイドロゲル薄膜を剥がす作業は、作業者の手技によってハイドロゲル薄膜が破れることもあるため、より簡便に細胞組織を回収することができる細胞培養装置の開発が求められていた。   The conventional culture method using such a hydrogel thin film is cultured in a state where it is attached to a dish (usually 35 mm), and then the hydrogel thin film on which the cells are cultured is peeled off from the dish and transferred / transplanted. Was to do. However, since the operation of peeling such a hydrogel thin film may break the hydrogel thin film by an operator's technique, the development of a cell culture device that can more easily collect cell tissues has been demanded.

本発明は、上述したような従来技術の問題点を解消するために提案されたものであり、その第１の目的は、細胞培養担体を破損することなく脱着可能な細胞培養用カセットを提供することにある。
また、本発明の第２の目的は、細胞培養用カセットからの細胞培養担体の着脱操作を容易に実施することができる細胞培養用カセット着脱用治具を提供することにある。
さらに、本発明の第３の目的は、細胞培養担体により培養液を分離することができる細胞培養装置を提供することにある。 The present invention has been proposed to solve the above-described problems of the prior art, and a first object thereof is to provide a cell culture cassette that can be detached without damaging the cell culture carrier. There is.
A second object of the present invention is to provide a cell culture cassette attaching / detaching jig capable of easily attaching / detaching a cell culture carrier from a cell culture cassette.
Furthermore, a third object of the present invention is to provide a cell culture apparatus that can separate a culture solution using a cell culture carrier.

上記課題を解決するために、請求項１に記載の細胞培養用カセットは、共に筒状部材の一端に内側に突出するフランジ状の突起が形成された外側筒状部材と内側筒状部材とからなり、前記内側筒状部材を前記外側筒状部材の他端から挿入して、外側筒状部材と内側筒状部材を嵌合することによって、前記フランジ状の突起同士が圧接状態となるように構成されていることを特徴とするものである。   In order to solve the above-mentioned problem, the cell culture cassette according to claim 1 is composed of an outer cylindrical member and an inner cylindrical member, both of which are formed with flange-shaped protrusions protruding inward at one end of the cylindrical member. The inner cylindrical member is inserted from the other end of the outer cylindrical member, and the outer cylindrical member and the inner cylindrical member are fitted together so that the flange-shaped projections are in a pressure contact state. It is characterized by being comprised.

上記のような構成を有する請求項１に記載の発明によれば、外側筒状部材と内側筒状部材に形成されたフランジ状の突起同士を圧接状態とすることにより、突起による挟む力、すなわち、両フランジ状の突起の間に設置される細胞培養担体に対して垂直の力で細胞培養担体を挟持することができるので、細胞培養担体自体に外側に引っ張られる力が働くことがない。また、フランジ状の突起同士に隙間が形成されないので、培養液のしみ出しや流入を防ぐことができる。   According to the first aspect of the present invention having the above-described configuration, the flange-shaped projections formed on the outer cylindrical member and the inner cylindrical member are brought into a pressure contact state, whereby the force sandwiched by the projections, that is, Since the cell culture carrier can be sandwiched with a force perpendicular to the cell culture carrier placed between the two flange-shaped projections, the cell culture carrier itself does not have a force pulled outward. In addition, since no gap is formed between the flange-shaped protrusions, it is possible to prevent the culture medium from seeping out and flowing in.

請求項２に記載の細胞培養用カセットは、共に筒状部材の一端に内側に突出するフランジ状の突起が形成された外側筒状部材と内側筒状部材とからなり、前記外側筒状部材と内側筒状部材の間に細胞培養担体を設置した後、前記内側筒状部材を前記外側筒状部材の他端から挿入して、外側筒状部材と内側筒状部材を嵌合することによって、前記外側筒状部材及び内側筒状部材のフランジ状の突起が前記細胞培養担体を挟持するように構成されていることを特徴とするものである。   The cassette for cell culture according to claim 2 comprises an outer cylindrical member and an inner cylindrical member, both of which are formed with flange-shaped protrusions protruding inward at one end of the cylindrical member, After installing the cell culture carrier between the inner cylindrical members, by inserting the inner cylindrical member from the other end of the outer cylindrical member, by fitting the outer cylindrical member and the inner cylindrical member, A flange-like projection of the outer cylindrical member and the inner cylindrical member is configured to sandwich the cell culture carrier.

上記のような構成を有する請求項２に記載の発明によれば、外側筒状部材と内側筒状部材に形成されたフランジ状の突起同士を圧接状態とすることにより、突起による挟む力、すなわち、両フランジ状の突起の間に設置される細胞培養担体に対して垂直の力で細胞培養担体を挟持することができるので、細胞培養担体自体に外側に引っ張られる力が働くことがない。また、フランジ状の突起同士に隙間が形成されないので、培養液のしみ出しや流入を防ぐことができる。   According to the invention described in claim 2 having the above-described configuration, the flange-shaped projections formed on the outer cylindrical member and the inner cylindrical member are brought into a pressure contact state, whereby the force between the projections, Since the cell culture carrier can be sandwiched with a force perpendicular to the cell culture carrier placed between the flange-shaped projections, the cell culture carrier itself is not subjected to a force pulled outward. In addition, since no gap is formed between the flange-shaped protrusions, it is possible to prevent the culture medium from seeping out and flowing in.

請求項３に記載の発明は、請求項２に記載の細胞培養用カセットにおいて、前記細胞培養担体が、リング状の支持体上に細胞培養担体膜を形成して構成され、前記リング状の支持体部分を、前記外側筒状部材及び内側筒状部材のフランジ状の突起によって挟持するように構成されていることを特徴とするものである。   The invention according to claim 3 is the cell culture cassette according to claim 2, wherein the cell culture carrier is formed by forming a cell culture carrier film on a ring-shaped support, and the ring-shaped support The body portion is configured to be sandwiched between flange-like protrusions of the outer cylindrical member and the inner cylindrical member.

上記のような構成を有する請求項３に記載の発明によれば、支持体部分を両フランジ状の突起で挟持することにより、フランジ状の突起と支持体との間に隙間を形成することなく取り付けることができるので、たとえ細胞培養担体を構成する細胞培養担体膜が薄くても、突起と支持体との間からの培養液のしみ出しや流入を防ぐことができる。また、挟持時の押圧力が細胞培養担体膜へ直接かからないので、細胞培養担体膜の破損の可能性を大幅に低減することができる。   According to the invention of claim 3 having the above-described configuration, the support portion is sandwiched between the flange-shaped projections, so that no gap is formed between the flange-shaped projection and the support. Since it can be attached, even if the cell culture carrier film constituting the cell culture carrier is thin, it is possible to prevent the culture medium from seeping out and flowing in between the protrusion and the support. Further, since the pressing force at the time of clamping is not directly applied to the cell culture carrier film, the possibility of breakage of the cell culture carrier film can be greatly reduced.

請求項４に記載の発明は、請求項１乃至請求項３のいずれか一に記載の細胞培養用カセットにおいて、前記外側筒状部材の内側面及び内側筒状部材の外側面に、互いに係合するネジ部が形成されていることを特徴とするものである。
上記のような構成を有する請求項４に記載の発明によれば、ネジ部を形成することにより、フランジ状の突起同士の圧接状態を容易に形成でき、且つ維持することができる。 According to a fourth aspect of the present invention, in the cassette for cell culture according to any one of the first to third aspects, the inner surface of the outer cylindrical member and the outer surface of the inner cylindrical member are engaged with each other. The screw part to be formed is formed.
According to invention of Claim 4 which has the above structures, the press-contact state of flange-shaped protrusions can be formed easily and can be maintained by forming a screw part.

請求項５に記載の発明は、請求項１乃至請求項４のいずれか一に記載の細胞培養用カセットにおいて、前記外側筒状部材に形成されたフランジ状の突起の上面、及び内側筒状部材に形成されたフランジ状の突起の下面の少なくとも一方に、クッション材が取付けられていることを特徴とするものである。   The invention according to claim 5 is the cell culture cassette according to any one of claims 1 to 4, wherein the upper surface of the flange-shaped protrusion formed on the outer cylindrical member, and the inner cylindrical member A cushioning material is attached to at least one of the lower surfaces of the flange-shaped projections formed in the above.

上記のような構成を有する請求項５に記載の発明によれば、フランジ状の突起にクッション材を取り付けることにより、それぞれの突起の圧接に関与する面の表面状態に影響されることなく、細胞培養担体を面で保持することができるようになる。また、密閉度も向上させることができるので、さらに培養液のしみ出しや流入を効果的に防ぐことができる。   According to the invention of claim 5 having the above-described configuration, by attaching a cushion material to the flange-shaped protrusion, the cell is not affected by the surface state of the surface involved in the pressure contact of each protrusion. The culture carrier can be held on the surface. Moreover, since the degree of sealing can be improved, it is possible to effectively prevent the culture medium from seeping out and flowing in.

請求項６に記載の発明は、請求項１乃至請求項５のいずれか一に記載の細胞培養用カセットにおいて、前記外側筒状部材のフランジ状の突起が形成されていない端部、及び内側筒状部材のフランジ状の突起が形成されていない端部の少なくとも一方に、筒状部材の外方に延びる突出部が形成されていることを特徴とするものである。   The invention according to claim 6 is the cell culture cassette according to any one of claims 1 to 5, wherein the outer cylindrical member is formed with an end portion on which the flange-shaped protrusion is not formed, and the inner cylinder. A protruding portion extending outward of the cylindrical member is formed on at least one of the end portions of the cylindrical member where the flange-shaped protrusion is not formed.

上記のような構成を有する請求項６に記載の発明によれば、突出部をピンセットその他無菌処理された器具で押さえつけることにより、外側筒状部材と内側筒状部材の着脱を容易に行うことができるようになる。   According to the invention of claim 6 having the above-described configuration, the outer cylindrical member and the inner cylindrical member can be easily attached and detached by pressing the protruding portion with tweezers or other aseptically processed instruments. become able to.

請求項７に記載の発明は、請求項６に記載の細胞培養用カセットにおいて、前記突出部に、少なくとも一つの孔が形成されていることを特徴とするものである。
上記のような構成を有する請求項７に記載の発明によれば、この孔に棒状の部材を挿入することにより、外側筒状部材あるいは内側筒状部材を回動させることが容易になるので、外側筒状部材と内側筒状部材の着脱をさらに容易に行うことができるようになる。 The invention according to claim 7 is the cell culture cassette according to claim 6, wherein at least one hole is formed in the protruding portion.
According to the invention of claim 7 having the above-described configuration, by inserting a rod-like member into this hole, it becomes easy to rotate the outer cylindrical member or the inner cylindrical member. The outer cylindrical member and the inner cylindrical member can be attached and detached more easily.

請求項８に記載の発明は、請求項１乃至請求項７のいずれか一に記載の細胞培養用カセットにおいて、前記外側筒状部材に形成されたフランジ状の突起の外面の少なくとも３箇所に、所定の高さを有する自立部材が形成されていることを特徴とするものである。
上記のような構成を有する請求項８に記載の発明によれば、培養液を入れた培養容器（例えば、シャーレやディッシュ）内に細胞培養用カセットを載置した場合でも、細胞培養用カセットの外底面に培養液を接触させることができる。 The invention according to claim 8 is the cell culture cassette according to any one of claims 1 to 7, wherein at least three locations on the outer surface of the flange-shaped protrusion formed on the outer cylindrical member, A self-supporting member having a predetermined height is formed.
According to the invention of claim 8 having the above-described configuration, even when the cell culture cassette is placed in a culture container (eg, a petri dish or a dish) containing a culture solution, The culture solution can be brought into contact with the outer bottom surface.

請求項９に記載の細胞培養用カセット着脱用治具は、請求項１乃至請求項８のいずれか一に記載の細胞培養用カセットの外側筒状部材と内側筒状部材の少なくとも一方を固定することができるように構成されていることを特徴とするものである。
上記のような構成を有する請求項９に記載の発明によれば、筒状部材の少なくともいずれか一方を固定することにより、人が直接細胞培養用カセットに触れる機会を減らすことができるので、無菌的な着脱操作の精度を向上させることができる。 The cell culture cassette attaching / detaching jig according to claim 9 fixes at least one of the outer cylindrical member and the inner cylindrical member of the cell culture cassette according to any one of claims 1 to 8. It is comprised so that it can be performed.
According to the invention described in claim 9 having the above-described configuration, by fixing at least one of the cylindrical members, it is possible to reduce the chance that a person directly touches the cell culture cassette. The accuracy of a typical detachment operation can be improved.

請求項１０に記載の細胞培養装置は、支持体に取り付けられた細胞培養担体膜であって、片面への細胞培養、両面への細胞培養、及びタンパク質透過の少なくとも一つが可能な細胞培養担体膜を、請求項１乃至請求項８のいずれか一に記載の細胞培養用カセットの外側筒状部材と内側筒状部材の間に設置したことを特徴とするものである。   The cell culture device according to claim 10, which is a cell culture carrier membrane attached to a support, and is capable of at least one of cell culture on one side, cell culture on both sides, and protein permeation. Is installed between the outer cylindrical member and the inner cylindrical member of the cassette for cell culture according to any one of claims 1 to 8.

上記のような構成を有する請求項１０に記載の発明によれば、細胞培養担体膜を支持体をつけたままの状態で培養することができるので、培養後に細胞培養装置から容易に細胞培養担体を取り出すことができ、細胞が培養された細胞培養担体膜の取り扱いを極めて容易に行なうことができる。また、細胞培養担体を一旦取り出して反転させ、再度、細胞培養装置にセットすることも容易に行なうことができる。   According to the invention of claim 10 having the above-described configuration, since the cell culture carrier membrane can be cultured with the support attached, the cell culture carrier can be easily obtained from the cell culture device after the culture. The cell culture carrier membrane in which the cells are cultured can be handled very easily. It is also possible to easily take out the cell culture carrier, invert it, and set it again in the cell culture apparatus.

請求項１１に記載の発明は、請求項１０に記載の細胞培養装置において、前記支持体が不織布から構成されていることを特徴とするものである。
上記のような構成を有する請求項１１に記載の発明によれば、支持体の強度が高まるので、細胞培養担体の取り扱いが容易なものとなる。 The invention according to claim 11 is the cell culture apparatus according to claim 10, wherein the support is made of a nonwoven fabric.
According to the invention of the eleventh aspect having the above-described configuration, the strength of the support is increased, so that the cell culture carrier can be easily handled.

本発明によれば、細胞培養担体を破損することなく脱着可能な細胞培養用カセットを提供することができる。また、細胞培養用カセットからの細胞培養担体の着脱操作を容易に実施することができる細胞培養用カセット着脱用治具を提供することができる。さらに、細胞培養担体により培養液を分離することができる細胞培養装置を提供することができる。   According to the present invention, it is possible to provide a cell culture cassette that can be detached without damaging the cell culture carrier. Further, it is possible to provide a cell culture cassette attaching / detaching jig capable of easily performing the operation of attaching / detaching the cell culture carrier from the cell culture cassette. Furthermore, the cell culture apparatus which can isolate | separate a culture solution with a cell culture carrier can be provided.

以下、本発明の実施の形態について、図面を参照して具体的に説明する。   Hereinafter, embodiments of the present invention will be specifically described with reference to the drawings.

（１）構成
（１−１）全体構成
本実施形態の細胞培養装置１０は、図１に示すように、細胞培養用カセット２０を構成する外側筒状部材１と内側筒状部材２との間に、細胞培養担体３を挟持して構成されている。なお、図１は本発明に係る細胞培養装置１０の分解斜視図、図２は細胞培養装置１０の全体像を示す斜視図、図３は細胞培養装置１０の全体像を示す断面図、図４は細胞培養装置１０を既存の培養容器３０内に設置した状態を示す斜視図である。以下、各構成要素について詳述する。 (1) Configuration (1-1) Overall Configuration As shown in FIG. 1, the cell culture apparatus 10 of the present embodiment is between an outer cylindrical member 1 and an inner cylindrical member 2 that constitute a cell culture cassette 20. In addition, the cell culture carrier 3 is sandwiched. 1 is an exploded perspective view of the cell culture device 10 according to the present invention, FIG. 2 is a perspective view showing an overall image of the cell culture device 10, FIG. 3 is a cross-sectional view showing the overall image of the cell culture device 10, and FIG. FIG. 2 is a perspective view showing a state where the cell culture device 10 is installed in an existing culture vessel 30. Hereinafter, each component will be described in detail.

（１−２）外側筒状部材
細胞培養用カセット２０を構成する外側筒状部材１は、円筒形状を有し、下側の開口部に内側に突出するフランジ状の突起１１が形成され、このフランジ状の突起１１の下面には、所定個数の自立部材１２が設けられている。また、外側筒状部材１の内側面には、所定の間隔をあけて、所定の長さを有する係合用ネジ部１３が形成されている。さらに、上側の開口部には、外方に延びる２つの鍔状の突出部１４が互いに対向する位置に形成され、この鍔状の突出部１４には孔１５が形成されている。なお、前記フランジ状の突起１１の上面に、シリコン等で形成されたクッション材を設置しても良い。 (1-2) Outer cylindrical member The outer cylindrical member 1 constituting the cell culture cassette 20 has a cylindrical shape, and a flange-shaped protrusion 11 protruding inward is formed in the lower opening. A predetermined number of self-supporting members 12 are provided on the lower surface of the flange-shaped protrusion 11. Further, on the inner side surface of the outer cylindrical member 1, an engagement screw portion 13 having a predetermined length is formed at a predetermined interval. Further, two ridge-like protrusions 14 extending outward are formed at positions facing each other in the upper opening, and a hole 15 is formed in the ridge-like protrusion 14. A cushion material made of silicon or the like may be installed on the upper surface of the flange-shaped protrusion 11.

（１−３）内側筒状部材
細胞培養用カセット２０を構成する内側筒状部材２は、円筒形状を有し、前記外側筒状部材１の上側の開口部から外側筒状部材１内に嵌め込むことができるように構成されている。また、内側筒状部材２の下側の開口部には、前記外側筒状部材１と同様に、内側に突出するフランジ状の突起２１が形成されている。そして、これらのフランジ状の突起１１、２１により、それらの間に設置された細胞培養担体３を挟持し、固定することができるように構成されている。 (1-3) Inner cylindrical member The inner cylindrical member 2 constituting the cell culture cassette 20 has a cylindrical shape and is fitted into the outer cylindrical member 1 from the upper opening of the outer cylindrical member 1. It is configured so that it can be included. Further, similarly to the outer cylindrical member 1, a flange-like projection 21 protruding inward is formed in the lower opening of the inner cylindrical member 2. The flange-shaped protrusions 11 and 21 are configured so that the cell culture carrier 3 placed between them can be sandwiched and fixed.

また、内側筒状部材２の外側面には、前記外側筒状部材１の内側面に形成された係合用ネジ部１３と係合する係合用ネジ部２３が形成されている。さらに、上側の開口部には、外側筒状部材１と同様に、外方に延びる２つの鍔状の突出部２４が互いに対向する位置に形成され、この鍔状の突出部２４には孔２５が形成されている。なお、前記フランジ状の突起２１の下面にも、シリコン等で形成されたクッション材を設置しても良い。   Further, on the outer surface of the inner cylindrical member 2, an engagement screw portion 23 that engages with the engagement screw portion 13 formed on the inner surface of the outer cylindrical member 1 is formed. Further, in the same manner as the outer cylindrical member 1, two outwardly extending hook-like protrusions 24 are formed at positions facing each other in the upper opening, and a hole 25 is formed in the hook-like protrusion 24. Is formed. Note that a cushion material made of silicon or the like may be provided on the lower surface of the flange-shaped protrusion 21.

（１−４）細胞培養担体
前記外側筒状部材１と内側筒状部材２の間に挿入される細胞培養担体３は、ナイロンメンブレン等をドーナツ状に切り抜いた支持体３１上に、コラーゲン等のハイドロゲルを乾燥してガラス化させ、その後水和させたハイドロゲル薄膜３２を形成して構成されている。 (1-4) Cell culture carrier The cell culture carrier 3 inserted between the outer cylindrical member 1 and the inner cylindrical member 2 is made of collagen or the like on a support 31 obtained by cutting a nylon membrane or the like into a donut shape. The hydrogel is dried and vitrified, and then hydrated hydrogel thin film 32 is formed.

そして、外側筒状部材１のフランジ状の突起１１の上に、細胞培養担体３の支持体３１部分を乗せ、その後、内側筒状部材２を外側筒状部材１に嵌め込み、外側筒状部材１のフランジ状の突起１１と内側筒状部材２のフランジ状の突起２１によって細胞培養担体３の支持体３１を挟持することによって、図２及び図３に示すような細胞培養装置１０が形成される。   Then, the support 31 portion of the cell culture carrier 3 is placed on the flange-shaped protrusion 11 of the outer cylindrical member 1, and then the inner cylindrical member 2 is fitted into the outer cylindrical member 1, and the outer cylindrical member 1 is inserted. The cell culture device 10 as shown in FIGS. 2 and 3 is formed by sandwiching the support 31 of the cell culture carrier 3 by the flange-shaped projection 11 and the flange-shaped projection 21 of the inner cylindrical member 2. .

また、細胞培養用カセット２０に挟まれる細胞培養担体３としては、片面または両面に細胞を培養することができ、細胞の顕微鏡観察に耐えうる透明度を持ち、種々の細胞産生因子を透過することができる膜を用いることが好ましい。さらに好ましくは細胞培養担体３自体の生体適合性が高く、そのまま移植に耐えうるものが良い。例えば、上記特許文献３に記載のハイドロゲル薄膜が好適である。   Further, as the cell culture carrier 3 sandwiched between the cell culture cassettes 20, cells can be cultured on one side or both sides, have transparency that can withstand microscopic observation of cells, and can transmit various cell production factors. It is preferable to use a film that can be used. More preferably, the cell culture carrier 3 itself is highly biocompatible and can withstand transplantation as it is. For example, the hydrogel thin film described in Patent Document 3 is suitable.

また、外側筒状部材１と内側筒状部材２の間に挟まれる細胞培養担体３の枚数は１枚でなくても良い。細胞培養担体３を複数枚とすることにより、ハイドロゲル薄膜を支持する支持体３１も複数枚となることから、クッション材がない状態でもフランジ状の両突起１１、２１間の密閉度を向上させることができる。また、２枚の細胞培養担体３を用いて両面培養を行った場合には、培養を行った各細胞について異なる評価をするために、細胞培養担体３ごとに所望の培地に移したり、別の処理を行うことができる。   The number of cell culture carriers 3 sandwiched between the outer cylindrical member 1 and the inner cylindrical member 2 may not be one. By using a plurality of cell culture carriers 3, a plurality of supports 31 for supporting the hydrogel thin film are also provided, so that the degree of sealing between the flange-shaped protrusions 11 and 21 is improved even without a cushioning material. be able to. In addition, when double-sided culture is performed using two cell culture carriers 3, each cell culture carrier 3 is transferred to a desired medium in order to perform different evaluation on each cultured cell, Processing can be performed.

なお、前記支持体としては、不織布を用いることが好ましい。この不織布には、人工繊維で成形されたものや、天然繊維で形成されたものが用いられる。具体的には、ナイロン、ポリフッ化ビニリデン、ニトロセルロース、綿、絹等が好ましい。   In addition, it is preferable to use a nonwoven fabric as the support. As this non-woven fabric, those formed from artificial fibers and those formed from natural fibers are used. Specifically, nylon, polyvinylidene fluoride, nitrocellulose, cotton, silk and the like are preferable.

（１−５）細胞培養装置
細胞培養装置１０の主要部である細胞培養用カセット２０の素材は、細胞培養に適するもの、細胞毒性の低いものであれば採用可能である。例えば、ポリエチレン、ポリスチレン、ポリエチレンテレフタラート及び、ポリプロピレンのような各種素材で成形可能である。その他、特に細胞培養装置１０に細胞が接着する必要性はないので、細胞毒性を持たない金属（具体的には、ステンレス鋼、チタン合金、金、銀及び白金等）でも作製可能であるが、量産性を考慮すると素材は多少の弾性を備える樹脂製ものが好適である。 (1-5) Cell Culture Device The material of the cell culture cassette 20 that is the main part of the cell culture device 10 can be used as long as it is suitable for cell culture and has low cytotoxicity. For example, it can be molded from various materials such as polyethylene, polystyrene, polyethylene terephthalate, and polypropylene. In addition, since it is not particularly necessary for the cells to adhere to the cell culture device 10, it can be made of a metal having no cytotoxicity (specifically, stainless steel, titanium alloy, gold, silver, platinum, etc.) In consideration of mass productivity, the material is preferably made of resin having some elasticity.

（１−６）培養容器
前記細胞培養装置１０は、図４に示すように、既存の培養容器３０内に設置されて用いられる。従って、前記細胞培養装置１０は、既存の培養容器３０内に挿入可能な形状・大きさとすることが好ましい。既存の培養容器３０としては、例えば、６０ｍｍディッシュ（IWAKI Code#３０１０−０６０X）、３５ｍｍディッシュ（IWAKI Code#３０００−０３５X）等が用いられる。既存の培養容器３０は円形であることが多いが、その他の形状でも良いことは言うまでもない。 (1-6) Culture container As shown in FIG. 4, the cell culture apparatus 10 is installed and used in an existing culture container 30. Therefore, it is preferable that the cell culture device 10 has a shape and size that can be inserted into the existing culture vessel 30. As the existing culture container 30, for example, a 60 mm dish (IWAKI Code # 3010-060X), a 35 mm dish (IWAKI Code # 3000-035X), or the like is used. The existing culture vessel 30 is often circular, but it goes without saying that other shapes may be used.

（１−７）係合用ネジ部
なお、本実施形態においては、細胞培養装置１０を構成する外側筒状部材１と内側筒状部材２とをより強靭に固定するための手段として、外側筒状部材１の内側面と、内側筒状部材２の外側面のそれぞれに、互いに係合する係合用ネジ部１３、２３が設けられている。 (1-7) Engagement screw portion In this embodiment, as a means for more firmly fixing the outer cylindrical member 1 and the inner cylindrical member 2 constituting the cell culture device 10, an outer cylindrical shape is used. Engagement thread portions 13 and 23 that engage with each other are provided on the inner side surface of the member 1 and the outer side surface of the inner cylindrical member 2.

（１−８）クッション材
また、本実施形態においては、上下培地の分離能を上昇させることを目的として、該細胞培養装置１０の外側筒状部材１のフランジ状の突起１１の上面、あるいは内側筒状部材２のフランジ状の突起２１の下面の少なくとも一方にクッション材が取り付けられている。これにより、細胞培養担体３をより密に挟み込むことができるので、上下の培地のコンタミネーションをより高度に防止することができる。なお、クッション材の素材としては、弾性を有し、細胞毒性を持たないものであれば採用可能である。例えば、シリコン樹脂等が好適である。 (1-8) Cushion material In the present embodiment, the upper surface or the inner side of the flange-shaped protrusion 11 of the outer cylindrical member 1 of the cell culture device 10 for the purpose of increasing the separation ability of the upper and lower culture media. A cushion material is attached to at least one of the lower surfaces of the flange-shaped protrusions 21 of the tubular member 2. Thereby, since the cell culture support | carrier 3 can be pinched | interposed more densely, the contamination of an upper and lower culture medium can be prevented more highly. As a material for the cushion material, any material that has elasticity and does not have cytotoxicity can be used. For example, silicon resin or the like is suitable.

（１−９）鍔状の突出部
さらに、本実施形態の細胞培養装置１０においては、内側筒状部材２を外側筒状部材１から着脱する際の無菌的操作を容易に実現するための手段として、少なくとも外側筒状部材１に、その頂部壁から外方に伸長する鍔状の突出部１４が設けられている。 (1-9) Sponge-shaped projecting portion Furthermore, in the cell culture device 10 of the present embodiment, means for easily realizing aseptic operation when the inner tubular member 2 is attached to and detached from the outer tubular member 1. As described above, at least the outer cylindrical member 1 is provided with a hook-shaped protrusion 14 extending outward from its top wall.

この外側筒状部材１に設けられた鍔状の突出部１４を、ピンセットその他無菌処理された器具で押さえつけた後、外側筒状部材１の筒状部より上に突き出している内側筒状部材２を鉗子その他の無菌処理された器具で挟んで、内側筒状部材２をその周方向に回動させ、係合用ネジ部１３、２３の係合を外すことにより、内側筒状部材２を外側筒状部材１から無菌的に引き出すことができる。   The inner tubular member 2 protruding above the tubular portion of the outer tubular member 1 after pressing the hook-shaped protruding portion 14 provided on the outer tubular member 1 with tweezers or other aseptically treated instruments. Is sandwiched between forceps and other sterilized instruments, the inner cylindrical member 2 is rotated in the circumferential direction, and the engagement screw portions 13 and 23 are disengaged. It can be aseptically pulled out from the shaped member 1.

なお、外側筒状部材１に内側筒状部材２を挿入する際には、上記と逆の操作を行えば良い。また、外側筒状部材１だけでなく、内側筒状部材２にもその頂部壁から外方に伸長する鍔状の突出部２４を設けても良いことは言うまでもない。   In addition, when inserting the inner cylindrical member 2 into the outer cylindrical member 1, an operation reverse to the above may be performed. Needless to say, not only the outer cylindrical member 1 but also the inner cylindrical member 2 may be provided with a hook-like protrusion 24 extending outward from its top wall.

また、外側筒状部材１の鍔状の突出部１４、内側筒状部材２の鍔状の突出部２４の少なくともいずれか一方に孔１５、２５を形成することにより、この孔１５、２５内にピンセットその他無菌処理された器具を挿入して筒状部材を捻じることができるので、外側筒状部材１と内側筒状部材２の着脱操作をさらに容易に行うことができるようになる。   Further, by forming holes 15 and 25 in at least one of the flange-shaped protrusion 14 of the outer cylindrical member 1 and the flange-shaped protrusion 24 of the inner cylindrical member 2, the holes 15 and 25 are formed in the holes 15 and 25. Since the cylindrical member can be twisted by inserting tweezers or other sterilized instrument, the attaching / detaching operation of the outer cylindrical member 1 and the inner cylindrical member 2 can be performed more easily.

（１−１０）自立部材
また、本実施形態においては、外側筒状部材１の下方開口端またはフランジ状の突起１１の下面に、少なくとも３個以上の自立部材１２が設けられている。この自立部材１２により、細胞培養装置１０に設置された細胞培養担体３と、細胞培養装置１０が収納配置される培養容器３０の内底面との間に隙間を形成することができる。 (1-10) Self-supporting member In the present embodiment, at least three or more self-supporting members 12 are provided on the lower opening end of the outer cylindrical member 1 or the lower surface of the flange-shaped protrusion 11. By this self-supporting member 12, a gap can be formed between the cell culture carrier 3 installed in the cell culture device 10 and the inner bottom surface of the culture vessel 30 in which the cell culture device 10 is accommodated.

このように、細胞培養装置１０の下部に隙間を設けることにより、培養液を入れた培養容器３０に細胞培養装置１０を入れて細胞を培養する場合に、培養容器３０中の培養液による影響を把握したり、細胞培養担体３の両面に細胞を培養することが可能となる。   Thus, by providing a gap at the lower part of the cell culture device 10, when the cell culture device 10 is placed in the culture vessel 30 containing the culture solution and the cells are cultured, the influence of the culture solution in the culture vessel 30 is affected. It becomes possible to grasp or to culture cells on both surfaces of the cell culture carrier 3.

（１−１１）細胞培養用カセット着脱用治具
前記係合用ネジ部１３、２３の開閉をより簡便にする手段として、細胞培養用カセット着脱用治具（以下、着脱用治具という）を用いることができる。この着脱用治具には、細胞培養用カセット２０の外側筒状部材１が嵌り込む凹部が形成されており、その凹部の底面には、外側筒状部材１の底面に形成された自立部材１２が嵌合する穴が形成されている。 (1-11) Cell culture cassette attaching / detaching jig A cell culture cassette attaching / detaching jig (hereinafter referred to as an attaching / detaching jig) is used as a means for making the engagement screw portions 13 and 23 easier to open and close. be able to. In this attachment / detachment jig, a concave portion into which the outer cylindrical member 1 of the cell culture cassette 20 is fitted is formed, and a self-standing member 12 formed on the bottom surface of the outer cylindrical member 1 is formed on the bottom surface of the concave portion. A hole for fitting is formed.

そして、この穴に外側筒状部材１の自立部材１２が嵌合することにより、着脱用治具と外側筒状部材１とがロックされるため、外側筒状部材１に内側筒状部材２を嵌め込んで、係合用ネジ部１３、２３を係合させるために捻るときに、着脱用治具内における外側筒状部材１の水平方向への回転を防ぐことができる。   Then, since the self-standing member 12 of the outer cylindrical member 1 is fitted into this hole, the attaching / detaching jig and the outer cylindrical member 1 are locked, so that the inner cylindrical member 2 is attached to the outer cylindrical member 1. When fitted and twisted to engage the engaging screw portions 13 and 23, the outer cylindrical member 1 can be prevented from rotating in the horizontal direction in the attaching / detaching jig.

また、外側筒状部材１が着脱用治具の凹部に完全に埋没する場合には、着脱用治具の凹部の上縁に、外側筒状部材１の鍔状の突出部１４が嵌合する切り欠きを形成しても良く、この切り欠きのみで着脱時の回転防止機構としても良い。   Further, when the outer cylindrical member 1 is completely buried in the concave portion of the attaching / detaching jig, the hook-like protruding portion 14 of the outer cylindrical member 1 is fitted to the upper edge of the concave portion of the attaching / detaching jig. A notch may be formed, and only this notch may serve as a rotation prevention mechanism at the time of attachment / detachment.

なお、外側筒状部材１から内側筒状部材２を着脱する際には、内側筒状部材２の鍔状の突出部２４に形成された孔２５にピンセット等を挿入し、外側筒状部材１に対して内側筒状部材２を所望の方向に回動させることにより、係合用ネジ部１３、２３の係合を容易に外すことができる。また、内側筒状部材２を着脱用治具で固定するように構成しても良いことは言うまでもない。   In addition, when attaching and detaching the inner cylindrical member 2 from the outer cylindrical member 1, tweezers or the like is inserted into the hole 25 formed in the flange-shaped protruding portion 24 of the inner cylindrical member 2, and the outer cylindrical member 1. On the other hand, by engaging the inner cylindrical member 2 in a desired direction, the engagement screw portions 13 and 23 can be easily disengaged. Moreover, it cannot be overemphasized that you may comprise so that the inner side cylindrical member 2 may be fixed with the jig | tool for attachment / detachment.

（２）使用態様
上記のような構成を有する本実施形態の細胞培養装置は、以下のような種々の培養方法に適用することができる。
（ａ）所定の培養液を入れた培養容器３０内に、細胞培養担体３を挟持した細胞培養装置１０を設置し、細胞培養担体３の上下いずれか一方の面に細胞を培養する。 (2) Usage Mode The cell culture device of the present embodiment having the above-described configuration can be applied to the following various culture methods.
(A) A cell culture apparatus 10 holding the cell culture carrier 3 is placed in a culture vessel 30 containing a predetermined culture solution, and cells are cultured on either the upper or lower surface of the cell culture carrier 3.

（ｂ）上記（ａ）のようにして培養した細胞に対して、その細胞が浸されている培養液、細胞培養担体３を介した培養液及び両方の培養液のいずれかの培養液に種々の因子を加えて培養する。なお、添加する因子としては、例えば、インターロイキン（ＩＬ）、コロニー刺激因子（ＣＳＦ）、幹細胞因子（ＳＣＦ）、腫瘍壊死因子（ＴＮＦ）、インターフェロン（ＩＦＮ）等、試験対象や試験項目に対応する種々の因子を添加することができる。 (B) For the cells cultured as described in (a) above, there are various types of culture solutions in which the cells are immersed, the culture solution via the cell culture carrier 3 and any one of both culture solutions. Add the above factors and culture. In addition, as a factor to be added, for example, interleukin (IL), colony stimulating factor (CSF), stem cell factor (SCF), tumor necrosis factor (TNF), interferon (IFN) and the like correspond to test subjects and test items. Various factors can be added.

（ｃ）培養容器３０内に細胞培養担体３を挟持した細胞培養装置１０を設置し、培養容器３０の底面と細胞培養担体３の上面とに異なる細胞を培養し、細胞培養担体３を介した双方の細胞間における影響を解析する。
（ｄ）細胞培養担体３の上下両面に異なる細胞を培養する。
（ｅ）上記（ｃ）及び（ｄ）において、（ｂ）のように培養液に種々の因子を加えて培養する。 (C) The cell culture device 10 having the cell culture carrier 3 sandwiched in the culture vessel 30 is installed, different cells are cultured on the bottom surface of the culture vessel 30 and the top surface of the cell culture carrier 3, and the cell culture carrier 3 is interposed. Analyze the effects between both cells.
(D) Different cells are cultured on the upper and lower surfaces of the cell culture carrier 3.
(E) In the above (c) and (d), various factors are added to the culture solution as shown in (b) and cultured.

このように、本発明に係る細胞培養装置１０を用いた細胞培養法として、細胞培養担体３の片面に細胞を培養し、細胞培養用カセット２０に挟み込んで細胞培養装置１０とし、この細胞培養装置１０を、シャーレ等の培養容器３０に入れ、さらにサイトカイン等の因子を加えることによって、細胞の機能解析が可能である。   As described above, as a cell culture method using the cell culture device 10 according to the present invention, cells are cultured on one side of the cell culture carrier 3 and sandwiched between the cell culture cassettes 20 to form the cell culture device 10. 10 is placed in a culture container 30 such as a petri dish, and a factor such as a cytokine is added to analyze the function of the cell.

また、細胞培養装置１０を用いた細胞培養法として、細胞培養担体３の片面に細胞を培養し、細胞培養面を上にした状態で細胞培養用カセット２０に挟み込んで細胞培養装置１０とし、この細胞培養装置１０を、別途培養した細胞が存在するシャーレ等の培養容器３０に入れることによって、細胞の機能解析及び細胞間の相互作用解析が可能である。   Further, as a cell culture method using the cell culture apparatus 10, cells are cultured on one side of the cell culture support 3, and the cell culture apparatus 10 is sandwiched between the cell culture cassettes 20 with the cell culture surface facing up. By inserting the cell culture device 10 into a culture container 30 such as a petri dish where separately cultured cells are present, it is possible to analyze the function of the cells and the interaction between the cells.

また、細胞培養装置１０を用いたさらなる細胞培養法として、細胞培養担体３の片面に細胞を培養し、細胞培養面を下にした状態で細胞培養用カセット２０に挟み込んで細胞培養装置１０とし、この細胞培養装置１０をシャーレ等の容器に入れ、さらに細胞培養担体３の細胞が培養されていない面に異なる細胞を播種することによって、簡便な両面培養が実現可能である。この培養モデルを用いて、細胞の機能解析及び細胞間の相互作用解析、さらには組織モデルの作製が可能である。   Further, as a further cell culture method using the cell culture device 10, cells are cultured on one side of the cell culture carrier 3, and the cell culture device 10 is sandwiched between the cell culture cassettes 20 with the cell culture surface facing down. A simple double-sided culture can be realized by placing the cell culture device 10 in a container such as a petri dish and further seeding different cells on the surface of the cell culture carrier 3 where the cells are not cultured. Using this culture model, it is possible to analyze the function of cells, analyze the interaction between cells, and create a tissue model.

上記細胞培養後、細胞培養担体３を細胞培養用カセット２０から外し、固定・染色・観察等の解析に供することができる。また、種々の処理が行われた細胞培養モデルをそのまま移植に供することも可能である。なお、ここでいう解析とは、細胞間相互作用、細胞接着、細胞分化、物質輸送・拡散・分泌、細胞極性といった現象の解析である。ただしこれらの使用態様は、本発明の使用範囲をなんら限定するものではない。   After the cell culture, the cell culture carrier 3 can be removed from the cell culture cassette 20 and subjected to analysis such as fixation, staining, and observation. In addition, a cell culture model subjected to various treatments can be directly used for transplantation. The analysis here is an analysis of phenomena such as cell-cell interaction, cell adhesion, cell differentiation, substance transport / diffusion / secretion, and cell polarity. However, these usage modes do not limit the usage range of the present invention.

（３）試験例
以下のようにして、本実施形態の細胞培養用カセット２０に細胞培養担体３を挟むことによって、上面と下面の培地が隔離されることを確認する試験を実施した。すなわち、直径３５ｍｍの円形に切り取ったポリエチレン膜（厚み：０．０４ｍｍ）を、歯科用ピンセットを用いて細胞培養用カセット２０を構成する外側筒状部材１のフランジ状の突起１１上に載置する。 (3) Test Example A test for confirming that the medium on the upper surface and the lower surface was isolated by sandwiching the cell culture carrier 3 between the cell culture cassettes 20 of the present embodiment was performed as follows. That is, a polyethylene film (thickness: 0.04 mm) cut into a circle having a diameter of 35 mm is placed on the flange-shaped protrusion 11 of the outer tubular member 1 constituting the cell culture cassette 20 using dental tweezers. .

次に、内側筒状部材２の鍔状の突出部２４をピンセットで掴み、外側筒状部材１の上方から内部に挿入する。その後、外側筒状部材１と内側筒状部材２の鍔状の突出部１４、２４の孔１５、２５にそれぞれピンセットを差し込み、外側筒状部材１及び内側筒状部材２をそれぞれ逆方向に回転させることにより、ポリエチレン膜を細胞培養用カセット２０に設置する。   Next, the hook-like projecting portion 24 of the inner cylindrical member 2 is grasped with tweezers and inserted into the outer cylindrical member 1 from above. Thereafter, tweezers are respectively inserted into the holes 15 and 25 of the flange-like protrusions 14 and 24 of the outer cylindrical member 1 and the inner cylindrical member 2, and the outer cylindrical member 1 and the inner cylindrical member 2 are rotated in opposite directions. By doing so, the polyethylene membrane is placed in the cell culture cassette 20.

その後、ポリエチレン膜を挟んだ細胞培養用カセット２０を、培養容器３０として用いられる６０ｍｍＤｉｓｈ（IWAKI Code#３０１０−０６０X）に入れ、細胞培養用カセット２０にフェノールレッド含有の水を入れ、培養容器３０に水を入れた状態で一昼夜放置した。その結果、培養容器３０に入れた水に、フェノールレッドの漏れ出しは確認されなかった。   Thereafter, the cell culture cassette 20 with the polyethylene membrane sandwiched between them is placed in a 60 mm dish (IWAKI Code # 3010-060X) used as the culture container 30, and phenol red-containing water is placed in the cell culture cassette 20. I left it for a day with water in it. As a result, no leakage of phenol red was confirmed in the water placed in the culture vessel 30.

（４）効果
本実施形態によれば、ピンセットまたは鉗子、もしくは前記着脱用治具等を使用して、細胞培養用カセット２０に滅菌状態で細胞培養担体３を挟み込んで細胞培養装置１０とし、この細胞培養装置１０をさらにシャーレ等の培養容器３０に挿入し、細胞培養またはその他解析終了後に、培養容器３０から細胞培養装置１０を取り出し、さらに細胞培養担体３を回収することが可能となる。 (4) Effect According to the present embodiment, the cell culture carrier 3 is sandwiched between the cell culture cassette 20 in a sterilized state using tweezers or forceps, or the attaching / detaching jig, etc. The cell culture device 10 is further inserted into a culture vessel 30 such as a petri dish, and after completion of cell culture or other analysis, the cell culture device 10 can be taken out from the culture vessel 30 and the cell culture carrier 3 can be further recovered.

このように、本発明の細胞培養装置は、外側筒状部材１と内側筒状部材２の間に細胞培養担体３を挟持し、この細胞培養担体３を着脱可能に構成したことにより、より簡便に細胞培養担体と細胞組織を回収することができ、また、上記の種々の培養方法により種々の処理を行った細胞組織を回収することも可能となる。   Thus, the cell culture device of the present invention is simpler by sandwiching the cell culture carrier 3 between the outer cylindrical member 1 and the inner cylindrical member 2 and detachably attaching the cell culture carrier 3. In addition, the cell culture carrier and the cell tissue can be recovered, and the cell tissue subjected to various treatments by the above-described various culture methods can also be recovered.

さらに、本発明の細胞培養装置を使用すれば、ハイドロゲル薄膜の片面に細胞を培養後に、細胞培養面を下にした状態で、細胞培養装置１０に挟持させ、さらに上方部分に異なる細胞を培養することで、効率的に両面培養を行うことも可能となる。   Furthermore, if the cell culture device of the present invention is used, after culturing the cells on one side of the hydrogel thin film, the cells are sandwiched by the cell culture device 10 with the cell culture surface down, and further different cells are cultured in the upper part. This makes it possible to perform double-sided culture efficiently.

（５）他の実施形態
本発明に係る細胞培養装置に使用される培養液は特に限定されない。培養液としては、例えば、ＤＭＥＭ、ＥＭＥＭ、ＲＰＭＩ１６４０等、培養細胞に適した種々の培養液を用いることができる。 (5) Other Embodiments The culture solution used for the cell culture device according to the present invention is not particularly limited. As the culture solution, for example, various culture solutions suitable for cultured cells such as DMEM, EMEM, RPMI1640, and the like can be used.

また、本発明は、上述したような実施形態に限定されるものではなく、各構成部材について、以下のような変形例にも適用することができる。   In addition, the present invention is not limited to the above-described embodiment, and can be applied to the following modified examples for each constituent member.

（５−１）筒状部材のフランジ状突起の形状
外側筒状部材１のフランジ状突起１１及び内側筒状部材２のフランジ状突起２１は、各筒状部材の下端部から内側に連設された、切れ目のないリング状に構成されていれば良く、その突起部分は平面でなくても良い。 (5-1) Shape of Flange-shaped Projection of Cylindrical Member The flange-shaped projection 11 of the outer tubular member 1 and the flange-shaped projection 21 of the inner tubular member 2 are connected inward from the lower end of each tubular member. In addition, it is only necessary to be configured in a ring shape without a break, and the protruding portion may not be a flat surface.

例えば、図５（Ａ）に示すように、内側筒状部材２のフランジ状突起２１を、下に凸となるような凹部２１ａとしても良い。この場合、内側筒状部材２のフランジ状突起２１と細胞培養担体３の支持体３１とは、面接触ではなく線接触となるが、線接触であっても支持体との間からの培養液のしみ出しや流入を防ぐことができる。   For example, as shown in FIG. 5A, the flange-like protrusion 21 of the inner cylindrical member 2 may be a recess 21a that protrudes downward. In this case, the flange-like projection 21 of the inner cylindrical member 2 and the support 31 of the cell culture carrier 3 are not in surface contact but in line contact, but even in line contact, the culture solution from between the support It can prevent exudation and inflow.

また、図５（Ｂ）に示すように、外側筒状部材１のフランジ状突起１１の先端部に凸部１１ｂを形成すると共に、内側筒状部材２のフランジ状突起２１の先端部にも、前記フランジ状突起１１の凸部１１ｂと嵌合する凸部２１ｂを形成しても良い。また、図５（Ｃ）に示すように、外側筒状部材１のフランジ状突起１１全体を凸部１１ｃとすると共に、内側筒状部材２のフランジ状突起２１も、前記フランジ状突起１１の凸部１１ｃと嵌合する凸部２１ｃとしても良い。   Further, as shown in FIG. 5 (B), a convex portion 11b is formed at the distal end portion of the flange-like projection 11 of the outer cylindrical member 1, and also at the distal end portion of the flange-like projection 21 of the inner cylindrical member 2. You may form the convex part 21b fitted with the convex part 11b of the said flange-shaped protrusion 11. FIG. As shown in FIG. 5C, the entire flange-shaped protrusion 11 of the outer cylindrical member 1 is a convex portion 11c, and the flange-shaped protrusion 21 of the inner cylindrical member 2 is also a convex portion of the flange-shaped protrusion 11. It is good also as the convex part 21c fitted with the part 11c.

このように、外側筒状部材１のフランジ状突起１１及び内側筒状部材２のフランジ状突起２１の一部あるいは全部を、互いに嵌合する凹凸形状とすることによって、フランジ状突起１１、２１と細胞培養担体３の支持体３１との接触面積を増やすことができるので、支持体との間からの培養液のしみ出しや流入をより効果的に防ぐことができる。   In this way, by forming a part or all of the flange-like protrusion 11 of the outer tubular member 1 and the flange-like protrusion 21 of the inner tubular member 2 into an uneven shape that fits together, the flange-like protrusions 11, 21 and Since the contact area of the cell culture carrier 3 with the support 31 can be increased, it is possible to more effectively prevent the culture medium from seeping out and flowing in from the support.

（５−２）クッション材の形状
外側筒状部材１のフランジ状突起１１の上面、あるいは内側筒状部材２のフランジ状突起２１の下面に取り付けられるクッション材４０は、切れ目のないリング状とすることが好ましい。また、このクッション材４０は、図６（Ａ）に示したように、内側筒状部材２のフランジ状突起２１の下面と細胞培養担体３との間に設置しても良く、図６（Ｂ）に示したように、外側筒状部材１のフランジ状突起１１の上面と細胞培養担体３との間に設置しても良い。また、細胞培養担体３の上下に２枚設置しても良い。 (5-2) Shape of Cushion Material The cushion material 40 attached to the upper surface of the flange-shaped protrusion 11 of the outer cylindrical member 1 or the lower surface of the flange-shaped protrusion 21 of the inner cylindrical member 2 has a continuous ring shape. It is preferable. Further, as shown in FIG. 6 (A), the cushion material 40 may be installed between the lower surface of the flange-like projection 21 of the inner cylindrical member 2 and the cell culture carrier 3, and FIG. As shown in FIG. 5, it may be installed between the upper surface of the flange-shaped protrusion 11 of the outer cylindrical member 1 and the cell culture carrier 3. Further, two pieces may be installed above and below the cell culture carrier 3.

（５−３）細胞培養担体の支持体の有無
上述したように、図１に示した実施形態のように細胞培養担体３に支持体３１が設けられている場合には、外側筒状部材１のフランジ状突起１１に支持体付きの細胞培養担体３を乗せた後、内側筒状部材２を外側筒状部材１に嵌め込んでセットすれば良い。 (5-3) Presence / absence of support for cell culture carrier As described above, when the support 31 is provided on the cell culture carrier 3 as in the embodiment shown in FIG. After placing the cell culture carrier 3 with the support on the flange-shaped protrusion 11, the inner cylindrical member 2 may be fitted into the outer cylindrical member 1 and set.

一方、細胞培養担体３に支持体３１が設けられていない場合には、図７（Ａ）に示したように、内側筒状部材２のフランジ状突起２１及び筒状部分の外側面の下端部に、湿らせた細胞培養担体を貼り付けた後、内側筒状部材２を外側筒状部材１に嵌め込んでセットする。なお、この場合、細胞培養担体が外側筒状部材１及び内側筒状部材２の係合用ネジ部１３、２３まで達しないような位置に貼り付けることが好ましい。   On the other hand, when the cell culture carrier 3 is not provided with the support 31, as shown in FIG. 7A, the flange-like protrusion 21 of the inner cylindrical member 2 and the lower end portion of the outer surface of the cylindrical portion. After the wet cell culture carrier is pasted, the inner cylindrical member 2 is fitted into the outer cylindrical member 1 and set. In this case, it is preferable that the cell culture carrier is attached at a position where it does not reach the engaging screw portions 13 and 23 of the outer cylindrical member 1 and the inner cylindrical member 2.

また、図７（Ｂ）に示したように、外側筒状部材１のフランジ状突起１１及び筒状部分の内側面の下端部に、湿らせた細胞培養担体を貼り付けた後、内側筒状部材２を外側筒状部材１に嵌め込んでセットしても良い。なお、この場合も、細胞培養担体が外側筒状部材１及び内側筒状部材２の係合用ネジ部１３、２３まで達しないような位置に貼り付けることが好ましい。   Moreover, as shown in FIG. 7 (B), after attaching a wet cell culture carrier to the flange-like protrusion 11 of the outer cylindrical member 1 and the lower end portion of the inner side surface of the cylindrical portion, the inner cylindrical shape is attached. The member 2 may be set by being fitted to the outer cylindrical member 1. In this case as well, it is preferable that the cell culture carrier is attached at a position where it does not reach the engaging screw portions 13 and 23 of the outer cylindrical member 1 and the inner cylindrical member 2.

（５−４）挟持される細胞培養担体の数
細胞培養用カセットを構成する外側筒状部材１及び内側筒状部材２に挟持される細胞培養担体３は、図８（Ａ）に示すように１枚でも良く、また、図８（Ｂ）に示すように２枚でも良い。 (5-4) Number of sandwiched cell culture carriers As shown in FIG. 8A, the cell culture carrier 3 sandwiched between the outer cylindrical member 1 and the inner cylindrical member 2 constituting the cell culture cassette is as shown in FIG. One may be sufficient, and two may be sufficient as shown in FIG.8 (B).

すなわち、図８（Ｂ）に示すように、１枚の細胞培養担体３に１種類の細胞を培養後、それを２枚重ねることで両面培養を行うことができる。その後、各細胞について異なる方法で形態観察や機能評価を行うことができる。この方法は、例えば、片面細胞の増殖効率や、酵素活性、薬物代謝測定等、片方のみの値を知りたいときに有効である。   That is, as shown in FIG. 8B, double-sided culture can be performed by culturing one type of cell on one cell culture carrier 3 and then stacking two cells. Thereafter, morphological observation and functional evaluation can be performed for each cell by different methods. This method is effective, for example, when it is desired to know the value of only one side such as the proliferation efficiency of single-sided cells, enzyme activity, and drug metabolism measurement.

本発明に係る細胞培養装置の構成を示す分解斜視図である。It is a disassembled perspective view which shows the structure of the cell culture apparatus which concerns on this invention. 本発明に係る細胞培養装置の全体像を示す斜視図である。It is a perspective view which shows the whole image of the cell culture apparatus which concerns on this invention. 図２に示した細胞培養装置の側方断面図である。It is a sectional side view of the cell culture apparatus shown in FIG. 本発明に係る細胞培養装置を既存の培養容器内に設置した状態を示す斜視図である。It is a perspective view which shows the state which installed the cell culture apparatus based on this invention in the existing culture container. （Ａ）〜（Ｃ）は共に、筒状部材のフランジ状突起の形状の変形例を示す断面図である。(A)-(C) are sectional drawings which show the modification of the shape of the flange-shaped protrusion of a cylindrical member. （Ａ）〜（Ｃ）は共に、クッション材の設置例を示す断面図である。(A)-(C) are sectional drawings which show the example of installation of a cushioning material. （Ａ）（Ｂ）は共に、支持体がないときの細胞培養担体の設置例を示す断面図である。(A) (B) is sectional drawing which shows the example of installation of the cell culture carrier when there is no support body. （Ａ）（Ｂ）は共に、細胞培養方法の一例を示す断面図である。(A) (B) is sectional drawing which shows an example of a cell culture method.

符号の説明Explanation of symbols

１…外側筒状部材
２…内側筒状部材
３…細胞培養担体
１０…細胞培養装置
１１、２１…フランジ状の突起
１２…自立部材
１３、２３…係合用ネジ部
１４、２４…鍔状の突出部
１５、２５…孔
２０…細胞培養用カセット
３０…培養容器
３１…支持体
３２…ハイドロゲル薄膜
４０…クッション材 DESCRIPTION OF SYMBOLS 1 ... Outer cylindrical member 2 ... Inner cylindrical member 3 ... Cell culture support | carrier 10 ... Cell culture apparatus 11, 21 ... Flange-like protrusion 12 ... Self-supporting member 13, 23 ... Screw part for engagement 14, 24 ... Sponge-like protrusion Portions 15, 25 ... Hole 20 ... Cell culture cassette 30 ... Culture vessel 31 ... Support 32 ... Hydrogel thin film 40 ... Cushion material

Claims (11)

共に筒状部材の一端に内側に突出するフランジ状の突起が形成された外側筒状部材と内側筒状部材とからなり、前記内側筒状部材を前記外側筒状部材の他端から挿入して、外側筒状部材と内側筒状部材を嵌合することによって、前記フランジ状の突起同士が圧接状態となるように構成されていることを特徴とする細胞培養用カセット。   Both are composed of an outer cylindrical member and an inner cylindrical member in which a flange-like projection protruding inward is formed at one end of the cylindrical member, and the inner cylindrical member is inserted from the other end of the outer cylindrical member. A cassette for cell culture, wherein the flange-like projections are brought into a pressure contact state by fitting the outer cylindrical member and the inner cylindrical member. 共に筒状部材の一端に内側に突出するフランジ状の突起が形成された外側筒状部材と内側筒状部材とからなり、前記外側筒状部材と内側筒状部材の間に細胞培養担体を設置した後、前記内側筒状部材を前記外側筒状部材の他端から挿入して、外側筒状部材と内側筒状部材を嵌合することによって、前記外側筒状部材及び内側筒状部材のフランジ状の突起が前記細胞培養担体を挟持するように構成されていることを特徴とする細胞培養用カセット。   Both are composed of an outer cylindrical member and an inner cylindrical member in which a flange-like projection protruding inward is formed at one end of the cylindrical member, and a cell culture carrier is installed between the outer cylindrical member and the inner cylindrical member Then, the inner cylindrical member is inserted from the other end of the outer cylindrical member, and the outer cylindrical member and the inner cylindrical member are fitted to each other, whereby the outer cylindrical member and the flange of the inner cylindrical member are fitted. A cell culture cassette, wherein the cell-shaped protrusion is configured to sandwich the cell culture carrier. 前記細胞培養担体が、リング状の支持体上に細胞培養担体膜を形成して構成され、前記リング状の支持体部分を、前記外側筒状部材及び内側筒状部材のフランジ状の突起によって挟持するように構成されていることを特徴とする請求項２に記載の細胞培養用カセット。   The cell culture carrier is configured by forming a cell culture carrier film on a ring-shaped support, and the ring-shaped support portion is sandwiched between flange-shaped protrusions of the outer cylindrical member and the inner cylindrical member. The cell culture cassette according to claim 2, wherein the cassette for cell culture is configured as described above. 前記外側筒状部材の内側面及び内側筒状部材の外側面に、互いに係合するネジ部が形成されていることを特徴とする請求項１乃至請求項３のいずれか一に記載の細胞培養用カセット。   The cell culture according to any one of claims 1 to 3, wherein threaded portions that engage with each other are formed on an inner surface of the outer cylindrical member and an outer surface of the inner cylindrical member. Cassette. 前記外側筒状部材に形成されたフランジ状の突起の上面、及び内側筒状部材に形成されたフランジ状の突起の下面の少なくとも一方に、クッション材が取付けられていることを特徴とする請求項１乃至請求項４のいずれか一に記載の細胞培養用カセット。   The cushion material is attached to at least one of the upper surface of the flange-shaped projection formed on the outer cylindrical member and the lower surface of the flange-shaped projection formed on the inner cylindrical member. The cell culture cassette according to any one of claims 1 to 4. 前記外側筒状部材のフランジ状の突起が形成されていない端部、及び内側筒状部材のフランジ状の突起が形成されていない端部の少なくとも一方に、筒状部材の外方に延びる突出部が形成されていることを特徴とする請求項１乃至請求項５のいずれか一に記載の細胞培養用カセット。   A projecting portion extending outward of the tubular member at least one of an end portion of the outer tubular member where the flange-like projection is not formed and an end portion of the inner tubular member where the flange-like projection is not formed. The cell culture cassette according to any one of claims 1 to 5, wherein the cassette is formed. 前記突出部に、少なくとも一つの孔が形成されていることを特徴とする請求項６に記載の細胞培養用カセット。   The cassette for cell culture according to claim 6, wherein at least one hole is formed in the protruding portion. 前記外側筒状部材に形成されたフランジ状の突起の外面の少なくとも３箇所に、所定の高さを有する自立部材が形成されていることを特徴とする請求項１乃至請求項７のいずれか一に記載の細胞培養用カセット。   8. The self-standing member having a predetermined height is formed in at least three locations on the outer surface of the flange-shaped protrusion formed on the outer cylindrical member. A cassette for cell culture as described in 1. 請求項１乃至請求項８のいずれか一に記載の細胞培養用カセットの外側筒状部材と内側筒状部材の少なくとも一方を固定することができるように構成されていることを特徴とする細胞培養用カセット着脱用治具。   A cell culture comprising the outer cylindrical member and the inner cylindrical member of the cell culture cassette according to any one of claims 1 to 8, wherein at least one of the outer cylindrical member and the inner cylindrical member can be fixed. Cassette mounting / demounting jig. 支持体に取り付けられた細胞培養担体膜であって、片面への細胞培養、両面への細胞培養、及びタンパク質透過の少なくとも一つが可能な細胞培養担体膜を、請求項１乃至請求項８のいずれか一に記載の細胞培養用カセットの外側筒状部材と内側筒状部材の間に設置したことを特徴とする細胞培養装置。   A cell culture carrier membrane attached to a support, wherein the cell culture carrier membrane is capable of at least one of cell culture on one side, cell culture on both sides, and protein permeation. A cell culturing apparatus, which is installed between an outer cylindrical member and an inner cylindrical member of the cell culture cassette according to any one of the above. 前記支持体が、不織布から構成されていることを特徴とする請求項１０に記載の細胞培養装置。   The cell culture device according to claim 10, wherein the support is made of a nonwoven fabric.
JP2005370657A 2005-12-22 2005-12-22 Cassette for cell culture, tool for attaching and detaching cassette for cell culture and cell culture apparatus Pending JP2007167002A (en)

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