JP2010158214A - Culture vessel - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To prevent contact of once formed aggregates having a uniform size before transplantation to form a larger aggregate. <P>SOLUTION: The culture vessel 100 is provided with a membrane filter 3 for holding cells 5 and having a number of through-holes 3a to inhibit the passage of the cells 5 and allow the passage of a culture liquid, a partition wall 7 placed on the membrane filter 3 and forming a plurality of mutually separated recesses 25 to use the membrane filter 3 as a bottom face, a liquid storing part 9 placed above the membrane filter 3 and storing the culture liquid, and a lid member 11 vertically movable in the liquid storing part 9 to a lower limit almost contacting with the upper end of the partition wall 7. <P>COPYRIGHT: (C)2010,JPO&INPIT

Description

  The present invention relates to a culture vessel.

Conventionally, in order to culture seed cells into a spherical cell tissue of a uniform size, a culture vessel provided with regularly arranged cell culture cells is known (see, for example, Patent Document 1).
According to this culture container, the seed cells are aggregated into a cell cluster of a certain size, thereby improving the cell activity and providing a cell tissue body having a high therapeutic effect.

JP-A-2005-27598

  However, in the conventional culture container, a plurality of partitioned cell culture cells are open to the upper space, so that it is especially transported after being cultured into a spherical cell tissue of uniform size in each cell culture cell. When the spherical cell tissue comes out of the cell culture cell due to vibration or the like at times, there is an inconvenience that a larger aggregate is formed in contact with the spherical cell tissue cultured in another cell culture cell. . A large agglomerate cannot be passed through a thin portion such as an injection needle during injection using a syringe or the like, and is clogged.

  The present invention has been made in view of the above-described circumstances, and it is possible to prevent the aggregates of uniform size once generated from coming into contact with each other until transplantation to become larger aggregates. It aims at providing the culture vessel which can be performed.

In order to solve the above problems, the present invention employs the following means.
The present invention has a number of through-holes that restrict the passage of cells and allow the passage of a culture solution, and are arranged on the upper surface of the filter member and partitioned from each other. A partition part that forms a plurality of recesses having a filter member as a bottom surface, a liquid storage part that stores a culture solution above the filter member, and is provided so as to be movable in the vertical direction in the liquid storage part. A culture vessel is provided that includes a lid member that is lowered to a position that substantially contacts the upper end.

  According to the present invention, by storing the culture medium in the liquid storage part and placing the cells on the filter member, the cells are aggregated for each of the plurality of recesses formed by the partition wall part, and the activity of the cells is improved. Cells can be cultured as aggregates of uniform size for each recess.

  In addition, by lowering the lid member to a position where it almost contacts the upper end of the partition wall, the cells are trapped in each recess, and the cell aggregate moves between the recesses even if vibration or the like occurs during transportation. Can be prevented. As a result, it is possible to prevent the agglomerates of uniform size once generated from coming into contact with agglomerates in other recesses until becoming a larger agglomerate until transplantation.

In addition, when recovering cultured cells, the liquid that can pass through the through hole is supplied from the side opposite to the side where the cells of the filter member are placed, with the lid member raised. Can be separated from the filter member and easily recovered.
In addition, as a filter member, a membrane filter etc. are mentioned, for example.

In the said invention, the aperture diameter of the said through-hole is good also as being set to the range of about 0.4-8 micrometers.
By comprising in this way, only a liquid can be passed efficiently, hold | maintaining a cell on a filter member.

Moreover, in the said invention, it is good also as the aperture diameter of the said through-hole being set to the range of about 1-4 micrometers.
In the invention described above, the upper surface of the filter member and the surface of the partition wall may be subjected to a hydrophilic treatment.
By comprising in this way, it can suppress that a cell adheres to the upper surface of a filter member, or the surface of a partition part, and can produce | generate the aggregate of the cell of a desired magnitude | size.

Moreover, in the said invention, it is good also as providing the protective member holding the said filter member.
By comprising in this way, a filter member can be arrange | positioned stably.

  According to the present invention, there is an effect that it is possible to prevent the agglomerates of uniform size once generated from coming into contact with each other until transplantation to become larger agglomerates.

It is the schematic of the culture container which concerns on one Embodiment of this invention. It is the schematic which showed the state which lowered | hung the cover member of the culture container of FIG. It is the schematic which showed the state which raised the cover member of the culture container of FIG. It is the schematic which provided the protective filter in the culture container of FIG.

Hereinafter, a culture vessel 100 according to an embodiment of the present invention will be described with reference to the drawings.
As shown in FIG. 1, the culture container 100 according to the present embodiment includes a container body 1 that stores cells 5 and a culture solution. Examples of cells 5 to be cultured include ES cells, iPS cells, stem cells (derived from bone marrow, fat, muscle, nerves, liver, placenta, cord blood, peripheral blood), chondrocytes, hepatocytes, bone cells, epithelial cells, cardiac muscle. Examples include cells, skeletal muscle cells, vascular endothelial cells, and nerve cells.

  The container body 1 includes a membrane filter (filter member) 3 on which cells 5 are placed, a plurality of partition walls (partition walls) 7 disposed on the upper surface of the membrane filter 3 and extending vertically upward, and a culture solution above the membrane filter 3. The liquid storage part 9 which stores liquid and the lid member 11 which seals the liquid storage part 9 are provided.

  The membrane filter 3 is for holding the cells 5, and has a large number of through holes 3a that restrict passage of the cells 5 and allow passage of the culture solution. For example, the through hole 3a has a diameter of about 0.4 to 8 μm, more preferably about 1 to 4 μm.

  Further, the membrane filter 3 is provided with a predetermined gap from the bottom surface 1a of the container body 1, and is disposed so as to divide the liquid reservoir 9 vertically. Further, the upper surface of the membrane filter 3, that is, the surface on which the cells 5 are placed is subjected to hydrophilic treatment.

  The partition wall 7 defines a space above the membrane filter 3 and forms a plurality of recesses 25 having the membrane filter 3 as a bottom surface. Between the partition walls 7, a plurality of columnar stoppers 17 having a height substantially equal to that of the partition walls 7 are arranged at intervals. The surfaces of the partition wall 7 and the stopper 17 are subjected to hydrophilic treatment in the same manner as the upper surface of the membrane filter 3.

  Above the membrane filter 3 of the liquid reservoir 9, an openable and closable liquid inlet 13 and a liquid outlet 15 serving as a liquid inlet / outlet are provided. The liquid inlet 13 and the liquid outlet 15 are arranged at a position higher than the stopper 17. Further, a recovery solution inlet 18 for supplying a recovery solution for recovering the cells 5 and a waste liquid port 19 for discharging the culture solution and the like are provided below the membrane filter 3 of the liquid reservoir 9 so as to be openable and closable. It has been.

  The lid member 11 constitutes the ceiling of the liquid storage unit 9 and has a flat ceiling surface 11a. The lid member 11 is provided so as to be movable in the vertical direction in the liquid storage portion 9 and can be lowered to a position where it comes into contact with the stopper 17, that is, to a position where it substantially contacts the upper end of the partition wall 7.

Next, the operation of the culture vessel 100 according to the present embodiment configured as described above will be described.
In order to culture the cells 5 using the culture vessel 100, first, a cell suspension containing the cells 5 is injected from the liquid inlet 13 with the lid member 11 raised, and the cells 5 are placed on the membrane filter 3. Sowing. It is desirable to inject the cells 5 so as to be distributed almost evenly into the respective recesses 25 defined by the partition walls 7.

  Subsequently, the culture solution is injected from the liquid inlet 13, and the culture solution is stored in the liquid storage unit 9. When the culture solution is stored in the liquid storage unit 9, the lid member 11 is lowered and abutted against the stopper 17 as shown in FIG. 2. At this time, the culture solution on the membrane filter 3 is pushed out in the direction passing through the membrane filter 3 by the ceiling surface 11 a of the lid member 11 and discharged from the waste liquid port 19. In this state, the culture vessel 100 is stored or transported to culture the cells 5.

  Since the upper surface of the membrane filter 3 and the surfaces of the partition walls 7 and the stoppers 17 are subjected to hydrophilic treatment, it is possible to prevent the cells 5 from adhering to the upper surface of the membrane filter 3 and the surfaces of the partition walls 7 and the stoppers 17. it can. Thereby, the activity of the cells 5 can be improved by efficiently aggregating the cells 5 for each of the plurality of recesses 25. And the aggregate 50 of the cell 5 of the uniform magnitude | size can be produced | generated for every recessed part 25. FIG.

  Moreover, the cell 5 is confined for every recessed part 25 by the ceiling surface 11a by descend | falling the cover member 11 to the position which contacts the upper end of the partition 7 substantially. Therefore, for example, it is possible to prevent the generated aggregate 50 of the cells 5 from moving between the recesses 25 even if vibration or the like occurs during transportation. Thereby, the cell 5 can be stably cultured as the aggregate 50 of a desired size.

  Next, in order to collect the cells 5 cultured on the membrane filter 3, first, the waste liquid port 19 is opened, and the culture solution stored in the liquid storage unit 9 is discharged. Then, as shown in FIG. 3, the lid member 11 is raised, and a recovered solution (liquid) that can pass through the through hole 3 a is supplied from the recovered solution inlet 18 and discharged from the liquid outlet 15.

  By supplying the recovery solution from below the membrane filter 3, the aggregate 50 of the cells 5 is peeled off from the membrane filter 3 by the recovery solution passing through the through holes 3a, and floats in the recovery solution. Therefore, the aggregate 50 of the cells 5 can be taken out from the liquid outlet 15 together with the recovered solution, and the cultured cells 5 can be easily recovered.

  As described above, according to the culturing device 100 according to the present embodiment, the cell 50 is confined for each recess 25 by the lid member 11, so that the aggregate 50 of the cells 5 moves between the recesses 25. Can be prevented. Thereby, it is possible to prevent the aggregate 50 having a uniform size once generated from coming into contact with the aggregate 50 in the other recess 25 and becoming a larger aggregate until transplantation. .

  In the present embodiment, the membrane filter 3 is disposed with a predetermined gap from the bottom surface 1a of the container body 1. For example, as shown in FIG. A flat protective frame (protective member) 21 may be provided, and the membrane filter 3 may be disposed on the protective frame 21. In this case, the protective frame 21 may be formed with a plurality of through holes 21a penetrating in the thickness direction at predetermined intervals. By doing in this way, the membrane filter 3 can be stably hold | maintained, enabling distribution | circulation of a culture solution and a collection | recovery solution.

In the present embodiment, the height of the partition wall 7 and the stopper 17 has been described as being substantially equal. However, the agglomerate 50 of the cells 5 housed in the recess 25 in a state where the lid member 11 is lowered is another recess. For example, the stopper 17 may be set higher than the partition wall 7 in a range smaller than the size of the aggregate 50 of the cells 5.
Moreover, in this embodiment, although the membrane filter 3 was illustrated and demonstrated as a filter member, it is not limited to this, Many through-holes which restrict | limit passage of the cell 5 and permit passage of a culture solution are provided. What is necessary is just to have. For example, in place of the membrane filter 3, a non-woven fabric, a porous body (such as a sponge) or the like may be employed.

3 Membrane filter (filter member)
3a through-hole 5 cell 7 partition (partition wall)
9 Liquid storage part 11 Lid member 21 Protective frame (protective member)
25 Recess 100 Culture container

Claims (5)

A filter member for limiting the passage of cells and having a large number of through-holes allowing passage of the culture solution, and for placing the cells;
A partition wall portion that is disposed on the upper surface of the filter member and forms a plurality of recesses with the filter members partitioned from each other as a bottom surface;
A liquid storage section for storing a culture solution above the filter member;
A culture vessel comprising: a lid member which is provided so as to be movable in the vertical direction in the liquid storage part and is lowered to a position where it substantially contacts the upper end of the partition wall part.   The culture vessel according to claim 1, wherein the diameter of the through hole is set in a range of about 0.4 to 8 µm.   The culture container according to claim 2, wherein the diameter of the through hole is set in a range of about 1 to 4 μm.   The culture container according to any one of claims 1 to 3, wherein the upper surface of the filter member and the surface of the partition wall are subjected to a hydrophilic treatment.   The culture container in any one of Claims 1-4 provided with the protection member holding the said filter member.

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