JP2004089136A - Culture vessel - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To keep cultural environment of a mesenchymal stem cell by a simple vessel and efficiently proliferate the cell. <P>SOLUTION: A cultural vessel 1 cultures a cell growing along a bottom surface 2 and can enlarge the area of the bottom surface 2 growing the cell. <P>COPYRIGHT: (C)2004,JPO

Description

[0001]
TECHNICAL FIELD OF THE INVENTION
TECHNICAL FIELD The present invention relates to a culture vessel for culturing mesenchymal stem cells and the like growing along a bottom surface.
[0002]
[Prior art]
Conventionally, stem cells such as mesenchymal stem cells have been known as cells capable of differentiating into various tissues and regenerating those tissues. Mesenchymal stem cells are contained in bone marrow fluid. However, only a very small amount of mesenchymal stem cells can be collected from the bone marrow fluid, and in order to obtain the amount of mesenchymal stem cells required for tissue regeneration, it is necessary to proliferate by culturing the bone marrow fluid.
[0003]
Conventionally, to culture mesenchymal stem cells, bone marrow fluid collected from a patient is inoculated on a flat culture vessel and cultured in an appropriate medium. Hematopoietic cells such as red blood cells and white blood cells in the bone marrow fluid float in the medium, while mesenchymal stem cells adhere to the bottom of the culture vessel and proliferate. Therefore, by discarding the hematopoietic cells by replacing the medium, it becomes possible to extract only the mesenchymal stem cells that have adhered and proliferated to the bottom surface of the culture vessel.
[0004]
[Patent Document 1]
Japanese Patent Publication No. 3-69508 (Page 1, Figure 1)
[Patent Document 2]
Japanese Patent Publication No. 3-57744 (page 7, FIG. 4)
[0005]
[Problems to be solved by the invention]
However, the growth rate of mesenchymal stem cells that adhere to and grow on the bottom surface of the culture vessel in this way tends to be influenced by the spatial culture environment in which they are placed. In other words, the mesenchymal stem cells adhere to the bottom of the culture vessel and proliferate, so it goes without saying that a bottom area that is large enough to attach is necessary, but if it is too wide, the proliferation rate decreases, There is a disadvantage that it cannot be grown efficiently.
[0006]
In order to avoid such inconveniences, it is considered that an appropriate culture environment for mesenchymal stem cells is maintained by sequentially transferring to a culture container having a large volume according to the degree of proliferation of the mesenchymal stem cells. I have.
However, the work of transferring to a plurality of culture vessels is complicated, and when this is automatically performed, means for moving cells and culture media between the culture vessels is required, and a large number of vessels are stored. A space is required, and there is a disadvantage that the device becomes large. Further, for culturing mesenchymal stem cells, different culture vessels are used for each patient, and the culture vessels that have been used once are not reused. Therefore, if a large number of culture vessels are used for culturing mesenchymal stem cells in one patient, there is a disadvantage that waste is increased and resources are wasted.
Also, an apparatus for automatically culturing cells has been proposed (for example, see Patent Documents 1 and 2).
[0007]
The present invention has been made in view of the above-described circumstances, and has as its object to provide a culture container that can maintain a culture environment of mesenchymal stem cells and grow it efficiently with a single container. And
[0008]
[Means for Solving the Problems]
In order to achieve the above object, the present invention provides the following means.
The invention according to claim 1 is a culture container for culturing cells growing along the bottom surface, and provides a culture container capable of expanding the area of the bottom surface for growing cells.
According to the present invention, by increasing the area of the bottom surface in accordance with the growth of the cells, it becomes possible to grow the culture efficiently in a single container.
[0009]
According to a second aspect of the present invention, there is provided the culture vessel according to the first aspect, wherein the vessel wall constituting the bottom surface can be extended.
According to the present invention, in accordance with the degree of cell growth, for example, external force, heat or the like is applied to the container to extend the container wall constituting the bottom surface, thereby expanding the bottom area, thereby increasing the efficiency of cells. It is possible to secure the bottom area necessary for growth.
[0010]
According to a third aspect of the present invention, there is provided the culture vessel according to the first aspect, wherein the culture vessel has a plurality of inner surfaces that are arranged at an angle to each other and have different areas each of which can be a bottom surface.
According to the present invention, it is possible to change the area of the inner surface serving as the bottom surface from a small one to a large one by changing the angle of the culture vessel according to the degree of cell growth. As a result, the bottom area can be increased to achieve efficient cell growth.
[0011]
According to a fourth aspect of the present invention, in the culture vessel according to the first aspect, a step capable of increasing a bottom area immersed in the culture solution by increasing an amount of the culture solution stored therein is provided on the bottom surface. A culture vessel is provided.
According to the present invention, by increasing the amount of the culture solution as the cell growth progresses, the culture solution is stored over the step provided on the bottom surface, and as a result, the bottom area contributing to the cell culture Can be increased.
[0012]
The invention according to claim 5 is the culture vessel according to claim 1, further comprising at least two container pieces provided so as to be relatively movable, and maintaining a gap between the adjacent container pieces in a sealed state. The present invention provides a culture container provided with a sealing means.
According to this invention, the bottom area can be increased by disposing the bottom surface along the relative movement direction of the two container pieces and relatively moving the positions of the two container pieces. At this time, since the gap is maintained in a sealed state by the sealing means disposed between the container pieces, the leakage of the culture solution inside is prevented.
[0013]
The invention according to claim 6 is the culture vessel according to claim 5, wherein the container piece comprises a cylinder and a piston fitted to the cylinder, and the sealing means is provided between the cylinder and the piston. Provided is a culture vessel which is a seal member to be arranged.
According to the present invention, the internal volume of the container can be increased by moving the piston fitted to the cylinder in the cylinder. Therefore, by disposing the bottom surface along the movement direction of the piston, the bottom area can be enlarged by the movement of the piston. In addition, the sealing member disposed between the cylinder and the piston prevents the culture solution or the like from leaking from the inside of the culture vessel.
[0014]
BEST MODE FOR CARRYING OUT THE INVENTION
Before describing the culture vessel according to each embodiment of the present invention, a manufacturing process of a bone filling body as a living tissue filling body will be schematically described. In order to produce a bone filling, as shown in FIG. 15, first, bone marrow fluid is collected from a patient's iliac bone or the like. The collected bone marrow fluid is centrifuged and swirled to extract bone marrow cells having a high specific gravity.
[0015]
The extracted bone marrow cells are put into a culture vessel together with a previously prepared medium and mixed. A portion of the medium is removed and sent for infection testing.
Thereafter, the mixed bone marrow fluid and culture medium are maintained under culture conditions such as a predetermined temperature (for example, 37 ± 0.5 ° C.) and a CO ₂ concentration (for example, 5%), so that constant culture conditions are maintained for a predetermined time. The cells are primarily cultured in. At a predetermined exchange time during the culturing of the cells, the medium is discarded from the inside of the culture vessel. Then, the medium is mixed again, and the culturing step is repeated and continued. A portion of the discarded medium is sent for infection testing.
[0016]
After a predetermined culture period is over, after the medium is discarded from the culture vessel, a protease such as trypsin is charged and mixed into the culture vessel. As a result, the mesenchymal stem cells attached to and grown on the bottom surface of the culture container are detached from the bottom surface of the main culture container. The mesenchymal stem cells thus detached are extracted by being subjected to a centrifugal separator.
[0017]
The extracted mesenchymal stem cells are mixed in a culture vessel in which a bone filling material and an appropriate medium have been charged after the cell number is adjusted. In practice, mesenchymal stem cells are attached to a bone substitute and injected into a medium. Then, in the same manner as described above, the mixed mesenchymal stem cells and the medium are maintained at culture conditions such as a predetermined temperature (for example, 37 ± 0.5 ° C.) and a CO ₂ concentration (for example, 5%). The cells are subcultured under a constant culture condition for a predetermined time.
[0018]
In the secondary culture step as well, as in the primary culture step, the medium is changed periodically, and a part of the medium to be supplied and a part of the medium to be discarded are each sent for an infection test. Then, after a lapse of a predetermined culture period, sample extraction for quality inspection and infection inspection for shipping is performed, and the manufactured bone replacement material is sealed and provided as a product.
[0019]
The culture vessel according to each embodiment of the present invention described below is mainly used in the primary culture step among the culture steps described above, but may be used in the secondary culture step.
As shown in FIG. 1, the culture vessel 1 according to the present embodiment is made of a stretchable polymer film (for example, polystyrene), and is provided with an external force to apply the external force (in FIG. 1). The container wall itself constituting the bottom surface 2 can be extended in the direction of the arrow).
[0020]
The operation of the thus-configured culture vessel 1 according to the present embodiment will be described below.
When primary culture of mesenchymal stem cells is performed using the culture vessel 1 according to the present embodiment, at the beginning of the start of culture of mesenchymal stem cells, a relatively narrow line as shown by a solid line in FIGS. 1 and 2. The culture is performed for a predetermined period while maintaining the state having the bottom surface 2. Next, when the growth of the mesenchymal stem cells progresses and the space for growing the mesenchymal stem cells in the culture vessel 1 in the state shown by the solid line becomes narrow, an external force is applied to the culture vessel 1 and the culture vessel 1 is shown by the dashed line. Extend to the position.
This makes it possible to prepare a comfortable spatial culture environment that is neither too wide nor too narrow for mesenchymal stem cells, and allows mesenchymal stem cells to grow efficiently.
[0021]
Instead of the above-described extensible culture vessel 1, for example, an elastically deformable tubular culture vessel 3 as shown in FIG. 3 and a clip 4 for squeezing the culture vessel 3 at an arbitrary position. And may be adopted. The inner volume of the culture vessel 3 may be increased by shifting the position of the clip 4 in accordance with the growth of the cells.
[0022]
Next, a culture vessel 10 according to a second embodiment of the present invention will be described below with reference to FIGS.
The culture vessel 10 according to the present embodiment includes two vessel pieces 11 and 12 that are slidably combined with each other, and a seal member 13 that is disposed therebetween.
[0023]
Each of the container pieces 11 and 12 has a rectangular bottom wall 11a, 12a and three side walls 11b, 12b rising from three sides of the bottom wall 11a, 12a. It is a box-shaped member that opens. Then, the other container piece 12 is formed inside the opposing side wall portion 11b of the one container piece 11 so as to be fitted into the other container piece 12 exactly. On the inner surface of the outer container 11 piece, a guide groove 11c formed parallel to the bottom surface 11a on the opposing side wall 11b, and a seal groove 11d formed continuously with the side wall 11b and the bottom wall 11a. It has. A projection 12c is formed on the outer surface of the inner container piece 12 so as to be slidably inserted into the guide groove 11c. Further, the seal member 13 is disposed in the seal groove 11 d, and at any relative position between the two container pieces 11, 12, the inner surface of the seal groove 11 d of the outer container piece 11 and the outer surface of the inner container piece 12 are formed. , So that the inside culture solution does not leak from between the two.
[0024]
According to the culture container 10 according to the present embodiment configured as described above, at the initial stage of the start of the culture of the mesenchymal stem cells, the container pieces 11 and 12 are relatively contracted as shown by solid lines in FIGS. 4 and 5. Culture is performed for a predetermined period while maintaining the positional relationship. Then, when the growth of the mesenchymal stem cells progresses and the space for growing the mesenchymal stem cells in the culture vessel in the state shown by the solid line becomes narrow, an external force is applied to the container pieces 11 and 12 of the culture vessel 10. Then, by sliding the projection 12c of the outer container piece 12 along the guide groove 11c on the inner surface of the outer container piece 11, the two container pieces 11, 12 are relatively moved to the position indicated by the chain line.
[0025]
Thereby, similarly to the culture container 1 according to the first embodiment, it is possible to prepare a comfortable spatial culture environment that is not too wide and not too narrow for the mesenchymal stem cells. It can be grown efficiently. Moreover, according to the culture container 10 according to the present embodiment, the bottom area of the culture container 10 can be easily increased only by sliding the other container piece 12 with respect to the one container piece 11. Therefore, the bottom area of the culture vessel 10 can be finely adjusted according to the cell growth status, and more efficient culture of mesenchymal stem cells can be performed.
[0026]
Next, a culture vessel 20 according to a third embodiment of the present invention will be described below with reference to FIG.
The culture container 20 according to the present embodiment can increase the bottom area of the container 20 by slidably combining the two container pieces 21 and 22 similarly to the culture container 10 according to the second embodiment. It is what was constituted. Specifically, as shown in FIG. 6, the culture vessel 20 according to the present embodiment includes a cylindrical cylinder 21, a piston 22 fitted in the cylinder 21, and the cylinder 21 and the piston 22. And a sealing member 23 for sealing the space therebetween.
In the figure, reference numeral 24 denotes a lid for closing the culture medium supply port 25 provided in the cylinder 21, reference numeral 26 denotes a seal groove for attaching the ring-shaped seal member 23, and reference numeral 27 denotes a piston connected to a driving unit (not shown). Shown is a rod for driving 22.
[0027]
According to the culture vessel 20 according to the present embodiment configured as described above, in the initial stage of the culture start, culture is performed for a predetermined period with the piston 22 arranged at the position shown by the solid line in the figure with respect to the cylinder 21. As the culture proceeds, the size of the bottom area can be increased by pulling out the piston 22 as shown by a chain line. With the configuration including the cylinder 21 and the piston 22, the sealing by the seal member 23 can be easily performed.
[0028]
Next, a culture vessel 30 according to a fourth embodiment of the present invention will be described below with reference to FIG.
As shown in FIG. 7, the culture vessel 30 according to the present embodiment has a plurality of steps 31, 32, and 33 at the bottom. In the example shown in FIG. 7, a step is formed such that the height gradually increases from the center toward the outer periphery.
[0029]
The case where the mesenchymal stem cells are cultured using the culture vessel 30 according to the present embodiment configured as described above will be described below.
First, at the beginning of the culture start, culture is performed by supplying a culture solution up to level A so that only the lowermost bottom surface indicated by reference numeral 34 in the figure is immersed. As a result, the mesenchymal stem cells proliferate along the bottom surface 34 until the bottom surface 34 feels narrow.
[0030]
Next, in a state where the mesenchymal stem cells have proliferated to some extent, culture is performed by supplying a culture solution to level B so as to be immersed to the second bottom surface 35 from the bottom. Thus, the bottom area in which the mesenchymal stem cells can proliferate is the sum of the bottom areas of the bottom surfaces 34 and 35. Therefore, the mesenchymal stem cells are provided with a newly proliferable bottom surface 35, so that proliferation can be advanced. By repeating this operation up to levels C and D, the bottom area can be increased by three steps up to the bottom surfaces 36 and 37, and efficient growth of mesenchymal stem cells can be promoted.
[0031]
In the culture container according to the present embodiment, by increasing the amount of medium to be stored on the bottom surface having the plurality of steps 31, 32, and 33, the bottom area in which cells can proliferate is increased. However, instead of this, as shown in FIG. 8, the bottom surfaces 34, 35, 36 constituting the steps 31, 32, 33 are configured to be able to move up and down, and the culture of the cells on the bottom surface 34 at the lowest position is performed. At the end, the bottom area 34 required for cell culture may be increased by raising the lowest bottom face 34 to the height of the bottom face 35 one step above. In this case, means for raising and lowering the bottom surfaces 34, 35, 36 may be any mechanism such as a piston, a motor, or the like.
[0032]
Further, in the culture vessel according to the present embodiment, a plurality of steps are provided at one place on the bottom surface of the culture vessel. Instead, as shown in FIGS. 9 and 10, the steps are formed one step lower. A plurality of bottom surfaces 38 may be provided at intervals. According to this, an appropriate culture environment can be realized at a plurality of locations in the culture vessel at the initial stage of the culture, so that the efficiency of the culture can be improved.
[0033]
Also, instead of the step extending in the vertical direction as shown in FIG. 7, as shown in FIG. 10, the bottom surfaces 37 and 38 with height differences are connected by a gentle slope 39. Is also good. Thus, the cells that have grown on the lower bottom surface 38 at the beginning of the culture can climb and grow to the upper bottom surface 37 in the next culture stage.
In addition, as shown in FIG. 11, the lower bottom surface may be a relatively narrow groove-shaped bottom surface 38, which may be formed continuously over the entire bottom surface of the culture vessel. Thus, even if the cells are introduced in the initial stage of the culture at one location of the groove-shaped bottom surface 38, the culture can be performed on the entire bottom surface 38, so that the processing is easy.
[0034]
Next, a culture vessel 40 according to a fifth embodiment of the present invention will be described below with reference to FIG.
As shown in FIG. 12, the culture container 40 according to the present embodiment includes a container main body 41 having a flat bottom surface, and a cylindrical container wall member 42 placed on the bottom surface of the container main body 41. The container wall member 42 is, for example, a cylindrical body formed in a cylindrical shape, and one end thereof is formed flat so as to be able to be in close contact with the bottom surface of the container main body.
[0035]
In order to culture mesenchymal stem cells using the culture container 40 according to the present embodiment configured as described above, first, a cylindrical container wall member 42 is placed on the bottom surface of the container body 41 and The space inside is divided into two. Then, the storage and the cells are put into the container wall member 42 and placed under predetermined culture conditions. Thereby, the mesenchymal stem cells in the container wall member 42 are not too wide, and can efficiently grow along the appropriately narrow bottom surface 43a.
[0036]
Next, after a predetermined culture period has elapsed, the compartment in the container body 41 is removed by removing the container wall member 42 from the inside of the container body 41. As a result, the mesenchymal stem cells can obtain a wider bottom surface 43, and can be efficiently proliferated in a comfortable culture environment.
[0037]
In the culture container 40 according to the present embodiment, a seal member is provided at the lower end of the container wall member 42 to seal the inside and outside of the container wall member 42 when placed on the bottom surface 43 of the container body 41. Is also good. According to this, in the initial stage of the culture, the culture is performed using only the medium inside the container wall member 42, and by providing the container wall member 42, a new medium outside the container wall member 42 is provided to the cells. Can be.
[0038]
Alternatively, a plurality of container wall members 42 may be arranged in the container body 41, and cells may be cultured at a plurality of locations. In addition, as shown by a chain line in FIG. 12, by surrounding the container wall member 42 with one or more other container wall members having different sizes, and sequentially removing the container wall member 42 from the inner container wall member 42, The area of the bottom surface on which the culture is performed may be gradually increased.
[0039]
Next, a culture vessel 50 according to a sixth embodiment of the present invention will be described below with reference to FIG.
The culture container 50 according to the present embodiment has, for example, a plurality of wall surfaces 51, 52, 53 adjacent to each other at an angle as shown in FIG. The areas of the wall surfaces 51, 52, 53 are different from each other. For example, the area of the second wall 52 is larger than that of the first wall 51, and the area of the third wall 53 is larger than the area of the second wall 52. These first to third wall surfaces 51, 52, 53 are arranged at positions away from the medium supply port 54 provided in the culture vessel 50, and are all arranged horizontally at the lowest level so that the culture vessel 50 can be configured.
[0040]
The case where the mesenchymal stem cells are cultured using the culture container 50 according to the present embodiment configured as described above will be described below.
First, in the initial stage of the start of culture, as shown in FIG. 13A, culture is performed with the narrowest first wall surface 51 horizontally arranged at the lowest position. Thereby, the mesenchymal stem cells in the culture solution can be efficiently expanded along the appropriately narrow bottom surface 51 without being too wide.
[0041]
Next, after the culturing has progressed for a predetermined culturing period, as shown in FIG. 13B, the culturing is performed with the second wall surface 52 arranged horizontally at the lowest position. Since the second wall surface 52 has a larger area than the first wall surface 51, efficient growth of mesenchymal stem cells proceeds. Further, after culturing for a predetermined culturing period, as shown in FIG. 13 (c), culturing is performed with the third wall surface 53 arranged horizontally at the lowest position. Thereby, the efficient proliferation of the mesenchymal stem cells is continued.
[0042]
As described above, according to the culture container 50 according to the present embodiment, the culture environment suitable for the proliferation of mesenchymal stem cells can be obtained only by rotating the culture container 50 and selecting the wall surfaces 51, 52, and 53 to be the bottom surfaces. Therefore, efficient culture can be performed extremely easily. Therefore, a required amount of mesenchymal stem cells can be rapidly produced.
[0043]
In addition, in the said embodiment, although the culture container 50 which made the planar wall surface which differs in area adjacent was demonstrated as an example, as shown in FIG. It may be adopted. In this way, as shown in FIGS. 14A to 14C, the area of the bottom surface immersed in the culture medium can be increased by changing the inclination angle of the culture vessel 54.
[0044]
In each of the above embodiments, only the culture step of culturing mesenchymal stem cells while increasing the bottom area has been described, but actually, the bottom surface 2, 11a, 12a, 34, 35, 36, 37 is described. , 51, 52, 53, a step of detaching mesenchymal stem cells growing in a state of being attached to the bottom surfaces 2, 11a, 12a, 34, 35, 36, 37, 51, 52, 53, and a step of periodically culturing the culture solution. It is necessary that the step of exchanging to be performed during the culture step.
[0045]
The step of detaching the mesenchymal stem cells from the bottom surface of the culture vessels 1, 10, 20, 30, 40, 50, 60 is performed by, for example, using a protease such as trypsin in the culture vessels 1, 10, 20, 30, 40, 60. It is carried out by putting it into 50 and 60. In this case, for example, a centrifugation step is required to extract the mesenchymal stem cells mixed in the trypsin. Instead, the inside surfaces of the culture vessels 1, 10, 20, 30, 40, 50, and 60 may be subjected to a temperature responsive process of switching between hydrophobicity and hydrophilicity at a predetermined temperature as a boundary.
[0046]
The temperature-responsive treatment is performed by immobilizing a temperature-responsive polymer poly (N-isopropylacrylamide) with a covalent bond. The region subjected to the temperature responsive treatment has a boundary temperature of 32 ° C., above which a weak hydrophobicity similar to that of a commercially available cell culture vessel is exhibited, but a high hydrophilicity is obtained by cooling the temperature below the boundary temperature. Is an area which comes to exhibit. Therefore, for example, by culturing at 37 ° C. and then cooling to 32 ° C. or lower, the inner surfaces of the culture vessels 1, 10, 20, 30, 40, 50, and 60 are changed to exhibit high hydrophilicity, and easily mesenchyme. Lineage stem cells can be detached.
In this way, the mesenchymal stem cells can be expanded in a single culture vessel 1, 10, 20, 30, 40, 50, 60 only by replacing the culture solution without performing a trypsin treatment or a centrifugation step. You can keep it going.
[0047]
In each of the above embodiments, the open-type and sealed-type containers are separately illustrated and described. However, the present invention is not limited to this, and both types of containers may be adopted in each embodiment. it can.
The number and height of the steps on the bottom surface of the culture vessel 30 in the fourth embodiment, and the numbers and sizes of the wall surfaces 51, 52, and 53 of the culture vessel 50 in the sixth embodiment are not particularly limited. Needless to say, it can be set appropriately.
[0048]
Further, in each of the above-described embodiments, bone was used as an example of a biological tissue, and a case where mesenchymal stem cells extracted from bone marrow fluid were cultured was described. It may be. The cells are not limited to mesenchymal stem cells, and can be used for culturing somatic cells such as ES cells, somatic stem cells, bone cells, chondrocytes, and nerve cells.
[0049]
【The invention's effect】
As described above, according to the culture container according to the present invention, the cells grown in a single container can provide a bottom surface having a bottom area that gradually increases with growth, This provides an effect that a comfortable culture environment is provided to the cells to promote efficient growth, and a required amount of cells can be obtained quickly.
[Brief description of the drawings]
FIG. 1 is a perspective view showing a culture vessel according to a first embodiment of the present invention.
FIG. 2 is a longitudinal sectional view showing the culture vessel of FIG.
FIG. 3 is a longitudinal sectional view showing a modified example of the culture vessel of FIG.
FIG. 4 is a perspective view showing a culture vessel according to a second embodiment of the present invention.
FIG. 5 is a longitudinal sectional view showing the culture vessel of FIG.
FIG. 6 is a longitudinal sectional view showing a culture container according to a third embodiment of the present invention.
FIG. 7 is a longitudinal sectional view showing a culture vessel according to a fourth embodiment of the present invention.
8 is a longitudinal sectional view showing a modification of the culture vessel of FIG.
FIG. 9 is a plan view showing another modified example of the culture container of FIG. 7;
FIG. 10 is a longitudinal sectional view showing the culture container of FIG. 9;
FIG. 11 is a plan view showing another modification of the culture vessel of FIG.
FIG. 12 is a longitudinal sectional view showing a culture vessel according to a fifth embodiment of the present invention.
FIG. 13 is a longitudinal sectional view showing a culture vessel according to a sixth embodiment of the present invention, divided for each culture state.
FIG. 14 is a longitudinal sectional view showing a modification of the culture vessel of FIG.
FIG. 15 is a schematic diagram illustrating a culture step in which a culture vessel according to each embodiment of the present invention is used.
[Explanation of symbols]
1, 10, 20, 30, 40, 50, 60 Culture vessels 2, 11a, 12a, 34, 35, 36, 37 Bottom surface 11, 12 Container pieces 13, 23 Seal member (sealing means)
21 cylinders (container pieces)
22 piston (container piece)
39 Inclined surface 41 Container main body 42 Container wall members 51, 52, 53 Wall surfaces (bottom surface, inner surface)

Claims (10)

A culture vessel for culturing cells growing along the bottom surface,
A culture vessel that can expand the area of the bottom surface for growing cells. The culture container according to claim 1, wherein the container wall constituting the bottom surface can be extended. The culture vessel according to claim 1, comprising a plurality of inner surfaces that are arranged at an angle to each other and have different areas each of which can be a bottom surface. The culture vessel according to claim 1, wherein a step is provided on the bottom surface of the bottom surface, the bottom area being immersed in the culture solution being increased by increasing the amount of the culture solution stored therein. The culture container according to claim 4, wherein a plurality of regions that are formed lower by the steps are provided at intervals from each other. The culture container according to claim 4 or 5, wherein adjacent areas provided with a height difference by the step are connected by an inclined surface. The culture vessel according to claim 4, wherein the region formed lower by the step comprises a band-shaped groove. The culture container according to claim 1, further comprising at least two container pieces provided so as to be relatively movable, and a sealing means for maintaining a gap between the adjacent container pieces in a sealed state. The container piece comprises a cylinder and a piston fitted to the cylinder,
The culture vessel according to claim 5, wherein the sealing means is a seal member disposed between the cylinder and the piston. A culture container comprising: a container main body; and a cylindrical container wall member placed on the bottom surface of the container main body and defining the inside of the container main body.

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