WO2016047751A1 - Culture vessel - Google Patents

Culture vessel Download PDF

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Publication number
WO2016047751A1
WO2016047751A1 PCT/JP2015/077086 JP2015077086W WO2016047751A1 WO 2016047751 A1 WO2016047751 A1 WO 2016047751A1 JP 2015077086 W JP2015077086 W JP 2015077086W WO 2016047751 A1 WO2016047751 A1 WO 2016047751A1
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WO
WIPO (PCT)
Prior art keywords
culture
peripheral edge
barrier surface
inner peripheral
barrier
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PCT/JP2015/077086
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French (fr)
Japanese (ja)
Inventor
林　大輔
春男 大久保
魚住　孝之
宏昭 紀伊
Original Assignee
住友ベークライト株式会社
株式会社ニコン
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Application filed by 住友ベークライト株式会社, 株式会社ニコン filed Critical 住友ベークライト株式会社
Publication of WO2016047751A1 publication Critical patent/WO2016047751A1/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M1/00—Apparatus for enzymology or microbiology
    • C12M1/22—Petri dishes

Definitions

  • the present invention relates to a culture vessel. More specifically, the present invention relates to a culture vessel having a meniscus effect.
  • This application claims priority based on Japanese Patent Application No. 2014-195164 filed in Japan on September 25, 2014, the contents of which are incorporated herein by reference.
  • Patent Document 1 discloses that a solution that becomes a gel state by heating containing a biological substance on the bottom surface of a culture vessel is not yet available. Disclosed is a cell culture device having a layer A frozen in a gelled state, and a layer B frozen on an aqueous solution that does not become a gel state by heating. Has been.
  • the conventional technique aimed at solving the meniscus problem is performed by means for producing an action for reducing or preventing meniscus generation itself.
  • an object of the present invention is to provide a culture vessel that allows generation of meniscus and can solve the problem of meniscus.
  • the inventor has found that the object of the present invention can be achieved by providing the culture vessel with a structure that allows the meniscus to escape outside the range to be cultured and observed, and to complete the present invention. It came.
  • the culture container of the present invention includes a culture chamber having a bottom surface and a peripheral side wall surface provided on the periphery of the bottom surface.
  • a barrier surface is provided along the periphery of the bottom surface. The barrier surface rises with the inner peripheral edge inside the peripheral edge on the bottom surface as a base end, and is formed so as to go to the peripheral side wall surface and to form a boundary with the peripheral side wall surface.
  • the barrier surface is set to a height that allows a drop to be formed to a degree that suppresses cell survival on the barrier surface.
  • the shape of the barrier surface in a cross-sectional view from the lateral direction may be any of a straight line, a curved line, and a combination of a straight line and a curved line.
  • the barrier surface is composed of a lower barrier surface portion that rises from the inner peripheral edge, and an upper barrier surface portion that is connected to the lower barrier surface portion and forms a boundary with the peripheral side wall surface. It is preferable that the lower barrier surface portion rising from the angle forms an angle of 90 degrees or more and 150 degrees or less with the bottom surface.
  • the angle of the lower barrier surface portion is 90 degrees or more, the observation range of the collected cultured cells can be expanded without obstructing the field of view except for the peripheral portion in the culture chamber, which is 150 degrees or less. Thus, it is possible to further reduce the invasion of the cultured cells to the peripheral portion in the culture chamber that is difficult to observe.
  • the lower barrier surface portion and the upper barrier surface portion having the same inclination may be continuously provided without a boundary in a cross-sectional view from the lateral direction.
  • the structure of the barrier surface is simple, which is preferable from the viewpoint of manufacturing.
  • the curve has a shape in which an angle formed between the tangent and the bottom surface increases from the bottom surface side toward the peripheral side wall surface side. That is, the shape of the barrier surface in a cross-sectional view from the lateral direction (direction parallel to the bottom surface) is a curve, and the curve is an angle formed between the tangent line of the curve and the bottom surface from the bottom surface side toward the peripheral side wall surface side.
  • the shape may gradually increase.
  • a top view shape of the peripheral side wall surface and the inner peripheral edge may be a circle, and in this case, the diameter of the inner peripheral edge may be not less than 30 mm and not more than 170 mm.
  • the diameter of the inner peripheral edge is 30 mm or more, a space on the bottom surface for culturing and observation can be preferably obtained, and when it is 170 mm or less, an effect of collecting most of the cultured cells in an observable part. Can be enjoyed preferably.
  • a culture container in which a shadow formed by the meniscus does not hinder the observation of cultured cells can be provided.
  • FIG. 3 shows a partially enlarged view of the schematic cross-sectional view of FIG. 2.
  • the principal part enlarged view of the typical sectional drawing of FIG. 2 is shown.
  • the typical sectional view at the time of use of the culture container of the 1st modification of a 1st embodiment is shown.
  • the typical sectional view at the time of use of the culture container of the 2nd modification of a 1st embodiment is shown.
  • the typical sectional view at the time of use of the culture container of the 3rd modification of a 1st embodiment is shown.
  • the typical sectional view at the time of use of the culture container of the 4th modification of a 1st embodiment is shown.
  • the typical sectional view at the time of use of the culture container of the 5th modification of a 1st embodiment is shown.
  • the principal part enlarged view of the culture container of 2nd Embodiment is shown.
  • the principal part enlarged view of the culture container of 3rd Embodiment is shown.
  • the principal part enlarged view of the culture container of 4th Embodiment is shown.
  • the principal part enlarged view of the culture container of 5th Embodiment is shown.
  • the principal part enlarged view of the culture container of 6th Embodiment is shown.
  • the principal part enlarged view of the culture container of 7th Embodiment is shown.
  • FIG. 2 is a vertical view photograph after adding a culture medium to the culture container prepared in Example 1.
  • FIG. 2 is a vertically viewed photograph (partially enlarged) after cell culture in the culture vessel prepared in Example 1.
  • FIG. 2 is a vertically viewed photograph (partially enlarged) after cell culture in the culture vessel prepared in Example 1.
  • FIG. 4 is a vertical view photograph after adding a culture medium to the culture container produced in Example 2.
  • FIG. 4 is a vertically viewed photograph (partially enlarged) after cell culture in a culture vessel prepared in Example 2.
  • FIG. 4 is a vertically viewed photograph (partially enlarged) after cell culture in a culture vessel prepared in Example 2.
  • FIG. 4 is a vertically viewed photograph (partially enlarged) after cell culture in a culture vessel prepared in Example 2.
  • the typical sectional view of the culture container produced by comparative example 1 is shown. It is an up-down direction photograph after adding a culture medium to the culture container produced by the comparative example 1. It is an up-down direction view photograph (partially enlarged) after cell culture in the culture container produced in Comparative Example 1. It is an up-down direction view photograph (partially enlarged) after cell culture in the culture container produced in Comparative Example 1.
  • FIG. 1 the typical external appearance perspective view of the culture container of 1st Embodiment is shown.
  • FIG. 2 the typical sectional drawing at the time of use of the culture container of 1st Embodiment is shown.
  • the schematic cross-sectional view of FIG. 2 is a view of a plane cut along a vertical plane including the line AA of FIG.
  • FIG. 3 shows a partial enlargement of the schematic cross-sectional view of FIG.
  • FIG. 4 shows an enlarged view of a main part (circled portion) of the schematic cross-sectional view of FIG.
  • the upper side of each drawing is referred to as the upper side
  • the lower side is referred to as the lower side.
  • the culture vessel 100 includes a bottom surface 200, a peripheral wall surface 300, and a barrier surface 400 as surfaces constituting the inside thereof.
  • the culture chamber C is defined by the bottom surface 200, the peripheral side wall surface 300 and the barrier surface 400.
  • the culture container of the present embodiment includes a bottom surface 200, a barrier surface 400 provided outside along the peripheral edge B2 of the bottom surface 200, and a peripheral side wall surface provided further outward along the peripheral edge of the barrier surface 400. 300 culture chamber C.
  • the bottom surface 200 has a circular shape when viewed in the vertical direction, and is provided with a peripheral side wall surface 300 that rises from the entire periphery of the peripheral edge O and opens upward as shown in FIG.
  • the barrier surface 400 provided around the entire periphery of the peripheral edge O of the bottom surface 200 at a predetermined height h rises from the peripheral edge O of the bottom surface 200 to the inside with a predetermined distance w as a base end, While going to the peripheral side wall surface 300, it is formed so as to form a boundary B3 with the peripheral side wall surface 300 at a predetermined height h.
  • the predetermined distance w corresponds to the distance between the peripheral edge O and the internal peripheral edge B2.
  • the predetermined height h corresponds to the distance between the peripheral edge O and the boundary B3, and is set so as to be able to configure a shape that suppresses cell remaining on the barrier surface 400 in relation to the predetermined distance w.
  • the temperature is set to be equal to or lower than the lowest water level H of the culture solution to be stored in the culture chamber C.
  • the field of view from the vertical direction is difficult to observe because the culture liquid surface S in the peripheral part of the culture chamber C is curved due to the meniscus M phenomenon.
  • the culture vessel 100 by providing the barrier surface 400 having a predetermined height h rising from the inner peripheral edge B2, the culture cell is prevented from moving from the inner peripheral edge B2 to the barrier surface 400, and the barrier surface 400 is moved from the inner peripheral edge B2.
  • the barrier surface 400 By separating the inner peripheral edge B2 and the peripheral side wall surface 300 by a predetermined distance w in the outward direction, at least a part of the portion where the meniscus M is generated escapes from the inner peripheral edge B2. Therefore, while allowing the generation of the meniscus M, most of the cultured cells can be present inside the observable inner peripheral edge B2.
  • the predetermined distance w is not particularly limited.
  • the larger the predetermined distance w the more the influence of the meniscus M tends to be reduced.
  • the smaller the predetermined distance w the more the space on the bottom surface 200 for culture and observation tends to be sufficient.
  • the predetermined distance w may be 2 mm or more and 3 mm or less.
  • the predetermined distance w is set to be equal to or larger than the width W of the meniscus M.
  • the predetermined height h is not particularly limited, as long as it can be set so as to be able to configure a drop that suppresses cell remaining on the barrier surface 400 in relation to the predetermined distance w.
  • the predetermined height h is 1 mm or more and 21 mm or less, preferably 1 mm or more and 15 mm or less.
  • the predetermined height h is set to be equal to or lower than the water level H of the culture solution to be stored in the culture chamber C from the viewpoint of taking a sufficient volume to store the culture solution.
  • the boundary E between the culture liquid surface S and the inner wall surface of the culture vessel 100 is on the peripheral wall surface 300.
  • the barrier surface 400 includes a lower barrier surface portion 410 and an upper barrier surface portion 420.
  • the lower barrier surface portion 410 rises with the inner peripheral edge B2 as a base end so as to form an angle ⁇ 1 with the bottom surface 200, and is connected to the upper barrier surface portion 420 at a height h1.
  • the boundary between the lower barrier surface portion 410 and the upper barrier surface portion 420 forms a ridge line.
  • the upper barrier surface portion 420 is inclined toward the peripheral side wall surface 300 at an angle ⁇ 2 with respect to the bottom surface 200, and forms a boundary B3 with the peripheral side wall surface 300.
  • the angle ⁇ 1 between the lower barrier surface portion 410 and the bottom surface 200 is 90 degrees. Thereby, it is possible to effectively prevent the cultured cells on the bottom surface 200 from moving beyond the inner peripheral edge B2.
  • the angle ⁇ 2 that is the inclination angle of the upper barrier surface portion 420 is not particularly limited. For example, as the angle ⁇ 2 is larger, the height h1 of the lower barrier surface portion 410 is sufficiently secured, and the invasion of the cultured cells into the outer portion of the inner peripheral edge B2 that is difficult to observe in the vertical direction can be reduced. It can be determined appropriately by those skilled in the art based on the tendency that the smaller the angle ⁇ 2 is, the more the volume for storing the culture solution can be taken.
  • the height h1 of the lower barrier surface portion 410 is not particularly limited.
  • the height h1 may be set to, for example, 0.5 mm or 1 mm.
  • the diameter ⁇ of the inner peripheral edge B2 may be equal to the diameter of the inner bottom surface of the conventional culture vessel. For example, it is 30 mm or more and 170 mm or less, preferably 32 mm or more and 90 mm or less. By being above the lower limit value, it is possible to preferably take a space on the bottom surface 200 for culture and observation, and by being below the upper limit value, an effect of collecting most of the cultured cells in an observable portion. Can be enjoyed preferably.
  • the culture vessel 100 can be made of a transparent material.
  • the material having transparency include inorganic substances typified by glass and quartz, and organic substances typified by synthetic resins.
  • Synthetic resins include polystyrene (PS), polypropylene (PP), polymethylpentene (PMP), polycarbonate (PC), polymethyl methacrylate (PMMA), polymethylacrylmethylimide (PMMI), and cycloolefin copolymer (COC). And the like.
  • combined from two or more of the monomer units of these polymers is also mentioned.
  • the culture vessel 100 may be a structure formed by integral molding, or may be a structure formed by a combination of a member forming the bottom surface 200 and members forming the peripheral wall surface 300 and the barrier surface 400, for example.
  • each member may be made of different materials.
  • the bottom surface 200, the peripheral wall surface 300, and the barrier surface 400 of the culture vessel 100 may be subjected to a surface treatment from the viewpoint of physical, chemical, and / or biochemistry.
  • a surface treatment can be appropriately selected by those skilled in the art.
  • the barrier surface 400 or the barrier surface 400 and the peripheral wall surface 300 may be subjected to a surface treatment for inhibiting cell culture.
  • an additional structure may be provided depending on the purpose of use of the culture vessel 100 or the like.
  • Such an additional structure can be appropriately selected by those skilled in the art.
  • the bottom surface 200 such as a lid for placing the culture chamber C in a covered state, a partition for partitioning the contents, and a grid pattern for specifying the amount and position of the contents (particularly cell groups).
  • the guide structure provided in the above.
  • the culture vessel 100 can be variously modified in addition to the illustrated one.
  • at least a part of the meniscus M only needs to escape to the outside of the inner peripheral edge B2, so that the boundary E is 90 degrees with respect to the bottom surface 200 with the inner peripheral edge B2 being the base end of the barrier surface 400.
  • Any structure is acceptable as long as it is not formed on the rising lower barrier surface 410. That is, in the modification, when the culture solution is accommodated in the culture chamber C, the boundary E between the liquid surface S of the culture solution and the inner wall of the culture vessel 100 is outside the inner peripheral edge B2. Good.
  • FIG. 5 shows a first modification of the first embodiment.
  • the barrier surface 400a of the culture vessel 100a shown in FIG. 5 is set such that the predetermined distance wa is shorter than the predetermined distance w in the first embodiment, so that the inner peripheral edge B2a serving as the base end is the inner space in the first embodiment. It is outside the peripheral edge B2.
  • the meniscus M that escapes to the outside from the inner peripheral edge B2a becomes a part thereof, but the influence of the meniscus M can be reduced. That is, also in this modified example, the meniscus M is escaped to the outside from the inner peripheral edge B2a, so that the influence of the meniscus M can be reduced.
  • FIG. 6 shows a second modification of the first embodiment.
  • the barrier surface 400b of the culture vessel 100b shown in FIG. 6 has a predetermined height hb set to be larger than the predetermined height h in the first embodiment, so that the boundary B3b with the peripheral wall surface 300b is the first implementation. It is above the boundary B3 in the form. As a result, it is possible to further reduce the invasion of the cultured cells into the outer portion of the inner peripheral edge B2, which is difficult to observe when viewed in the vertical direction.
  • the predetermined height hb is equal to or higher than the water level H of the culture solution to be accommodated in the culture chamber C, but less than the height of the boundary E between the culture solution surface S and the peripheral side wall surface 300b.
  • FIG. 7 shows a third modification of the first embodiment.
  • the barrier surface 400c of the culture vessel 100c shown in FIG. 7 has an inner peripheral edge serving as a base end when the predetermined distance wa is set shorter than the predetermined distance w in the first embodiment.
  • B2a is outside the inner peripheral edge B2 in the first embodiment.
  • the barrier surface 400c has a boundary B3b with the peripheral side wall surface 300b above the boundary B3 in the first embodiment, as in the second modification.
  • the predetermined height hb is equal to or higher than the water level H of the culture solution to be accommodated in the culture chamber C, but less than the height of the boundary E between the culture solution surface S and the peripheral side wall surface 300b.
  • FIG. 8 shows a fourth modification of the first embodiment.
  • the barrier surface 400d of the culture vessel 100d shown in FIG. 8 is set such that the predetermined height hd is larger than the predetermined height hb in the second modified example, so that the boundary B3d with the peripheral wall surface 300d is the second. It is further above the boundary B3b in the modification. As a result, it is possible to further reduce the invasion of the cultured cells into the outer portion of the inner peripheral edge B2, which is difficult to observe when viewed in the vertical direction.
  • the predetermined height hd of the barrier surface 400d is to be accommodated in the culture chamber C.
  • the liquid level exceeds H and exceeds the height of the boundary Ed.
  • the meniscus M can escape to the outside of the inner peripheral edge B2, and the influence of the meniscus M can be avoided.
  • FIG. 9 shows a fifth modification of the first embodiment.
  • the barrier surface 400e of the culture vessel 100e shown in FIG. 9 is set such that the predetermined height hd is larger than the predetermined height hb in the second and third modifications.
  • the boundary B3d with the peripheral side wall surface 300d is further above the boundary B3b in the second and third modified examples.
  • the predetermined height hd of the barrier surface 400e is to be accommodated in the culture chamber C.
  • the liquid level exceeds H and exceeds the boundary Ee.
  • the predetermined distance wa is set shorter than the predetermined distance w in the first embodiment, so that the inner peripheral edge B2a serving as the base end is more than the inner peripheral edge B2 in the first embodiment.
  • the meniscus M that escapes to the outside from the inner peripheral edge B2a becomes a part thereof, but the influence of the meniscus M can be reduced. That is, also in this modified example, the meniscus M is escaped to the outside from the inner peripheral edge B2a, so that the influence of the meniscus M can be reduced.
  • the barrier surface 400 only needs to have a shape that passes between the inner peripheral edge B2 and the boundary B3 (a shape that connects the inner peripheral edge B2 and the boundary B3) in a cross-sectional view from the lateral direction. It is not limited to the shape shown in one embodiment.
  • the boundary between the lower barrier surface portion 410 and the upper barrier surface portion 420 may form a ridge line as shown in FIG. 4, or the boundary may be C chamfered or R chamfered.
  • the cross-sectional shape viewed from the lateral direction of the barrier surface 400 may be a straight line, a curved line, or a combination of a straight line and a curved line, and various shapes are widely acceptable. Is done.
  • the barrier surface 400 has the inner peripheral edge B2 and the boundary B3 in the cross-sectional view from the lateral direction, from the viewpoint of more surely preventing the cultured cells from being present on the outer portion of the inner peripheral edge B2, which is difficult to observe in the vertical direction.
  • FIGS. 10 to 15 correspond to FIG. 4 of the first embodiment.
  • differences from the first embodiment will be mainly described, and description of the same points will be omitted.
  • FIG. 10 shows an enlarged view of the main part of the culture vessel of the second embodiment.
  • the barrier surface 400f shown in FIG. 10 includes a lower barrier surface portion 410f and an upper barrier surface portion 420f.
  • the lower barrier surface portion 410f rises with the inner peripheral edge B2 as a base end so as to form an angle ⁇ 1 (90 degrees) with the bottom surface 200, and is connected to the upper barrier surface portion 420f parallel to the bottom surface 200 (angle ⁇ 2 is 180 degrees).
  • angle ⁇ 1 90 degrees
  • the boundary between the lower barrier surface portion 410f and the upper barrier surface portion 420f forms a ridge line, but the boundary may be C-chamfered or R-chamfered.
  • FIG. 11 shows the principal part enlarged view of the culture container of 3rd Embodiment.
  • the barrier surface 400g shown in FIG. 11 includes a lower barrier surface portion 410g and an upper barrier surface portion 420g.
  • the lower barrier surface portion 410g rises with the inner peripheral edge B2 as a base end so as to form an angle ⁇ 1 with the bottom surface 200, and is connected to the upper barrier surface portion 420g parallel to the bottom surface 200 (angle ⁇ 2 is 180 degrees).
  • the angle ⁇ 1 may be greater than 90 degrees and less than or equal to 150 degrees.
  • the boundary between the lower barrier surface portion 410g and the upper barrier surface portion 420g forms a ridge line, but the boundary may be C-chamfered or R-chamfered.
  • FIG. 12 shows an enlarged view of the main part of the culture vessel of the fourth embodiment.
  • the barrier surface 400h shown in FIG. 12 includes a lower barrier surface portion 410h and an upper barrier surface portion 420h. Since the angle ⁇ 1 formed by the lower barrier surface portion 410h and the bottom surface 200 is the same as the angle ⁇ 2 formed by the upper barrier surface portion 420h and the bottom surface 200, both surfaces are connected without boundaries. For this reason, the structure of the barrier surface 400h is simple, which is preferable from the viewpoint of manufacturing.
  • FIG. 13 shows the principal part enlarged view of the culture container of 5th Embodiment.
  • the barrier surface 400i shown in FIG. 13 has a curved surface when viewed from the lateral direction.
  • the angles ⁇ , ⁇ ′, ⁇ ′′ formed by the tangent to the curved surface and the bottom surface 200 gradually increase from the bottom surface 200 side toward the peripheral side wall surface 300 ( ⁇ ⁇ ′ ⁇ ′′).
  • the angles ⁇ , ⁇ ′, and ⁇ ′′ can be changed in the range of 90 degrees to 180 degrees.
  • FIG. 14 shows the principal part enlarged view of the culture container of 6th Embodiment.
  • the barrier surface 400j shown in FIG. 14 includes a lower barrier surface portion 410j and an upper barrier surface portion 420j.
  • the lower barrier surface portion 410j has a straight cross-sectional shape viewed from the lateral direction, rises with the inner peripheral edge B2 as the base end so as to form an angle ⁇ 1 with the bottom surface 200, and the cross-sectional shape is a curve at a position of height h1. It is connected to a certain upper barrier surface portion 420j. At the position of the height h1, the boundary between the lower barrier surface portion 410j and the upper barrier surface portion 420j forms a ridge line.
  • the angle ⁇ 1 may be not less than 90 degrees and not more than 150 degrees.
  • the cross-sectional shape of the upper barrier surface portion 420j may be a curved surface similar to the barrier surface 400i in the fifth embodiment.
  • FIG. 15 shows the principal part enlarged view of the culture container of 7th Embodiment.
  • the barrier surface 400k shown in FIG. 15 includes a lower barrier surface portion 410k and an upper barrier surface portion 420k.
  • the lower barrier surface portion 410k rises with the inner peripheral edge B2 as a base end so as to form an angle ⁇ 1 with the bottom surface 200, and is continuous with the upper barrier surface portion 420k at a height h1.
  • the boundary between the lower barrier surface portion 410k and the upper barrier surface portion 420k forms a ridge line.
  • the upper barrier surface portion 420k is inclined toward the peripheral side wall surface 300 at an angle ⁇ 2 with respect to the bottom surface 200, and forms a boundary B3 with the peripheral side wall surface 300.
  • the angle ⁇ 1 may be not less than 90 degrees and not more than 150 degrees.
  • the entire region within the inner peripheral edge B2 to be cultured and observed can be observed with good visibility from the vertical direction view. It is possible to reduce the entry of cultured cells into the outer portion of the inner peripheral edge B2, which is difficult.
  • the angle ⁇ 2 is not particularly limited.
  • the angle ⁇ 2 is larger, the height h1 of the lower barrier surface portion 410k is sufficiently secured, and the tendency of the cultured cells to enter the outer portion of the inner peripheral edge B2, which is difficult to observe in the vertical direction, can be reduced. Therefore, the smaller the angle ⁇ 2, the more appropriate it can be determined by those skilled in the art based on the tendency that a sufficient volume for accommodating the culture solution tends to be taken.
  • the boundary between the lower barrier surface portion 410k and the upper barrier surface portion 420k forms a ridge line, but the boundary may be C-chamfered or R-chamfered.
  • FIG. 16 is a schematic external perspective view of the culture vessel of the eighth embodiment.
  • the culture vessel 100l has a plurality of culture chambers C.
  • one culture vessel 100l has six culture chambers C, but the number of culture chambers C is not limited to this.
  • the number of the culture chambers C is not limited to this.
  • Those skilled in the art can appropriately determine the space on the bottom surface 200 for observation based on the appropriate space.
  • the structure of the culture chamber C of the culture vessel 100l the structure in any of the first to seventh embodiments and their modifications may be applied.
  • Example 1 A prototype of a culture vessel having the structure shown in FIG. 9 was prepared by laminating a polystyrene disc to a cut product using polycarbonate (in which the inner peripheral surface side of an open and bottomless cylinder was cut). (Note that the actual product of the culture vessel may be produced by integral molding. Hereinafter, the prototype produced by bonding is also simply referred to as “culture vessel”). ⁇ 1 was 90 degrees and ⁇ 2 was 120 degrees.
  • FIG. 17 shows a vertical view photograph.
  • the culture container has a barrier surface 400e, so that most of the meniscus M is generated outside the inner peripheral edge B2a, so that the culture container is well visible inside the inner peripheral edge B2a for culture and observation. It was confirmed that the range which can be expanded from the comparative example 1 mentioned later.
  • FIG. 18 shows a microscopic observation photograph of the center of the culture chamber
  • FIG. 19 shows the periphery of the culture chamber.
  • the barrier surface 400e causes most of the meniscus M to be generated outside the inner peripheral edge B2a, and the range that can be observed inside the inner peripheral edge B2a for culture and observation is Comparative Example 1 described later. It was confirmed that it was expanding.
  • Example 2 A prototype of a culture vessel having the structure shown in FIG. 9 was prepared by laminating a polystyrene disc to a cut product using polycarbonate (in which the inner peripheral surface of an open-bottomed bottomless cylinder was cut). .
  • the culture container of this example is the same as the culture container of Example 1 except that ⁇ 2 is 150 degrees.
  • FIG. 20 shows a vertical view photograph.
  • the culture vessel has a barrier surface 400e, so that a shadow caused by the meniscus M is generated outside the inner peripheral edge B2a, and there is a range that can be favorably seen inside the inner peripheral edge B2a for culture and observation. It was confirmed that it was larger than Comparative Example 1 described later.
  • FIG. 21 shows a microscope observation photograph of the center of the culture chamber and FIG. 22 shows the periphery of the culture chamber. As shown in FIG. 21, it was confirmed that the cells normally adhered to the bottom surface 200. Further, as shown in FIG. 22, the barrier surface 400e causes most of the meniscus M to be generated outside the inner peripheral edge B2a, and the range that can be observed inside the inner peripheral edge B2a for culture and observation is Comparative Example 1 described later. It was confirmed that it was expanding.
  • Example 2 Comparison of the microscopic observation photographs (FIGS. 19 and 22) in the periphery of the culture chamber of Example 1 and Example 2 confirms that Example 2 (FIG. 22) has a wider range in which the cells can be seen well. It was done.
  • the boundary Ee between the culture liquid surface S and the inner wall surface of the culture vessel 100e is on the upper peripheral wall surface portion 420e, and the predetermined height hd of the barrier surface 400e is accommodated in the culture chamber C.
  • the level of the culture broth was below the level H and the height of the boundary E was also below.
  • Example 2 in which ⁇ 2 is 150 degrees, the range of meniscus M generated outside the inner peripheral edge B2a becomes larger, and this Example 2 ( In FIG. 22), the range in which the cells can be visually recognized is widened.
  • a culture vessel having no barrier surface was prepared by attaching a polystyrene disk to a polycarbonate open-bottomed bottomless cylinder. A schematic cross-sectional view of this culture vessel is shown in FIG.
  • FIG. 24 shows a vertical view photograph. As shown in FIG. 24, all of the meniscus M occurred inside the peripheral edge B, and a shadow due to the meniscus M was confirmed inside the peripheral edge B for culture and observation.
  • FIG. 25 shows a microscope observation photograph of the center of the culture chamber
  • FIG. 26 shows the periphery of the culture chamber. As shown in FIG. 25, it was confirmed that the cells normally adhered to the bottom surface 200.
  • FIG. 25 shows a microscope observation photograph of the center of the culture chamber
  • FIG. 26 shows the periphery of the culture chamber. As shown in FIG. 25, it was confirmed that the cells normally adhered to the bottom surface 200.
  • the culture vessels 100 and 100l correspond to “culture vessels”
  • the bottom surface 200 corresponds to “bottom surface”
  • the peripheral side wall surfaces 300, 300b, and 300d correspond to “peripheral side wall surfaces”
  • the barrier surface 400 400a, 400b, 400c, 400d, 400e, 400f, 400g, 400h, 400i, 400j, 400k correspond to the “barrier surface”
  • the lower barrier surface portions 410, 410f, 410g, 410h, 410j, 410k are “lower side”.
  • the upper barrier surface portions 420, 420d, 420e, 420f, 420g, 420h, 420j, 420k correspond to the “upper barrier surface portion”
  • the peripheral edge O corresponds to the “periphery”
  • the inner peripheral edges B2, B2a corresponds to the “inner periphery”
  • the boundaries B3, B3b, B3d correspond to “boundaries” (between the barrier surface and the peripheral side wall surface)
  • the culture chamber C is the “culture chamber”.
  • the angle ⁇ 1 corresponds to the “angle” (the lower barrier surface portion and the bottom surface form), and the angles ⁇ , ⁇ ′, and ⁇ ′′ (the tangent of the cross-sectional shape of the barrier surface and the bottom surface form) “ Corresponds to “Angle”.
  • a culture container in which a shadow formed by the meniscus does not hinder the observation of cultured cells can be provided.

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Abstract

This culture vessel includes a culture chamber having a bottom surface and a peripheral side wall surface disposed at the periphery of the bottom surface, wherein a barrier surface of a predetermined height is disposed along the periphery inside the culture chamber, and the barrier surface is formed so as to rise from an inner periphery, which serves as the base end and is inward from the periphery of the bottom surface, to the peripheral side wall surface, and so as to form a boundary with the peripheral side wall surface.

Description

培養容器Culture vessel
　本発明は、培養容器に関する。より具体的には、本発明は、対メニスカス効果を有する培養容器に関する。本願は、２０１４年９月２５日に、日本に出願された特願２０１４－１９５１６４号に基づき優先権を主張し、その内容をここに援用する。 The present invention relates to a culture vessel. More specifically, the present invention relates to a culture vessel having a meniscus effect. This application claims priority based on Japanese Patent Application No. 2014-195164 filed in Japan on September 25, 2014, the contents of which are incorporated herein by reference.
　培養容器内に培養溶液を入れた場合、溶液の界面張力の働きにより、界面には凹型のメニスカスが生じる。特に、培養にゲルを用いる場合、ゲル化前の溶液の粘性が高く培養容器の内壁面に溶液が付着しやすいことから、この傾向はより顕著となる。培養面の観察において、メニスカスは、それが発生する部分に影を形成するため、当該部分に存在する培養細胞の観察の支障になる。 When a culture solution is placed in a culture vessel, a concave meniscus is generated at the interface due to the interfacial tension of the solution. In particular, when a gel is used for culturing, this tendency becomes more prominent because the viscosity of the solution before gelation is high and the solution tends to adhere to the inner wall surface of the culture vessel. In the observation of the culture surface, the meniscus forms a shadow in the portion where it occurs, which hinders observation of the cultured cells present in the portion.
　メニスカスの問題を解決することを目的とした技術として、特開２００１－３４００７０号公報（特許文献１）に、培養容器底面に、生体由来物質を含み加温することによりゲル状態となる溶液を未ゲル化状態で凍結した層Ａを有し、かつ加温することによりゲル状態とならない水溶液で凍結した層Ｂを層Ａの上に重ねて有していることを特徴とする細胞培養器が開示されている。 As a technique aimed at solving the meniscus problem, Japanese Patent Application Laid-Open No. 2001-340070 (Patent Document 1) discloses that a solution that becomes a gel state by heating containing a biological substance on the bottom surface of a culture vessel is not yet available. Disclosed is a cell culture device having a layer A frozen in a gelled state, and a layer B frozen on an aqueous solution that does not become a gel state by heating. Has been.
特開２００１－３４００７０号公報JP 2001-340070 A
　メニスカスの問題を解決することを目的とした従来の技術は、メニスカスの発生自体を低減または防止する作用を生じさせる手段によってなされる。 The conventional technique aimed at solving the meniscus problem is performed by means for producing an action for reducing or preventing meniscus generation itself.
　一方、本発明の目的は、メニスカスの発生を許容するとともに、メニスカスの問題を解決することができる培養容器を提供することにある。 On the other hand, an object of the present invention is to provide a culture vessel that allows generation of meniscus and can solve the problem of meniscus.
　本発明者は、鋭意検討の結果、培養容器に、培養および観察すべき範囲の外側にメニスカスを逃がす構造を持たせることにより、上記本発明の目的が達成できることを見出し、本発明を完成するに至った。 As a result of intensive studies, the inventor has found that the object of the present invention can be achieved by providing the culture vessel with a structure that allows the meniscus to escape outside the range to be cultured and observed, and to complete the present invention. It came.
（１）
　本発明の培養容器は、底面と、当該底面の周縁に設けられた周側壁面とを有する培養室を含む。培養室内には、底面の周縁に沿って障壁面が周設されている。障壁面は、底面上の、周縁より内側の内部周縁を基端として立ち上がり、周側壁面に向かうとともに、周側壁面との境界をなすように形成されている。 (1)
The culture container of the present invention includes a culture chamber having a bottom surface and a peripheral side wall surface provided on the periphery of the bottom surface. In the culture chamber, a barrier surface is provided along the periphery of the bottom surface. The barrier surface rises with the inner peripheral edge inside the peripheral edge on the bottom surface as a base end, and is formed so as to go to the peripheral side wall surface and to form a boundary with the peripheral side wall surface.
　周縁より内側に隔てた位置に内部周縁が存在するように構成することにより、培養室内に培養液を収容した場合に発生するメニスカスの影響を少なくとも低減させる。障壁面は、障壁面上における細胞残存を抑制する程度の落差を形成可能とする高さに設定される。
　上記の構成によって、培養液面におけるメニスカスの発生により観察が困難となる培養室内の周縁部分における培養細胞の存在を低減させることができる。したがって、メニスカスの発生を許容しながらも、培養した細胞の観察範囲を広げることが可能となる。 By configuring the inner periphery to be present at a position separated from the periphery, the influence of the meniscus generated when the culture solution is stored in the culture chamber is at least reduced. The barrier surface is set to a height that allows a drop to be formed to a degree that suppresses cell survival on the barrier surface.
With the above configuration, it is possible to reduce the presence of cultured cells in the peripheral portion of the culture chamber that is difficult to observe due to generation of a meniscus on the culture surface. Therefore, it is possible to expand the observation range of the cultured cells while allowing the generation of meniscus.
（２）
　横方向からの断面視における障壁面の形状が、直線、曲線、および直線と曲線との組み合わせのいずれかであってよい。 (2)
The shape of the barrier surface in a cross-sectional view from the lateral direction may be any of a straight line, a curved line, and a combination of a straight line and a curved line.
　このように、様々な形状の障壁面が広く許容される。 Thus, variously shaped barrier surfaces are widely accepted.
（３）
　横方向からの断面視において、障壁面が、内部周縁から立ち上がる下側障壁面部と、当該下部障壁面部に連設されかつ周側壁面との境界をなす上側障壁面部と、から構成され、内部周縁から立ち上がる下側障壁面部が、底面と９０度以上１５０度以下の角度をなすことが好ましい。 (3)
In a cross-sectional view from the lateral direction, the barrier surface is composed of a lower barrier surface portion that rises from the inner peripheral edge, and an upper barrier surface portion that is connected to the lower barrier surface portion and forms a boundary with the peripheral side wall surface. It is preferable that the lower barrier surface portion rising from the angle forms an angle of 90 degrees or more and 150 degrees or less with the bottom surface.
　下側障壁面部の当該角度が９０度以上であることにより、培養室内の周縁部分を除いた部分の視界が遮られることなく集めた培養細胞の観察範囲を広げることができ、１５０度以下であることにより、観察困難な培養室内の周縁部分への培養細胞の進入をより少なくすることができる。 When the angle of the lower barrier surface portion is 90 degrees or more, the observation range of the collected cultured cells can be expanded without obstructing the field of view except for the peripheral portion in the culture chamber, which is 150 degrees or less. Thus, it is possible to further reduce the invasion of the cultured cells to the peripheral portion in the culture chamber that is difficult to observe.
（４）
　上記（３）の培養容器では、横方向からの断面視において、互いに同じ傾斜の下側障壁面部と上側障壁面部とが境界なく連設していてよい。 (4)
In the culture vessel of (3) above, the lower barrier surface portion and the upper barrier surface portion having the same inclination may be continuously provided without a boundary in a cross-sectional view from the lateral direction.
　この場合、障壁面の構造が簡素であり、製造上の観点からも好ましい。 In this case, the structure of the barrier surface is simple, which is preferable from the viewpoint of manufacturing.
（５）
　横方向からの断面視における障壁面の形状が曲線である場合、当該曲線は、底面側から周側壁面側に向かって接線と前記底面とのなす角度が大きくなる形状であることが好ましい。すなわち、横方向（底面に対して平行な方向）からの断面視における障壁面の形状が曲線であり、当該曲線は、底面側から周側壁面側に向かって曲線の接線と底面とのなす角度が漸次大きくなる形状であってもよい。 (5)
When the shape of the barrier surface in a cross-sectional view from the lateral direction is a curve, it is preferable that the curve has a shape in which an angle formed between the tangent and the bottom surface increases from the bottom surface side toward the peripheral side wall surface side. That is, the shape of the barrier surface in a cross-sectional view from the lateral direction (direction parallel to the bottom surface) is a curve, and the curve is an angle formed between the tangent line of the curve and the bottom surface from the bottom surface side toward the peripheral side wall surface side. The shape may gradually increase.
　この場合、観察困難な培養室内の周縁部分への培養細胞の不在を好ましく担保するとともに、製造上の観点からも好ましい。 In this case, it is preferable to ensure the absence of cultured cells in the peripheral portion of the culture chamber which is difficult to observe, and from the viewpoint of manufacturing.
（６）
　周側壁面および内部周縁の上面視形状は円であってよく、この場合、内部周縁の直径は、３０ｍｍ以上１７０ｍｍ以下であってよい。 (6)
A top view shape of the peripheral side wall surface and the inner peripheral edge may be a circle, and in this case, the diameter of the inner peripheral edge may be not less than 30 mm and not more than 170 mm.
　内部周縁の直径が３０ｍｍ以上であることにより、培養および観察のための底面上のスペースを好ましくとることができ、１７０ｍｍ以下であることにより、培養した細胞の大部分を観察可能な部分に集める効果を好ましく享受することができる。 When the diameter of the inner peripheral edge is 30 mm or more, a space on the bottom surface for culturing and observation can be preferably obtained, and when it is 170 mm or less, an effect of collecting most of the cultured cells in an observable part. Can be enjoyed preferably.
　本発明によれば、メニスカスが発生しても、当該メニスカスにより形成される影が、培養細胞の観察の支障にならない培養容器を提供することができる。 According to the present invention, even when a meniscus is generated, a culture container in which a shadow formed by the meniscus does not hinder the observation of cultured cells can be provided.
第１実施形態の培養容器の模式的外観斜視図を示す。The typical external appearance perspective view of the culture container of a 1st embodiment is shown. 第１実施形態の培養容器の使用時における模式的断面図を示す。The typical sectional view at the time of use of the culture vessel of a 1st embodiment is shown. 図２の模式的断面図の一部拡大図を示す。FIG. 3 shows a partially enlarged view of the schematic cross-sectional view of FIG. 2. 図２の模式的断面図の要部拡大図を示す。The principal part enlarged view of the typical sectional drawing of FIG. 2 is shown. 第１実施形態の第１変形例の培養容器使用時における模式的断面図を示す。The typical sectional view at the time of use of the culture container of the 1st modification of a 1st embodiment is shown. 第１実施形態の第２変形例の培養容器使用時における模式的断面図を示す。The typical sectional view at the time of use of the culture container of the 2nd modification of a 1st embodiment is shown. 第１実施形態の第３変形例の培養容器使用時における模式的断面図を示す。The typical sectional view at the time of use of the culture container of the 3rd modification of a 1st embodiment is shown. 第１実施形態の第４変形例の培養容器使用時における模式的断面図を示す。The typical sectional view at the time of use of the culture container of the 4th modification of a 1st embodiment is shown. 第１実施形態の第５変形例の培養容器使用時における模式的断面図を示す。The typical sectional view at the time of use of the culture container of the 5th modification of a 1st embodiment is shown. 第２実施形態の培養容器の要部拡大図を示す。The principal part enlarged view of the culture container of 2nd Embodiment is shown. 第３実施形態の培養容器の要部拡大図を示す。The principal part enlarged view of the culture container of 3rd Embodiment is shown. 第４実施形態の培養容器の要部拡大図を示す。The principal part enlarged view of the culture container of 4th Embodiment is shown. 第５実施形態の培養容器の要部拡大図を示す。The principal part enlarged view of the culture container of 5th Embodiment is shown. 第６実施形態の培養容器の要部拡大図を示す。The principal part enlarged view of the culture container of 6th Embodiment is shown. 第７実施形態の培養容器の要部拡大図を示す。The principal part enlarged view of the culture container of 7th Embodiment is shown. 第８実施形態の培養容器の模式的外観斜視図を示す。The typical external appearance perspective view of the culture container of 8th Embodiment is shown. 実施例１で作製された培養容器に培地を加えた後の上下方向視写真である。2 is a vertical view photograph after adding a culture medium to the culture container prepared in Example 1. FIG. 実施例１で作製された培養容器内で細胞培養した後の上下方向視写真（一部拡大）である。FIG. 2 is a vertically viewed photograph (partially enlarged) after cell culture in the culture vessel prepared in Example 1. FIG. 実施例１で作製された培養容器内で細胞培養した後の上下方向視写真（一部拡大）である。FIG. 2 is a vertically viewed photograph (partially enlarged) after cell culture in the culture vessel prepared in Example 1. FIG. 実施例２で作製された培養容器に培地を加えた後の上下方向視写真である。4 is a vertical view photograph after adding a culture medium to the culture container produced in Example 2. FIG. 実施例２で作製された培養容器内で細胞培養した後の上下方向視写真（一部拡大）である。4 is a vertically viewed photograph (partially enlarged) after cell culture in a culture vessel prepared in Example 2. FIG. 実施例２で作製された培養容器内で細胞培養した後の上下方向視写真（一部拡大）である。4 is a vertically viewed photograph (partially enlarged) after cell culture in a culture vessel prepared in Example 2. FIG. 比較例１で作製された培養容器の模式的断面図を示す。The typical sectional view of the culture container produced by comparative example 1 is shown. 比較例１で作製された培養容器に培地を加えた後の上下方向視写真である。It is an up-down direction photograph after adding a culture medium to the culture container produced by the comparative example 1. 比較例１で作製された培養容器内で細胞培養した後の上下方向視写真（一部拡大）である。It is an up-down direction view photograph (partially enlarged) after cell culture in the culture container produced in Comparative Example 1. 比較例１で作製された培養容器内で細胞培養した後の上下方向視写真（一部拡大）である。It is an up-down direction view photograph (partially enlarged) after cell culture in the culture container produced in Comparative Example 1.
　以下、図面を参照しつつ、本発明の実施の形態について説明する。以下の説明では、同一の要素には同一の符号を付しており、特に断りのない限り、それらの名称および機能も同じである。したがって、基本的にそれらについての詳細な説明は繰り返さない。 Hereinafter, embodiments of the present invention will be described with reference to the drawings. In the following description, the same reference numerals are given to the same elements, and their names and functions are the same unless otherwise specified. Therefore, basically, detailed description thereof will not be repeated.
［第１実施形態］
　図１に、第１実施形態の培養容器の模式的外観斜視図を示す。図２に、第１実施形態の培養容器の使用時における模式的断面図を示す。図２の模式的断面図は、図１のＡ－Ａ線を含む鉛直面で切断した面を横方向から見た図である。図３に、図２の模式的断面図の一部拡大を示す。図４に、図２の模式的断面図の要部（円囲み部分）の拡大図を示す。なお、説明の便宜上、各図面の上側を上、下側を下と呼称する。 [First Embodiment]
In FIG. 1, the typical external appearance perspective view of the culture container of 1st Embodiment is shown. In FIG. 2, the typical sectional drawing at the time of use of the culture container of 1st Embodiment is shown. The schematic cross-sectional view of FIG. 2 is a view of a plane cut along a vertical plane including the line AA of FIG. FIG. 3 shows a partial enlargement of the schematic cross-sectional view of FIG. FIG. 4 shows an enlarged view of a main part (circled portion) of the schematic cross-sectional view of FIG. For convenience of explanation, the upper side of each drawing is referred to as the upper side, and the lower side is referred to as the lower side.
　図１及び図２に示すように、培養容器１００は、その内側を構成する面として、底面２００、周側壁面３００および障壁面４００を含む。これら底面２００、周側壁面３００および障壁面４００によって、培養室Ｃが規定される。いいかえると、本実施形態の培養容器は、底面２００と、底面２００の周縁Ｂ２に沿って外側に設けられた障壁面４００と、障壁面４００の周縁に沿って更に外側に設けられた周側壁面３００とを備える培養室Ｃを含む。 As shown in FIGS. 1 and 2, the culture vessel 100 includes a bottom surface 200, a peripheral wall surface 300, and a barrier surface 400 as surfaces constituting the inside thereof. The culture chamber C is defined by the bottom surface 200, the peripheral side wall surface 300 and the barrier surface 400. In other words, the culture container of the present embodiment includes a bottom surface 200, a barrier surface 400 provided outside along the peripheral edge B2 of the bottom surface 200, and a peripheral side wall surface provided further outward along the peripheral edge of the barrier surface 400. 300 culture chamber C.
　底面２００は、上下方向視円形であり、図３に示すように、その周縁Ｏの全周から立ち上がり上方開口した周側壁面３００が設けられている。底面２００の周縁Ｏの全周に沿って所定の高さｈで周設された障壁面４００は、底面２００の周縁Ｏから内側へ所定の距離ｗを隔てた内部周縁Ｂ２を基端として立ち上がり、周側壁面３００へ向かうとともに、所定の高さｈの位置で周側壁面３００との境界Ｂ３をなすように形成されている。 The bottom surface 200 has a circular shape when viewed in the vertical direction, and is provided with a peripheral side wall surface 300 that rises from the entire periphery of the peripheral edge O and opens upward as shown in FIG. The barrier surface 400 provided around the entire periphery of the peripheral edge O of the bottom surface 200 at a predetermined height h rises from the peripheral edge O of the bottom surface 200 to the inside with a predetermined distance w as a base end, While going to the peripheral side wall surface 300, it is formed so as to form a boundary B3 with the peripheral side wall surface 300 at a predetermined height h.
　図３に示すように、所定の距離ｗは、周縁Ｏと内部周縁Ｂ２との距離に相当する。所定の高さｈは、周縁Ｏと境界Ｂ３との距離に相当し、所定の距離ｗとの関係で障壁面４００への細胞残存を抑制する形状を構成可能となるように設定される。本実施形態では、培養室Ｃ内に収容すべき培養液の最低水位Ｈ以下となるように設定される。 As shown in FIG. 3, the predetermined distance w corresponds to the distance between the peripheral edge O and the internal peripheral edge B2. The predetermined height h corresponds to the distance between the peripheral edge O and the boundary B3, and is set so as to be able to configure a shape that suppresses cell remaining on the barrier surface 400 in relation to the predetermined distance w. In the present embodiment, the temperature is set to be equal to or lower than the lowest water level H of the culture solution to be stored in the culture chamber C.
　培養液面ＳにメニスカスＭが発生した場合、上下方向視からの視界は、培養室Ｃの周縁部分の培養液面ＳがメニスカスＭ現象によって曲面状となっているため、当該部分において観察困難となる。培養容器１００では、内部周縁Ｂ２から立ち上がる所定の高さｈの障壁面４００を設けることで、内部周縁Ｂ２から障壁面４００側への培養細胞の移動を阻止し、障壁面４００を内部周縁Ｂ２から外方向へ向かわせて内部周縁Ｂ２と周側壁面３００とを所定の距離ｗだけ離間させることで、メニスカスＭが発生する部分の少なくとも一部を、内部周縁Ｂ２の外へ逃がす。したがって、メニスカスＭの発生を許容しながらも、培養した細胞の大部分を観察可能な内部周縁Ｂ２内部に存在させることができる。 When the meniscus M is generated on the culture liquid surface S, the field of view from the vertical direction is difficult to observe because the culture liquid surface S in the peripheral part of the culture chamber C is curved due to the meniscus M phenomenon. Become. In the culture vessel 100, by providing the barrier surface 400 having a predetermined height h rising from the inner peripheral edge B2, the culture cell is prevented from moving from the inner peripheral edge B2 to the barrier surface 400, and the barrier surface 400 is moved from the inner peripheral edge B2. By separating the inner peripheral edge B2 and the peripheral side wall surface 300 by a predetermined distance w in the outward direction, at least a part of the portion where the meniscus M is generated escapes from the inner peripheral edge B2. Therefore, while allowing the generation of the meniscus M, most of the cultured cells can be present inside the observable inner peripheral edge B2.
　障壁面４００が所定の距離ｗに相当する幅を有することにより、培養室Ｃ内に培養液を収容した場合に培養液面Ｓに発生するメニスカスＭの影響が少なくとも低減される。したがって、当該所定の距離ｗは特に限定されない。たとえば、所定の距離ｗが大きいほど、メニスカスＭの影響をより低減する傾向にあり、所定の距離ｗが小さいほど、培養および観察のための底面２００上のスペースを十分にとることができる傾向にあることに基づいて、当業者が適宜決定することができる。具体例として、所定の距離ｗは２ｍｍ以上３ｍｍ以下であってよい。本実施形態においては、メニスカスＭの影響を回避する観点から、所定の距離ｗは、メニスカスＭの幅Ｗ以上となるように設定される。 When the barrier surface 400 has a width corresponding to the predetermined distance w, at least the influence of the meniscus M generated on the culture solution surface S when the culture solution is accommodated in the culture chamber C is reduced. Therefore, the predetermined distance w is not particularly limited. For example, the larger the predetermined distance w, the more the influence of the meniscus M tends to be reduced. The smaller the predetermined distance w, the more the space on the bottom surface 200 for culture and observation tends to be sufficient. Those skilled in the art can appropriately determine the situation based on the fact. As a specific example, the predetermined distance w may be 2 mm or more and 3 mm or less. In the present embodiment, from the viewpoint of avoiding the influence of the meniscus M, the predetermined distance w is set to be equal to or larger than the width W of the meniscus M.
　所定の高さｈは、所定の距離ｗとの関係で障壁面４００への細胞残存を抑制する落差を構成可能となるように設定されればよいため、特に限定されない。たとえば、所定の高さｈは、１ｍｍ以上２１ｍｍ以下、好ましくは１ｍｍ以上１５ｍｍ以下である。上記下限値以上であることにより、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の侵入をより少なくすることができ、上記上限値以下であることにより、培養液を収容すべき容積を十分にとることができる。本実施形態においては、培養液を収容すべき容積をより十分にとる観点から、所定の高さｈは、培養室Ｃ内に収容すべき培養液の水位Ｈ以下となるように設定される。この場合、培養液面Ｓと培養容器１００の内壁面との境界Ｅは、周側壁面３００上にある。 The predetermined height h is not particularly limited, as long as it can be set so as to be able to configure a drop that suppresses cell remaining on the barrier surface 400 in relation to the predetermined distance w. For example, the predetermined height h is 1 mm or more and 21 mm or less, preferably 1 mm or more and 15 mm or less. By being above the lower limit value, it is possible to further reduce the invasion of the cultured cells into the outer part of the inner peripheral edge B2, which is difficult to observe in the vertical direction. A sufficient volume can be taken. In the present embodiment, the predetermined height h is set to be equal to or lower than the water level H of the culture solution to be stored in the culture chamber C from the viewpoint of taking a sufficient volume to store the culture solution. In this case, the boundary E between the culture liquid surface S and the inner wall surface of the culture vessel 100 is on the peripheral wall surface 300.
　図４に示すように、障壁面４００は、下側障壁面部４１０と上側障壁面部４２０とから構成される。下側障壁面部４１０は、底面２００と角度θ１をなすように内部周縁Ｂ２を基端として立ち上がり、高さｈ１の位置で上側障壁面部４２０に連設されている。高さｈ１の位置において、下側障壁面部４１０と上側障壁面部４２０との境界は、稜線をなしている。上側障壁面部４２０は、底面２００に対して角度θ２で周側壁面３００に向かって傾斜し、周側壁面３００と境界Ｂ３をなす。 As shown in FIG. 4, the barrier surface 400 includes a lower barrier surface portion 410 and an upper barrier surface portion 420. The lower barrier surface portion 410 rises with the inner peripheral edge B2 as a base end so as to form an angle θ1 with the bottom surface 200, and is connected to the upper barrier surface portion 420 at a height h1. At the position of the height h1, the boundary between the lower barrier surface portion 410 and the upper barrier surface portion 420 forms a ridge line. The upper barrier surface portion 420 is inclined toward the peripheral side wall surface 300 at an angle θ2 with respect to the bottom surface 200, and forms a boundary B3 with the peripheral side wall surface 300.
　下側障壁面部４１０が底面２００となす角度θ１は、９０度である。これによって、底面２００における培養細胞が内部周縁Ｂ２を超えて外側へ移動することを効果的に防ぐことができる。上側障壁面部４２０の傾斜角である角度θ２は、特に限定されない。たとえば、角度θ２が大きいほど、下側障壁面部４１０の高さｈ１を十分確保して、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の侵入をより少なくすることができる傾向にあり、角度θ２が小さいほど、培養液を収容すべき容積を十分にとることができる傾向にあることに基づいて、当業者が適宜決定することができる。 The angle θ1 between the lower barrier surface portion 410 and the bottom surface 200 is 90 degrees. Thereby, it is possible to effectively prevent the cultured cells on the bottom surface 200 from moving beyond the inner peripheral edge B2. The angle θ2 that is the inclination angle of the upper barrier surface portion 420 is not particularly limited. For example, as the angle θ2 is larger, the height h1 of the lower barrier surface portion 410 is sufficiently secured, and the invasion of the cultured cells into the outer portion of the inner peripheral edge B2 that is difficult to observe in the vertical direction can be reduced. It can be determined appropriately by those skilled in the art based on the tendency that the smaller the angle θ2 is, the more the volume for storing the culture solution can be taken.
　下側障壁面部４１０の高さｈ１は、特に限定されない。たとえば、高さｈ１が大きいほど、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の侵入がより少なくなる傾向にあり、高さｈ１が小さいほど、培養液を収容すべき容積を十分にとることができる傾向に基づいて、当業者が適宜決定することができる。具体的には、高さｈ１は、たとえば０．５ｍｍに設定してもよいし、１ｍｍに設定してもよい。 The height h1 of the lower barrier surface portion 410 is not particularly limited. For example, the larger the height h1, the smaller the invasion of the cultured cells into the outer part of the inner peripheral edge B2, which is difficult to observe in the vertical direction, and the smaller the height h1, the more the culture solution should be accommodated. Those skilled in the art can appropriately determine the tendency based on the tendency to take a sufficient volume. Specifically, the height h1 may be set to, for example, 0.5 mm or 1 mm.
　内部周縁Ｂ２の直径φは、従来の培養容器の内底面の直径と同等であってよい。たとえば、３０ｍｍ以上１７０ｍｍ以下、好ましくは３２ｍｍ以上９０ｍｍ以下である。上記下限値以上であることにより、培養および観察のための底面２００上のスペースを好ましくとることができ、上記上限値以下であることにより、培養した細胞の大部分を観察可能な部分に集める効果を好ましく享受することができる。 The diameter φ of the inner peripheral edge B2 may be equal to the diameter of the inner bottom surface of the conventional culture vessel. For example, it is 30 mm or more and 170 mm or less, preferably 32 mm or more and 90 mm or less. By being above the lower limit value, it is possible to preferably take a space on the bottom surface 200 for culture and observation, and by being below the upper limit value, an effect of collecting most of the cultured cells in an observable portion. Can be enjoyed preferably.
　培養容器１００は、透明性を有する素材で構成されることができる。透明性を有する素材としては、ガラスおよび石英に代表される無機物、ならびに合成樹脂に代表される有機物が挙げられる。合成樹脂としては、ポリスチレン（ＰＳ）、ポリプロピレン（ＰＰ）、ポリメチルペンテン（ＰＭＰ）、ポリカーボネート（ＰＣ）、ポリメチルメタクリレート（ＰＭＭＡ）、ポリメチルアクリルメチルイミド（ＰＭＭＩ）、およびシクロオレフィンコポリマー（ＣＯＣ）などのポリマーが挙げられる。さらに、これらのポリマーのモノマーユニットの２以上から合成される共重合体も挙げられる。合成樹脂の場合、成形が容易であり、かつ割れにくい点で好ましい。 The culture vessel 100 can be made of a transparent material. Examples of the material having transparency include inorganic substances typified by glass and quartz, and organic substances typified by synthetic resins. Synthetic resins include polystyrene (PS), polypropylene (PP), polymethylpentene (PMP), polycarbonate (PC), polymethyl methacrylate (PMMA), polymethylacrylmethylimide (PMMI), and cycloolefin copolymer (COC). And the like. Furthermore, the copolymer synthesize | combined from two or more of the monomer units of these polymers is also mentioned. In the case of a synthetic resin, it is preferable because it is easy to mold and is difficult to break.
　培養容器１００は、一体成型による構造体であってもよいし、たとえば底面２００を構成する部材と周側壁面３００および障壁面４００を構成する部材との組み合わせによる構造体であってもよい。部材の組み合わせによる構造体である場合、それぞれの部材が異なる素材で構成されていてもよい。 The culture vessel 100 may be a structure formed by integral molding, or may be a structure formed by a combination of a member forming the bottom surface 200 and members forming the peripheral wall surface 300 and the barrier surface 400, for example. When the structure is a combination of members, each member may be made of different materials.
　培養容器１００の底面２００、周側壁面３００および障壁面４００には、物理的、化学的および／または生化学観点から、表面処理が施されてよい。このような表面処理は、当業者であれば適宜選択することができる。たとえば、障壁面４００、または障壁面４００および周側壁面３００に、細胞培養を阻害するための表面処理が施されていてよい。 The bottom surface 200, the peripheral wall surface 300, and the barrier surface 400 of the culture vessel 100 may be subjected to a surface treatment from the viewpoint of physical, chemical, and / or biochemistry. Such a surface treatment can be appropriately selected by those skilled in the art. For example, the barrier surface 400 or the barrier surface 400 and the peripheral wall surface 300 may be subjected to a surface treatment for inhibiting cell culture.
　さらに、培養容器１００の使用目的などに応じ、図示しない更なる付加的構造を設けてもよい。このような付加的構造は、当業者であれば適宜選択することができる。具体的には、培養室Ｃを施蓋状態とするための蓋体、内容物を仕切るための仕切りおよび内容物（特に細胞群）の量および位置を特定するためのグリッドパターンなどの、底面２００に設けられる案内構造、が挙げられる。 Furthermore, an additional structure (not shown) may be provided depending on the purpose of use of the culture vessel 100 or the like. Such an additional structure can be appropriately selected by those skilled in the art. Specifically, the bottom surface 200 such as a lid for placing the culture chamber C in a covered state, a partition for partitioning the contents, and a grid pattern for specifying the amount and position of the contents (particularly cell groups). The guide structure provided in the above.
＜変形例＞
　培養容器１００は、図示したもののほかに様々な変形例が許容される。変形例においては、メニスカスＭの少なくとも一部が内部周縁Ｂ２よりも外側に逃がされればよいため、境界Ｅが、障壁面４００のうち、内部周縁Ｂ２を基端として底面２００に対して９０度をなして立ち上がる下側障壁面４１０上で形成されない限り、いかなる構造も許容できる。つまり、変形例においては、培養室Ｃ内に培養液を収容した時に、培養液の液面Ｓと培養容器１００の内壁との境界Ｅが内部周縁Ｂ２よりも外側となるように構成されればよい。 <Modification>
The culture vessel 100 can be variously modified in addition to the illustrated one. In the modified example, at least a part of the meniscus M only needs to escape to the outside of the inner peripheral edge B2, so that the boundary E is 90 degrees with respect to the bottom surface 200 with the inner peripheral edge B2 being the base end of the barrier surface 400. Any structure is acceptable as long as it is not formed on the rising lower barrier surface 410. That is, in the modification, when the culture solution is accommodated in the culture chamber C, the boundary E between the liquid surface S of the culture solution and the inner wall of the culture vessel 100 is outside the inner peripheral edge B2. Good.
　以下に、所定の距離ｗおよび所定の高さｈの組み合わせが異なる第１実施形態の変形例について、図５から図９を挙げて説明する。図５から図９は、第１実施形態の図３に対応する。以下の変形例においては、主に第１実施形態と異なる点について説明し、同一点については説明を省略する。 Hereinafter, modified examples of the first embodiment in which combinations of the predetermined distance w and the predetermined height h are different will be described with reference to FIGS. 5 to 9 correspond to FIG. 3 of the first embodiment. In the following modifications, differences from the first embodiment will be mainly described, and description of the same points will be omitted.
［第１変形例］
　図５に、第１実施形態の第１変形例を示す。図５に示す培養容器１００ａの障壁面４００ａは、所定の距離ｗａが第１実施形態における所定の距離ｗより短く設定されていることにより、基端となる内部周縁Ｂ２ａが第１実施形態における内部周縁Ｂ２より外側にある。これによって、内部周縁Ｂ２ａより外側に逃がされるメニスカスＭはその一部となるが、メニスカスＭによる影響は低減することができる。すなわち、本変形例においても、メニスカスＭは内部周縁Ｂ２ａより外側に逃がされるため、メニスカスＭによる影響を低減することができる。 [First Modification]
FIG. 5 shows a first modification of the first embodiment. The barrier surface 400a of the culture vessel 100a shown in FIG. 5 is set such that the predetermined distance wa is shorter than the predetermined distance w in the first embodiment, so that the inner peripheral edge B2a serving as the base end is the inner space in the first embodiment. It is outside the peripheral edge B2. Thereby, the meniscus M that escapes to the outside from the inner peripheral edge B2a becomes a part thereof, but the influence of the meniscus M can be reduced. That is, also in this modified example, the meniscus M is escaped to the outside from the inner peripheral edge B2a, so that the influence of the meniscus M can be reduced.
［第２変形例］
　図６に、第１実施形態の第２変形例を示す。図６に示す培養容器１００ｂの障壁面４００ｂは、所定の高さｈｂが第１実施形態における所定の高さｈより大きく設定されていることにより、周側壁面３００ｂとの境界Ｂ３ｂが第１実施形態における境界Ｂ３より上側にある。これによって、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の侵入をより少なくすることができる。本変形例では、所定の高さｈｂは、培養室Ｃ内に収容すべき培養液の水位Ｈ以上となるが、培養液面Ｓと周側壁面３００ｂとの境界Ｅの高さ未満となる。 [Second Modification]
FIG. 6 shows a second modification of the first embodiment. The barrier surface 400b of the culture vessel 100b shown in FIG. 6 has a predetermined height hb set to be larger than the predetermined height h in the first embodiment, so that the boundary B3b with the peripheral wall surface 300b is the first implementation. It is above the boundary B3 in the form. As a result, it is possible to further reduce the invasion of the cultured cells into the outer portion of the inner peripheral edge B2, which is difficult to observe when viewed in the vertical direction. In this modification, the predetermined height hb is equal to or higher than the water level H of the culture solution to be accommodated in the culture chamber C, but less than the height of the boundary E between the culture solution surface S and the peripheral side wall surface 300b.
［第３変形例］
　図７に、第１実施形態の第３変形例を示す。図７に示す培養容器１００ｃの障壁面４００ｃは、第１変形例と同様に、所定の距離ｗａが第１実施形態における所定の距離ｗより短く設定されていることにより、基端となる内部周縁Ｂ２ａが第１実施形態における内部周縁Ｂ２より外側にある。これによって、内部周縁Ｂ２ａより外側に逃がされるメニスカスＭはその一部となるが、メニスカスＭによる影響は低減することができる。すなわち、本変形例においても、メニスカスＭは内部周縁Ｂ２ａより外側に逃がされるため、メニスカスＭによる影響を低減することができる。 [Third Modification]
FIG. 7 shows a third modification of the first embodiment. As in the first modification, the barrier surface 400c of the culture vessel 100c shown in FIG. 7 has an inner peripheral edge serving as a base end when the predetermined distance wa is set shorter than the predetermined distance w in the first embodiment. B2a is outside the inner peripheral edge B2 in the first embodiment. Thereby, the meniscus M that escapes to the outside from the inner peripheral edge B2a becomes a part thereof, but the influence of the meniscus M can be reduced. That is, also in this modified example, the meniscus M is escaped to the outside from the inner peripheral edge B2a, so that the influence of the meniscus M can be reduced.
　さらに、障壁面４００ｃは、第２変形例と同様に、周側壁面３００ｂとの境界Ｂ３ｂが第１実施形態における境界Ｂ３より上側にある。これによって、上下方向視で観察困難となる内部周縁Ｂ２ａの外側部分への培養細胞の侵入をより少なくすることができる。本変形例では、所定の高さｈｂは、培養室Ｃ内に収容すべき培養液の水位Ｈ以上となるが、培養液面Ｓと周側壁面３００ｂとの境界Ｅの高さ未満となる。 Furthermore, the barrier surface 400c has a boundary B3b with the peripheral side wall surface 300b above the boundary B3 in the first embodiment, as in the second modification. As a result, it is possible to further reduce the invasion of the cultured cells into the outer portion of the inner peripheral edge B2a, which is difficult to observe in the vertical direction. In this modification, the predetermined height hb is equal to or higher than the water level H of the culture solution to be accommodated in the culture chamber C, but less than the height of the boundary E between the culture solution surface S and the peripheral side wall surface 300b.
［第４変形例］
　図８に、第１実施形態の第４変形例を示す。図８に示す培養容器１００ｄの障壁面４００ｄは、所定の高さｈｄが第２変形例における所定の高さｈｂよりさらに大きく設定されていることにより、周側壁面３００ｄとの境界Ｂ３ｄが第２変形例における境界Ｂ３ｂよりさらに上側にある。これによって、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の侵入をより少なくすることができる。 [Fourth Modification]
FIG. 8 shows a fourth modification of the first embodiment. The barrier surface 400d of the culture vessel 100d shown in FIG. 8 is set such that the predetermined height hd is larger than the predetermined height hb in the second modified example, so that the boundary B3d with the peripheral wall surface 300d is the second. It is further above the boundary B3b in the modification. As a result, it is possible to further reduce the invasion of the cultured cells into the outer portion of the inner peripheral edge B2, which is difficult to observe when viewed in the vertical direction.
　本変形例では、培養液面Ｓと培養容器１００ｄの内壁面との境界Ｅｄが上側周側壁面部４２０ｄ上にあるため、障壁面４００ｄの所定の高さｈｄが培養室Ｃ内に収容すべき培養液の水位Ｈ超かつ境界Ｅｄの高さ以上となる。しかし、所定の距離ｗが十分量設けられているため、メニスカスＭを内部周縁Ｂ２の外側へ逃がすことができ、メニスカスＭの影響を回避することができる。 In this modification, since the boundary Ed between the culture liquid surface S and the inner wall surface of the culture vessel 100d is on the upper peripheral wall surface portion 420d, the predetermined height hd of the barrier surface 400d is to be accommodated in the culture chamber C. The liquid level exceeds H and exceeds the height of the boundary Ed. However, since a sufficient amount of the predetermined distance w is provided, the meniscus M can escape to the outside of the inner peripheral edge B2, and the influence of the meniscus M can be avoided.
［第５変形例］
　図９に、第１実施形態の第５変形例を示す。図９に示す培養容器１００ｅの障壁面４００ｅは、第４変形例と同様に、所定の高さｈｄが第２，第３変形例における所定の高さｈｂよりさらに大きく設定されていることにより、周側壁面３００ｄとの境界Ｂ３ｄが第２，第３変形例における境界Ｂ３ｂよりさらに上側にある。これによって、上下方向視で観察困難となる内部周縁Ｂ２ａの外側部分への培養細胞の侵入をより少なくすることができる。 [Fifth Modification]
FIG. 9 shows a fifth modification of the first embodiment. As in the fourth modification, the barrier surface 400e of the culture vessel 100e shown in FIG. 9 is set such that the predetermined height hd is larger than the predetermined height hb in the second and third modifications. The boundary B3d with the peripheral side wall surface 300d is further above the boundary B3b in the second and third modified examples. As a result, it is possible to further reduce the invasion of the cultured cells into the outer portion of the inner peripheral edge B2a, which is difficult to observe when viewed in the vertical direction.
　本変形例では、培養液面Ｓと培養容器１００ｅの内壁面との境界Ｅｅが上側周側壁面部４２０ｅ上にあるため、障壁面４００ｅの所定の高さｈｄが培養室Ｃ内に収容すべき培養液の水位Ｈ超かつ境界Ｅｅの高さ以上となる。さらに、第１変形例と同様に、所定の距離ｗａが第１実施形態における所定の距離ｗより短く設定されていることにより、基端となる内部周縁Ｂ２ａが第１実施形態における内部周縁Ｂ２より外側にある。これによって、内部周縁Ｂ２ａより外側に逃がされるメニスカスＭはその一部となるが、メニスカスＭによる影響は低減することができる。すなわち、本変形例においても、メニスカスＭは内部周縁Ｂ２ａより外側に逃がされるため、メニスカスＭによる影響を低減することができる。 In this modification, since the boundary Ee between the culture liquid surface S and the inner wall surface of the culture vessel 100e is on the upper peripheral wall surface portion 420e, the predetermined height hd of the barrier surface 400e is to be accommodated in the culture chamber C. The liquid level exceeds H and exceeds the boundary Ee. Further, as in the first modification, the predetermined distance wa is set shorter than the predetermined distance w in the first embodiment, so that the inner peripheral edge B2a serving as the base end is more than the inner peripheral edge B2 in the first embodiment. On the outside. Thereby, the meniscus M that escapes to the outside from the inner peripheral edge B2a becomes a part thereof, but the influence of the meniscus M can be reduced. That is, also in this modified example, the meniscus M is escaped to the outside from the inner peripheral edge B2a, so that the influence of the meniscus M can be reduced.
＜他の実施形態＞
　本発明において、障壁面４００は、横方向からの断面視において内部周縁Ｂ２と境界Ｂ３との間を通る形状（内部周縁Ｂ２と境界Ｂ３とを接続する形状）を有していればよく、第１実施形態に示した形状に限定されるものではない。たとえば、下側障壁面部４１０と上側障壁面部４２０との境界は、図４に示したように稜線をなしていてもよいし、当該境界がＣ面取り加工またはＲ面取り加工されていてもよい。 <Other embodiments>
In the present invention, the barrier surface 400 only needs to have a shape that passes between the inner peripheral edge B2 and the boundary B3 (a shape that connects the inner peripheral edge B2 and the boundary B3) in a cross-sectional view from the lateral direction. It is not limited to the shape shown in one embodiment. For example, the boundary between the lower barrier surface portion 410 and the upper barrier surface portion 420 may form a ridge line as shown in FIG. 4, or the boundary may be C chamfered or R chamfered.
　さらには、障壁面４００の横方向から見た断面形状は、直線であってもよいし、曲線であってもよいし、直線と曲線との組み合わせであってもよく、様々な形状が広く許容される。好ましくは、障壁面４００は、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の不在をより確実にする観点から、横方向からの断面視において、内部周縁Ｂ２と境界Ｂ３とを結ぶ直線またはそれより上に存在する形状である。 Furthermore, the cross-sectional shape viewed from the lateral direction of the barrier surface 400 may be a straight line, a curved line, or a combination of a straight line and a curved line, and various shapes are widely acceptable. Is done. Preferably, the barrier surface 400 has the inner peripheral edge B2 and the boundary B3 in the cross-sectional view from the lateral direction, from the viewpoint of more surely preventing the cultured cells from being present on the outer portion of the inner peripheral edge B2, which is difficult to observe in the vertical direction. A shape existing on or above a straight line connecting the two.
　以下において、障壁面４００の形状が第１実施形態と異なる第２実施形態から第７実施形態について、図１０から図１５を挙げて説明する。図１０から図１５は、第１実施形態の図４に対応する。以下の実施形態においては、主に第１実施形態と異なる点について説明し、同一点については説明を省略する。 Hereinafter, second to seventh embodiments, in which the shape of the barrier surface 400 is different from that of the first embodiment, will be described with reference to FIGS. 10 to 15. 10 to 15 correspond to FIG. 4 of the first embodiment. In the following embodiments, differences from the first embodiment will be mainly described, and description of the same points will be omitted.
［第２実施形態］
　図１０は、第２実施形態の培養容器の要部拡大図を示す。図１０に示す障壁面４００ｆは、下側障壁面部４１０ｆと上側障壁面部４２０ｆとから構成される。下側障壁面部４１０ｆは、底面２００と角度θ１（９０度）をなすように内部周縁Ｂ２を基端として立ち上がり、底面２００と並行な（角度θ２が１８０度）上側障壁面部４２０ｆに連設されている。なお、図１０では、下側障壁面部４１０ｆと上側障壁面部４２０ｆとの境界が稜線をなしているが、当該境界がＣ面取り加工またはＲ面取り加工されていてもよい。 [Second Embodiment]
FIG. 10 shows an enlarged view of the main part of the culture vessel of the second embodiment. The barrier surface 400f shown in FIG. 10 includes a lower barrier surface portion 410f and an upper barrier surface portion 420f. The lower barrier surface portion 410f rises with the inner peripheral edge B2 as a base end so as to form an angle θ1 (90 degrees) with the bottom surface 200, and is connected to the upper barrier surface portion 420f parallel to the bottom surface 200 (angle θ2 is 180 degrees). Yes. In FIG. 10, the boundary between the lower barrier surface portion 410f and the upper barrier surface portion 420f forms a ridge line, but the boundary may be C-chamfered or R-chamfered.
［第３実施形態］
　図１１は、第３実施形態の培養容器の要部拡大図を示す。図１１に示す障壁面４００ｇは、下側障壁面部４１０ｇと上側障壁面部４２０ｇとから構成される。下側障壁面部４１０ｇは、底面２００と角度θ１をなすように内部周縁Ｂ２を基端として立ち上がり、底面２００と並行な（角度θ２が１８０度）上側障壁面部４２０ｇに連設されている。 [Third Embodiment]
FIG. 11: shows the principal part enlarged view of the culture container of 3rd Embodiment. The barrier surface 400g shown in FIG. 11 includes a lower barrier surface portion 410g and an upper barrier surface portion 420g. The lower barrier surface portion 410g rises with the inner peripheral edge B2 as a base end so as to form an angle θ1 with the bottom surface 200, and is connected to the upper barrier surface portion 420g parallel to the bottom surface 200 (angle θ2 is 180 degrees).
　本実施形態において、角度θ１は、９０度超１５０度以下であってよい。当該下限値超であることにより、上下方向視から、培養および観察すべき内部周縁Ｂ２内の領域全体を視界良好に観察することができ、当該上限値以下であることにより、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の進入を少なくすることができる。なお、図１１では、下側障壁面部４１０ｇと上側障壁面部４２０ｇとの境界が稜線をなしているが、当該境界がＣ面取り加工またはＲ面取り加工されていてもよい。 In the present embodiment, the angle θ1 may be greater than 90 degrees and less than or equal to 150 degrees. By being above the lower limit value, it is possible to observe the entire region within the inner peripheral edge B2 to be cultured and observed from the vertical direction, and to observe the visual field well, and by being below the upper limit value, the vertical region is observed. It is possible to reduce the entry of cultured cells into the outer portion of the inner peripheral edge B2, which is difficult. In FIG. 11, the boundary between the lower barrier surface portion 410g and the upper barrier surface portion 420g forms a ridge line, but the boundary may be C-chamfered or R-chamfered.
［第４実施形態］
　図１２は、第４実施形態の培養容器の要部拡大図を示す。図１２に示す障壁面４００ｈは、下側障壁面部４１０ｈと上側障壁面部４２０ｈとから構成される。下側障壁面部４１０ｈが底面２００となす角度θ１と、上側障壁面部４２０ｈが底面２００となす角度θ２とは同一であるため、両面は境界なく連設されている。このため、障壁面４００ｈの構造が簡素であり、製造上の観点からも好ましい。 [Fourth Embodiment]
FIG. 12 shows an enlarged view of the main part of the culture vessel of the fourth embodiment. The barrier surface 400h shown in FIG. 12 includes a lower barrier surface portion 410h and an upper barrier surface portion 420h. Since the angle θ1 formed by the lower barrier surface portion 410h and the bottom surface 200 is the same as the angle θ2 formed by the upper barrier surface portion 420h and the bottom surface 200, both surfaces are connected without boundaries. For this reason, the structure of the barrier surface 400h is simple, which is preferable from the viewpoint of manufacturing.
［第５実施形態］
　図１３は、第５実施形態の培養容器の要部拡大図を示す。図１３に示す障壁面４００iは、横方向から見た断面形状が曲面である。当該曲面の接線と底面２００とのなす角度θ，θ’，θ’’は、底面２００の側から周側壁面３００の側へ向かって漸次大きくなる（θ＜θ’＜θ’’）。これによって、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の不在をより確実にすることができるとともに、障壁面４００iの構造自体も簡素であり、製造上の観点からも好ましい。なお、角度θ，θ’，θ’’は、９０度以上１８０度以下の範囲で変化させることができる。 [Fifth Embodiment]
FIG. 13: shows the principal part enlarged view of the culture container of 5th Embodiment. The barrier surface 400i shown in FIG. 13 has a curved surface when viewed from the lateral direction. The angles θ, θ ′, θ ″ formed by the tangent to the curved surface and the bottom surface 200 gradually increase from the bottom surface 200 side toward the peripheral side wall surface 300 (θ <θ ′ <θ ″). As a result, the absence of cultured cells in the outer portion of the inner peripheral edge B2, which is difficult to observe in the vertical direction, can be further ensured, and the structure of the barrier surface 400i itself is simple, and from the viewpoint of manufacturing. preferable. The angles θ, θ ′, and θ ″ can be changed in the range of 90 degrees to 180 degrees.
［第６実施形態］
　図１４は、第６実施形態の培養容器の要部拡大図を示す。図１４に示す障壁面４００ｊは、下側障壁面部４１０ｊと上側障壁面部４２０ｊとから構成される。下側障壁面部４１０ｊは、横方向から見た断面形状が直線であり、底面２００と角度θ１をなすように内部周縁Ｂ２を基端として立ち上がり、高さｈ１の位置で、当該断面形状が曲線である上側障壁面部４２０ｊに連設されている。高さｈ１の位置において、下側障壁面部４１０ｊと上側障壁面部４２０ｊとの境界は、稜線をなしている。 [Sixth Embodiment]
FIG. 14: shows the principal part enlarged view of the culture container of 6th Embodiment. The barrier surface 400j shown in FIG. 14 includes a lower barrier surface portion 410j and an upper barrier surface portion 420j. The lower barrier surface portion 410j has a straight cross-sectional shape viewed from the lateral direction, rises with the inner peripheral edge B2 as the base end so as to form an angle θ1 with the bottom surface 200, and the cross-sectional shape is a curve at a position of height h1. It is connected to a certain upper barrier surface portion 420j. At the position of the height h1, the boundary between the lower barrier surface portion 410j and the upper barrier surface portion 420j forms a ridge line.
　本実施形態において、角度θ１は、９０度以上１５０度以下であってよい。当該下限値以上であることにより、上下方向視から、培養および観察すべき内部周縁Ｂ２内の領域全体を視界良好に観察することができ、当該上限値以下であることにより、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の不在をより確実にすることができる。上側障壁面部４２０ｊの断面形状は、第５実施形態における障壁面４００iと同様の曲面であってよい。 In the present embodiment, the angle θ1 may be not less than 90 degrees and not more than 150 degrees. By being above the lower limit value, the entire region within the inner peripheral edge B2 to be cultured and observed can be observed with good visibility from the vertical direction view. The absence of cultured cells in the outer portion of the inner peripheral edge B2, which becomes difficult, can be further ensured. The cross-sectional shape of the upper barrier surface portion 420j may be a curved surface similar to the barrier surface 400i in the fifth embodiment.
［第７実施形態］
　図１５は、第７実施形態の培養容器の要部拡大図を示す。図１５に示す障壁面４００ｋは、下側障壁面部４１０ｋと上側障壁面部４２０ｋとから構成される。下側障壁面部４１０ｋは、底面２００と角度θ１をなすように内部周縁Ｂ２を基端として立ち上がり、高さｈ１の位置で上側障壁面部４２０ｋに連設されている。高さｈ１の位置において、下側障壁面部４１０ｋと上側障壁面部４２０ｋとの境界は、稜線をなしている。上側障壁面部４２０ｋは、底面２００に対して角度θ２で周側壁面３００に向かって傾斜し、周側壁面３００と境界Ｂ３をなす。 [Seventh Embodiment]
FIG. 15: shows the principal part enlarged view of the culture container of 7th Embodiment. The barrier surface 400k shown in FIG. 15 includes a lower barrier surface portion 410k and an upper barrier surface portion 420k. The lower barrier surface portion 410k rises with the inner peripheral edge B2 as a base end so as to form an angle θ1 with the bottom surface 200, and is continuous with the upper barrier surface portion 420k at a height h1. At the position of the height h1, the boundary between the lower barrier surface portion 410k and the upper barrier surface portion 420k forms a ridge line. The upper barrier surface portion 420k is inclined toward the peripheral side wall surface 300 at an angle θ2 with respect to the bottom surface 200, and forms a boundary B3 with the peripheral side wall surface 300.
　本実施形態において、角度θ１は、９０度以上１５０度以下であってよい。当該下限値以上であることにより、上下方向視から、培養および観察すべき内部周縁Ｂ２内の領域全体を視界良好に観察することができ、当該上限値以下であることにより、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の進入を少なくすることができる。 In the present embodiment, the angle θ1 may be not less than 90 degrees and not more than 150 degrees. By being above the lower limit value, the entire region within the inner peripheral edge B2 to be cultured and observed can be observed with good visibility from the vertical direction view. It is possible to reduce the entry of cultured cells into the outer portion of the inner peripheral edge B2, which is difficult.
　角度θ２は特に限定されない。たとえば、角度θ２が大きいほど、下側障壁面部４１０ｋの高さｈ１を十分確保して、上下方向視で観察困難となる内部周縁Ｂ２の外側部分への培養細胞の進入を少なくすることができる傾向にあり、角度θ２が小さいほど、培養液を収容すべき容積を十分にとることができる傾向にあることに基づいて、当業者が適宜決定することができる。
　なお、図１５では、下側障壁面部４１０ｋと上側障壁面部４２０ｋとの境界が稜線をなしているが、当該境界がＣ面取り加工またはＲ面取り加工されていてもよい。 The angle θ2 is not particularly limited. For example, as the angle θ2 is larger, the height h1 of the lower barrier surface portion 410k is sufficiently secured, and the tendency of the cultured cells to enter the outer portion of the inner peripheral edge B2, which is difficult to observe in the vertical direction, can be reduced. Therefore, the smaller the angle θ2, the more appropriate it can be determined by those skilled in the art based on the tendency that a sufficient volume for accommodating the culture solution tends to be taken.
In FIG. 15, the boundary between the lower barrier surface portion 410k and the upper barrier surface portion 420k forms a ridge line, but the boundary may be C-chamfered or R-chamfered.
＜変形例＞
　なお、上位の第２から第７実施形態においても、第１実施形態と同様に変形例が許容される。すなわち、メニスカスＭ（図３から図９参照）の少なくとも一部が内部周縁Ｂ２よりも外側に逃がされればよいため、境界Ｅ（図３から図９参照）が、障壁面４００ｆ，４００ｇ，４００ｈ，４００ｉ，４００ｊ，４００ｋのうち、内部周縁Ｂ２を基端として底面２００に対して９０度をなして立ち上がる下側障壁面４１０ｆ，４１０ｊ上で形成されない限り、いかなる構造も許容できる。つまり、変形例においては、培養室Ｃ（図３から図９参照）内に培養液を収容した時に、培養液の液面Ｓ（図３から図９参照）と培養容器の内壁との境界Ｅが内部周縁Ｂ２よりも外側となるように構成されればよい。 <Modification>
In the upper second to seventh embodiments, modifications are allowed as in the first embodiment. That is, since at least a part of the meniscus M (see FIGS. 3 to 9) needs to escape to the outside of the inner peripheral edge B2, the boundary E (see FIGS. 3 to 9) has the barrier surfaces 400f, 400g, 400h, Of 400i, 400j, and 400k, any structure is acceptable as long as it is not formed on the lower barrier surfaces 410f and 410j rising from the inner peripheral edge B2 at 90 degrees with respect to the bottom surface 200. That is, in the modified example, when the culture solution is stored in the culture chamber C (see FIGS. 3 to 9), the boundary E between the liquid surface S (see FIGS. 3 to 9) of the culture solution and the inner wall of the culture vessel. May be configured to be outside the inner peripheral edge B2.
［第８実施形態］
　図１６は、第８実施形態の培養容器の模式的外観斜視図である。培養容器１００ｌは、培養室Ｃを複数有する。本実施形態では１個の培養容器１００ｌが６個の培養室Ｃを有するが、培養室Ｃの数はこれに限定されない。たとえば、培養室Ｃの数が多いほど、培養した細胞の大部分を観察可能な部分に集める効果を好ましく享受することができる傾向にあり、培養室Ｃの数をある程度にとどめることで、培養および観察のための底面２００上のスペースを適切にとることができることに基づいて、当業者が適宜決定することができる。培養容器１００ｌの培養室Ｃの構造としては、第１実施形態から第７実施形態およびそれらの変形例のいずれにおけるものが適用されてもよい。 [Eighth Embodiment]
FIG. 16 is a schematic external perspective view of the culture vessel of the eighth embodiment. The culture vessel 100l has a plurality of culture chambers C. In this embodiment, one culture vessel 100l has six culture chambers C, but the number of culture chambers C is not limited to this. For example, as the number of the culture chambers C increases, it tends to be preferable to enjoy the effect of collecting most of the cultured cells in an observable portion. By limiting the number of the culture chambers C to some extent, Those skilled in the art can appropriately determine the space on the bottom surface 200 for observation based on the appropriate space. As the structure of the culture chamber C of the culture vessel 100l, the structure in any of the first to seventh embodiments and their modifications may be applied.
　以下に実施例を示し、本発明を具体的に説明するが、本発明は下記の実施例に制限されるものではない。 Hereinafter, the present invention will be specifically described with reference to examples. However, the present invention is not limited to the following examples.
＜実施例１＞
　ポリカーボネートを用いた切削加工品（無蓋無底円筒の内周面側を切削加工したもの）にポリスチレンの円板を貼り合わせることにより、図９に記載の構造を有する培養容器の試作品を作製した（なお、培養容器の本番品は、一体成形により作製してよい。以下、張り合わせにより製作した試作品も単に「培養容器」と記載する）。θ１は９０度、θ２は１２０度であった。 <Example 1>
A prototype of a culture vessel having the structure shown in FIG. 9 was prepared by laminating a polystyrene disc to a cut product using polycarbonate (in which the inner peripheral surface side of an open and bottomless cylinder was cut). (Note that the actual product of the culture vessel may be produced by integral molding. Hereinafter, the prototype produced by bonding is also simply referred to as “culture vessel”). θ1 was 90 degrees and θ2 was 120 degrees.
　作製した培養容器の培養室に、血清１０％含有培地３ｍＬを添加し、上下方向から培養室を観察した。図１７に上下方向視写真を示す。図１７に示すように、培養容器は、障壁面４００ｅを有することにより、メニスカスＭの大部分が内部周縁Ｂ２ａの外側に生じることにより、培養および観察のための内部周縁Ｂ２ａの内部において良好に視認できる範囲が後述の比較例１より拡大していることが確認された。 3 mL of medium containing 10% serum was added to the culture chamber of the prepared culture container, and the culture chamber was observed from the top and bottom. FIG. 17 shows a vertical view photograph. As shown in FIG. 17, the culture container has a barrier surface 400e, so that most of the meniscus M is generated outside the inner peripheral edge B2a, so that the culture container is well visible inside the inner peripheral edge B2a for culture and observation. It was confirmed that the range which can be expanded from the comparative example 1 mentioned later.
　作製した培養容器の培養室に、血清１０％含有培地３ｍLとヒト繊維芽細胞３８．４万個とを添加後、３７℃、５％ＣＯ２の条件下で２４時間培養し、倍率４０倍の顕微鏡で培養室中央部および周辺部を観察した。図１８に培養室中央部、図１９に培養室周辺部の顕微鏡観察写真を示す。図１８に示すように、細胞が正常に底面２００に接着していることが確認された。また、図１９に示すように、障壁面４００ｅにより、メニスカスＭの大部分が内部周縁Ｂ２ａの外側に生じ、培養および観察のための内部周縁Ｂ２ａの内部において観察可能な範囲が後述の比較例１より拡大していることが確認された。 After adding 3 mL of medium containing 10% serum and 3840 thousand human fibroblasts to the culture chamber of the culture vessel thus prepared, the culture was performed for 24 hours under conditions of 37 ° C. and 5% CO 2 . The center and periphery of the culture chamber were observed with a microscope. FIG. 18 shows a microscopic observation photograph of the center of the culture chamber and FIG. 19 shows the periphery of the culture chamber. As shown in FIG. 18, it was confirmed that the cells normally adhered to the bottom surface 200. Further, as shown in FIG. 19, the barrier surface 400e causes most of the meniscus M to be generated outside the inner peripheral edge B2a, and the range that can be observed inside the inner peripheral edge B2a for culture and observation is Comparative Example 1 described later. It was confirmed that it was expanding.
＜実施例２＞
　ポリカーボネートを用いた切削加工品（無蓋無底円筒の内周面側を切削加工したもの）にポリスチレンの円板を貼り合わせることにより、図９に記載の構造を有する培養容器の試作品を作成した。本実施例の培養容器は、θ２が１５０度であることを除き、実施例１の培養容器と同様である。 <Example 2>
A prototype of a culture vessel having the structure shown in FIG. 9 was prepared by laminating a polystyrene disc to a cut product using polycarbonate (in which the inner peripheral surface of an open-bottomed bottomless cylinder was cut). . The culture container of this example is the same as the culture container of Example 1 except that θ2 is 150 degrees.
　作製した培養容器の培養室に、血清１０％含有培地３ｍＬを添加し、上下方向から培養室を観察した。図２０に上下方向視写真を示す。図２０に示すように、培養容器は、障壁面４００ｅを有することにより、メニスカスＭによる影が内部周縁Ｂ２ａの外側に生じ、培養および観察のための内部周縁Ｂ２ａの内部において良好に視認できる範囲が後述の比較例１より拡大していることが確認された。 3 mL of medium containing 10% serum was added to the culture chamber of the prepared culture container, and the culture chamber was observed from the top and bottom. FIG. 20 shows a vertical view photograph. As shown in FIG. 20, the culture vessel has a barrier surface 400e, so that a shadow caused by the meniscus M is generated outside the inner peripheral edge B2a, and there is a range that can be favorably seen inside the inner peripheral edge B2a for culture and observation. It was confirmed that it was larger than Comparative Example 1 described later.
　作製した培養容器の培養室に、血清１０％含有培地３ｍLとヒト繊維芽細胞３８．４万個を添加後、３７℃、５％ＣＯ２の条件下で２４時間培養し、倍率４０倍の顕微鏡で培養室中央部および周辺部を観察した。図２１に培養室中央部、図２２に培養室周辺部の顕微鏡観察写真を示す。図２１に示すように、細胞が正常に底面２００に接着していることが確認された。また、図２２に示すように、障壁面４００ｅにより、メニスカスＭの大部分が内部周縁Ｂ２ａの外側に生じ、培養および観察のための内部周縁Ｂ２ａの内部において観察可能な範囲が後述の比較例１より拡大していることが確認された。 After adding 3 mL of medium containing 10% serum and 3840 human fibroblasts to the culture chamber of the culture vessel thus prepared, the culture was performed for 24 hours under conditions of 37 ° C. and 5% CO 2 , and a microscope with a magnification of 40 times. Then, the central part and the peripheral part of the culture chamber were observed. FIG. 21 shows a microscope observation photograph of the center of the culture chamber and FIG. 22 shows the periphery of the culture chamber. As shown in FIG. 21, it was confirmed that the cells normally adhered to the bottom surface 200. Further, as shown in FIG. 22, the barrier surface 400e causes most of the meniscus M to be generated outside the inner peripheral edge B2a, and the range that can be observed inside the inner peripheral edge B2a for culture and observation is Comparative Example 1 described later. It was confirmed that it was expanding.
　実施例１と本実施例２の培養室周辺部の顕微鏡観察写真（図１９および図２２）を比較すると、実施例２（図２２）の方が細胞を良好に視認できる範囲が広いことが確認された。本実施例２では、培養液面Ｓと培養容器１００ｅの内壁面との境界Ｅｅが上側周側壁面部４２０ｅ上にあり、さらに、障壁面４００ｅの所定の高さｈｄが、培養室Ｃ内に収容された培養液の水位Ｈを下回りかつ境界Ｅの高さも下回っていた。そのため、θ２が１２０度である実施例１に比べ、θ２が１５０度である本実施例２の方が、内部周縁Ｂ２ａの外側に生じさせるメニスカスＭの範囲がより大きくなり、本実施例２（図２２）の方が細胞を良好に視認できる範囲が広くなった。 Comparison of the microscopic observation photographs (FIGS. 19 and 22) in the periphery of the culture chamber of Example 1 and Example 2 confirms that Example 2 (FIG. 22) has a wider range in which the cells can be seen well. It was done. In Example 2, the boundary Ee between the culture liquid surface S and the inner wall surface of the culture vessel 100e is on the upper peripheral wall surface portion 420e, and the predetermined height hd of the barrier surface 400e is accommodated in the culture chamber C. The level of the culture broth was below the level H and the height of the boundary E was also below. Therefore, compared with Example 1 in which θ2 is 120 degrees, in Example 2 in which θ2 is 150 degrees, the range of meniscus M generated outside the inner peripheral edge B2a becomes larger, and this Example 2 ( In FIG. 22), the range in which the cells can be visually recognized is widened.
＜比較例１＞
　ポリカーボネートの無蓋無底円筒にポリスチレンの円板を貼り合わせるより、障壁面を有さない培養容器を作成した。この培養容器の模式的断面図を図２３に示す。 <Comparative Example 1>
A culture vessel having no barrier surface was prepared by attaching a polystyrene disk to a polycarbonate open-bottomed bottomless cylinder. A schematic cross-sectional view of this culture vessel is shown in FIG.
　作製した培養容器の培養室に、血清１０％含有培地３ｍＬを添加し、上下方向から培養室を観察した。図２４に、上下方向視写真を示す。図２４に示すように、メニスカスＭの全てが周縁Ｂの内側に生じ、培養および観察のための周縁Ｂの内部にメニスカスＭによる影が確認された。 3 mL of medium containing 10% serum was added to the culture chamber of the prepared culture container, and the culture chamber was observed from the top and bottom. FIG. 24 shows a vertical view photograph. As shown in FIG. 24, all of the meniscus M occurred inside the peripheral edge B, and a shadow due to the meniscus M was confirmed inside the peripheral edge B for culture and observation.
　作製した培養容器の培養室に、血清１０％含有培地３ｍLとヒト繊維芽細胞３８．４万個を添加後、３７℃、５％ＣＯ２の条件下で２４時間培養し、倍率４０倍の顕微鏡で培養室中央部および周辺部を観察した。図２５に培養室中央部、図２６に培養室周辺部の顕微鏡観察写真を示す。図２５に示すように、細胞が正常に底面２００に接着していることが確認された。また、図２６に示すように、メニスカスＭの全てが周縁Ｂの内側に生じ、培養および観察のための周縁Ｂの内部にメニスカスＭによる影が生じ、周縁Ｂ付近の細胞観察が困難であり、培養および観察のための周縁Ｂの内部において観察可能な範囲が狭いことが確認された。 After adding 3 mL of medium containing 10% serum and 3840 human fibroblasts to the culture chamber of the culture vessel thus prepared, the culture was performed for 24 hours under conditions of 37 ° C. and 5% CO 2 , and a microscope with a magnification of 40 times. Then, the central part and the peripheral part of the culture chamber were observed. FIG. 25 shows a microscope observation photograph of the center of the culture chamber, and FIG. 26 shows the periphery of the culture chamber. As shown in FIG. 25, it was confirmed that the cells normally adhered to the bottom surface 200. In addition, as shown in FIG. 26, all of the meniscus M is generated inside the peripheral edge B, and a shadow due to the meniscus M is generated inside the peripheral edge B for culture and observation, and cell observation in the vicinity of the peripheral edge B is difficult. It was confirmed that the observable range was narrow within the periphery B for culture and observation.
［第１実施形態から第８実施形態における各部と請求項の各構成要素との対応関係］
　本発明においては、培養容器１００，１００ｌが「培養容器」に相当し、底面２００が「底面」に相当し、周側壁面３００，３００ｂ，３００ｄが「周側壁面」に相当し、障壁面４００，４００ａ，４００ｂ，４００ｃ，４００ｄ，４００ｅ，４００ｆ，４００ｇ，４００ｈ，４００i，４００ｊ，４００ｋが「障壁面」に相当し、下側障壁面部４１０，４１０ｆ，４１０ｇ，４１０ｈ，４１０ｊ，４１０ｋが「下側障壁面部」に相当し、上側障壁面部４２０，４２０ｄ，４２０ｅ，４２０ｆ，４２０ｇ，４２０ｈ，４２０ｊ，４２０ｋが「上側障壁面部」に相当し、周縁Ｏが「周縁」に相当し、内部周縁Ｂ２，Ｂ２ａが「内部周縁」に相当し、境界Ｂ３，Ｂ３ｂ，Ｂ３ｄが（障壁面と周側壁面との）「境界」に相当し、培養室Ｃが「培養室」に相当し、角度θ１が（下側障壁面部と底面とがなす）「角度」に相当し、角度θ，θ’，θ’’が（障壁面の断面形状の接線と底面とがなす）「角度」に相当する。 [Correspondence Relationship Between Each Part in First to Eighth Embodiments and Each Component in Claims]
In the present invention, the culture vessels 100 and 100l correspond to “culture vessels”, the bottom surface 200 corresponds to “bottom surface”, the peripheral side wall surfaces 300, 300b, and 300d correspond to “peripheral side wall surfaces”, and the barrier surface 400. , 400a, 400b, 400c, 400d, 400e, 400f, 400g, 400h, 400i, 400j, 400k correspond to the “barrier surface”, and the lower barrier surface portions 410, 410f, 410g, 410h, 410j, 410k are “lower side”. The upper barrier surface portions 420, 420d, 420e, 420f, 420g, 420h, 420j, 420k correspond to the “upper barrier surface portion”, the peripheral edge O corresponds to the “periphery”, and the inner peripheral edges B2, B2a. Corresponds to the “inner periphery”, the boundaries B3, B3b, B3d correspond to “boundaries” (between the barrier surface and the peripheral side wall surface), and the culture chamber C is the “culture chamber”. The angle θ1 corresponds to the “angle” (the lower barrier surface portion and the bottom surface form), and the angles θ, θ ′, and θ ″ (the tangent of the cross-sectional shape of the barrier surface and the bottom surface form) “ Corresponds to “Angle”.
　本発明の好ましい実施形態は上記の通りであるが、本発明はそれらのみに限定されるものではなく、本発明の趣旨と範囲とから逸脱することのない様々な実施形態が他になされる。さらに、本実施形態において述べられる作用および効果は一例であり、本発明を限定するものではない。 Preferred embodiments of the present invention are as described above, but the present invention is not limited to them, and various other embodiments can be made without departing from the spirit and scope of the present invention. Furthermore, the operations and effects described in this embodiment are merely examples, and do not limit the present invention.
　本発明によれば、メニスカスが発生しても、当該メニスカスにより形成される影が、培養細胞の観察の支障にならない培養容器を提供することができる。 According to the present invention, even when a meniscus is generated, a culture container in which a shadow formed by the meniscus does not hinder the observation of cultured cells can be provided.
　１００，１００ｌ…培養容器、２００…底面、３００，３００ｂ，３００ｄ…周側壁面、４００，４００ａ，４００ｂ，４００ｃ，４００ｄ，４００ｅ，４００ｆ，４００ｇ，４００ｈ，４００i，４００ｊ，４００ｋ…障壁面、４１０，４１０ｆ，４１０ｇ，４１０ｈ，４１０ｊ，４１０ｋ…下側障壁面部、４２０，４２０ｄ，４２０ｅ，４２０ｆ，４２０ｇ，４２０ｈ，４２０ｊ，４２０ｋ…上側障壁面部、Ｏ，Ｂ…（底面の）周縁、Ｂ２，Ｂ２ａ…（底面の）内部周縁、Ｂ３，Ｂ３ｂ，Ｂ３ｄ…（障壁面と周側壁面との）境界、Ｃ…培養室、θ１…（下側障壁面部と底面とがなす）角度、θ，θ’，θ’’…（障壁面の断面形状の接線と底面とがなす）角度。 100, 100l ... culture vessel, 200 ... bottom surface, 300, 300b, 300d ... peripheral wall surface, 400, 400a, 400b, 400c, 400d, 400e, 400f, 400g, 400h, 400i, 400j, 400k ... barrier surface, 410, 410f, 410g, 410h, 410j, 410k ... lower barrier surface portion, 420, 420d, 420e, 420f, 420g, 420h, 420j, 420k ... upper barrier surface portion, O, B ... (bottom surface) peripheral edge, B2, B2a ... ( Inner peripheral edge (bottom surface), B3, B3b, B3d (boundary between barrier surface and peripheral wall surface), C ... culture chamber, θ1 (angle formed by lower barrier surface portion and bottom surface), θ, θ ', θ '' ... An angle formed by the tangent to the cross-sectional shape of the barrier surface and the bottom surface.

Claims (6)

  1. 　底面と、前記底面の周縁に設けられた周側壁面とを有する培養室を含み、
    　前記培養室内に、前記周縁に沿って所定の高さの障壁面が周設され、
    　前記障壁面が、前記底面上の、前記周縁より内側の内部周縁を基端として立ち上がり、前記周側壁面に向かうとともに、前記周側壁面との境界をなすように形成された、培養容器。     Including a culture chamber having a bottom surface and a peripheral side wall surface provided on a peripheral edge of the bottom surface,
    In the culture chamber, a barrier surface having a predetermined height is provided along the peripheral edge,
    A culture vessel in which the barrier surface rises from the inner peripheral edge inside the peripheral edge on the bottom surface as a base end, and is directed to the peripheral wall surface and forms a boundary with the peripheral wall surface.
  2. 　横方向からの断面視における前記障壁面の形状が、直線、曲線、および直線と曲線との組み合わせのいずれかである、請求項１に記載の培養容器。 The culture vessel according to claim 1, wherein the shape of the barrier surface in a cross-sectional view from the lateral direction is one of a straight line, a curved line, and a combination of a straight line and a curved line.
  3. 　横方向からの断面視において、前記障壁面が、前記内部周縁から立ち上がる下側障壁面部と、前記下部障壁面部に連設されかつ前記周側壁面との境界をなす上側障壁面部と、から構成され、前記内部周縁から立ち上がる下側障壁面部が、前記底面と９０度以上１５０度以下の角度をなす、請求項１または２に記載の培養容器。 In a cross-sectional view from the lateral direction, the barrier surface includes a lower barrier surface portion that rises from the inner peripheral edge, and an upper barrier surface portion that is connected to the lower barrier surface portion and forms a boundary with the peripheral side wall surface. The culture container according to claim 1 or 2, wherein the lower barrier surface portion rising from the inner peripheral edge forms an angle of 90 degrees to 150 degrees with the bottom surface.
  4. 　横方向からの断面視において、互いに同じ傾斜の前記下側障壁面部と前記上側障壁面部とが境界なく連設している、請求項３に記載の培養容器。 The culture vessel according to claim 3, wherein the lower barrier surface portion and the upper barrier surface portion having the same inclination in a cross-sectional view from the lateral direction are connected to each other without a boundary.
  5. 　前記断面視における前記障壁面の形状が曲線であり、前記底面側から前記周側壁面側に向かって前記曲線の接線と前記底面とのなす角度が漸次大きくなる、請求項２に記載の培養容器。 The culture vessel according to claim 2, wherein the shape of the barrier surface in the cross-sectional view is a curve, and an angle formed by a tangent to the curve and the bottom surface gradually increases from the bottom surface side toward the peripheral side wall surface side. .
  6. 　前記周側壁面および前記内部周縁の上面視形状が円であり、前記内部周縁の直径が３０ｍｍ以上１７０ｍｍ以下である、請求項１から５のいずれか１項に記載の培養容器。 The culture vessel according to any one of claims 1 to 5, wherein the shape of the top surface of the peripheral side wall surface and the inner peripheral edge is a circle, and the diameter of the inner peripheral edge is 30 mm or more and 170 mm or less.
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WO2017163378A1 (en) * 2016-03-24 2017-09-28 株式会社ニコン Culture vessel
CN112534038A (en) * 2018-05-30 2021-03-19 太阳生物科学股份公司 A hole for cultivating biomaterial

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