WO2018061795A1 - Récipient de manipulation de cellules - Google Patents

Récipient de manipulation de cellules Download PDF

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Publication number
WO2018061795A1
WO2018061795A1 PCT/JP2017/033209 JP2017033209W WO2018061795A1 WO 2018061795 A1 WO2018061795 A1 WO 2018061795A1 JP 2017033209 W JP2017033209 W JP 2017033209W WO 2018061795 A1 WO2018061795 A1 WO 2018061795A1
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WIPO (PCT)
Prior art keywords
drop
cell handling
handling container
drop forming
wall portion
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PCT/JP2017/033209
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English (en)
Japanese (ja)
Inventor
良至 藤枝
智紀 赤井
琢磨 馬塲
衛 山崎
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大日本印刷株式会社
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Application filed by 大日本印刷株式会社 filed Critical 大日本印刷株式会社
Priority to JP2018513026A priority Critical patent/JP6338037B1/ja
Publication of WO2018061795A1 publication Critical patent/WO2018061795A1/fr

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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
    • C12M3/00—Tissue, human, animal or plant cell, or virus culture apparatus

Definitions

  • Embodiments of the present disclosure relate to a cell handling container, and more particularly, to a cell handling container provided with a region for forming a drop.
  • the present application is Japanese Patent Application No. 2016-195121 filed on Sep. 30, 2016, Japanese Patent Application No. 2017-079258 filed on Apr. 12, 2017, and filed on Jun. 27, 2017. The priority is claimed based on Japanese Patent Application No. 2017-125395, which is incorporated herein by reference.
  • various effects are obtained by imparting hydrophilicity to the surface of the container and making the surface hydrophilic.
  • excellent initial adhesion and proliferation of cells can be obtained by making the surface of the container a hydrophilic surface.
  • the surface of the microwell is made hydrophilic so that bubbles generated in the microwell can be easily formed when the culture solution is added. It can be removed and workability can be improved.
  • a thick drop also referred to as a droplet
  • a pipette for washing and culturing.
  • the surface of the container is a hydrophilic surface, there is a problem that it is difficult to form a drop, and even if a drop is formed, the drop is easily crushed.
  • the embodiment of the present disclosure has been made in view of the above points, and an object thereof is to provide a cell handling container capable of easily forming a drop even on a hydrophilic surface.
  • a cell handling container is a cell handling container, and is provided with an upper surface that is a liquid drop formation region, and is provided continuously or intermittently along the circumferential direction of the upper surface and extends downward. And at least one drop forming portion having a side surface, wherein the side surface extends downward from the edge of the upper surface to a range of 5 ⁇ m or more in the vertical direction.
  • the minimum width of the upper surface is 0.5 mm or more and 10 mm or less in the horizontal direction.
  • the upper surface is a flat surface.
  • an angle formed between the upper surface and the side surface is 136 ° or less at an edge of the upper surface.
  • the cell handling container includes a container bottom, and a groove that surrounds the drop forming part is provided between the container bottom and the drop forming part, It is preferable that at least a part of the side surface of the drop forming portion constitutes one side surface of the groove portion.
  • the distance from the edge of the upper surface of the drop forming unit to the edge of the upper surface of the container bottom adjacent to the drop forming unit via the groove is 0. It is suitable that it is 01 mm or more.
  • the cell handling container includes a surrounding wall portion surrounding the drop forming portion, and an upper end surface of the surrounding wall portion is flush with the upper surface of the drop forming portion. It is preferable that
  • the distance from the edge of the upper surface of the drop forming portion to the outer edge of the upper end surface of the surrounding wall portion is preferably 1 mm or less. is there.
  • the cell handling container further includes a container bottom provided around the surrounding wall part, and between the container bottom part and the surrounding wall part, It is preferable that a groove portion surrounding the surrounding wall portion is provided.
  • the upper surface of the drop forming unit is flush with the upper surface of the container bottom.
  • At least the upper surface of the drop forming portion is a hydrophilic surface.
  • the water contact angle of the hydrophilic surface is less than 80 °.
  • the cell handling container it is preferable that there are a plurality of the drop forming portions, and the top surfaces of the plurality of drop forming portions are flush with each other.
  • the cell handling container further includes a microwell that contains a cell, and a liquid container that surrounds the microwell and contains a liquid, It is preferable that the bottom surface of the portion is flush with the top surface of the drop forming portion.
  • the cell handling container further includes a microwell that contains a cell, and a liquid container that surrounds the microwell and that contains a liquid. It is preferable that the bottom surface is flush with the top surface of the drop forming portion.
  • a cell handling container is a cell handling container, and includes a container bottom part, a drop forming part that is provided on the container bottom part and includes a recessed part that opens upward and a peripheral wall part that forms the recessed part. And the upper end portion and the concave portion of the peripheral wall portion serve as a liquid drop formation region, and in the vertical direction, the distance from the upper end portion of the peripheral wall portion to the upper surface of the container bottom portion is h1, and the peripheral wall portion When the distance from the upper end to the deepest part of the recess is h2, h1 + h2 ⁇ 0.2 mm.
  • a cell handling container is a cell handling container, and includes a container bottom, a drop that is provided on the container bottom, and has a recess that opens upward and a peripheral wall that forms the recess.
  • a forming portion, and a groove portion that is provided between the container bottom portion and the peripheral wall portion and surrounds the peripheral wall portion, and the upper end portion and the concave portion of the peripheral wall portion serve as a liquid drop forming region, and In the direction, h3 + h4 ⁇ 1 mm, where h3 is the distance from the upper end of the peripheral wall to the upper surface of the bottom of the container, and h4 is the distance from the upper end of the peripheral wall to the deepest part of the recess. It is characterized by that.
  • the outer wall surface of the peripheral wall portion extends downward from the edge of the upper end portion to a range of 5 ⁇ m or more in the vertical direction.
  • an angle formed between the upper end portion of the peripheral wall portion and the outer wall surface is 136 ° or less at an edge of the upper end portion.
  • a minimum width of the upper end portion of the peripheral wall portion is 0.5 mm or more and 10 mm or less in the horizontal direction.
  • the upper end portion of the peripheral wall portion is a flat surface and is flush with the upper surface of the container bottom portion.
  • the cell handling container includes a surrounding wall portion that surrounds the peripheral wall portion, and the upper end portion of the peripheral wall portion is a flat surface, and is located above the surrounding wall portion. It is preferable to be flush with the end face.
  • the distance from the edge of the upper end portion of the peripheral wall portion to the outer edge of the upper end surface of the surrounding wall portion is preferably 1 mm or less. is there.
  • At least the upper end portion of the peripheral wall portion is a hydrophilic surface.
  • the water contact angle of the hydrophilic surface is less than 80 °.
  • the cell handling container it is preferable that there are a plurality of the drop forming portions, and the upper end portions of the peripheral wall portions of the plurality of drop forming portions are flush with each other.
  • the cell handling container further includes a microwell that contains a cell, and a liquid container that surrounds the microwell and contains a liquid, It is preferable that the bottom surface of the portion is flush with the upper end portion of the peripheral wall portion of the drop forming portion.
  • the cell handling container further includes a microwell that contains a cell, and a liquid container that surrounds the microwell and that contains a liquid. It is preferable that the bottom surface is flush with the bottom surface of the recess of the drop forming portion.
  • a cell handling container includes at least one drop forming portion having an upper surface and a side surface extending downward and connected to the upper surface via a curved surface portion.
  • at least the upper surface is a liquid drop formation region, and the radius of curvature of the curved surface portion is less than 250 ⁇ m.
  • FIG. 3B is a sectional view taken along line BB in FIG. 3A. It is a fragmentary sectional view which shows the modification of a drop formation part and a groove part. It is a fragmentary sectional view which shows the modification of a drop formation part and a groove part.
  • FIG. 15A It is sectional drawing which follows the FF line
  • FIG. 20B is a cross-sectional view taken along line HH in FIG. 20A.
  • FIG. 20B is a cross-sectional view taken along line HH in FIG. 20A.
  • FIG. 21A It is a top view which shows the cell handling container of 10th Embodiment.
  • FIG. 22B is a cross-sectional view taken along line MM in FIG. 22A. It is a top view which shows the cell handling container of 11th Embodiment. It is sectional drawing which follows the NN line
  • FIG. 26 is a cross-sectional view taken along the line PP in FIG. 25. It is a bottom view which shows the container main body of a cell handling container.
  • FIG. 27A It is a top view which shows the cell handling container of 13th Embodiment.
  • FIG. 28B is a sectional view taken along line RR in FIG. 28A. It is an expanded sectional view of a drop formation part. It is an expanded sectional view showing a modification of a drop formation part.
  • 10 is a photograph showing the results of Example 8.
  • 14 is a photograph showing the results of Example 13.
  • It is a schematic fragmentary sectional view which shows the drop formed in the drop formation part of 1st Embodiment.
  • It is a model fragmentary sectional view which shows the relationship between angle (alpha) and water contact angle (theta). 2 is a photograph showing the results of some samples according to Example 1.
  • FIG. 1 shows the cell handling container of 13th Embodiment.
  • FIG. 28B is a sectional view taken along line RR in FIG. 28A.
  • It is an expanded sectional view of a drop formation part.
  • It is an expanded sectional view showing a modification of
  • FIG. 37B is a sectional view taken along line BB in FIG. 37A.
  • 2 is a photograph of a cell handling container used in Example 21.
  • FIG. 2 is a photograph of a cell handling container used in Example 21.
  • FIG. 2 is a photograph of a cell handling container used in Example 21.
  • FIG. 10 is a view showing the shape of a cell handling container used in Example 21. It is a schematic diagram which shows the drop shape change before the vibration in Example 21. It is a schematic diagram which shows the drop shape change after the vibration in Example 21.
  • FIG. 1A is a plan view showing the cell handling container of the first embodiment
  • FIG. 1B is a cross-sectional view taken along the line AA of FIG. 1A.
  • the cell handling container 1 of this embodiment is a container used for handling, for example, washing, culturing, and temporary storage of cells including fertilized eggs, egg cells, ES cells (embryonic stem cells) and iPS cells (artificial pluripotent stem cells). It is.
  • the egg cell refers to an unfertilized egg cell, and includes an immature oocyte and a mature oocyte.
  • the fertilized egg increases in number of cells from the 2 cell stage, the 4 cell stage, and the 8 cell stage by cleavage, and develops into a blastocyst through a morula.
  • Fertilized eggs include early embryos such as 2-cell embryos, 4-cell embryos and 8-cell embryos, morulas, blastocysts (including early blastocysts, expanded blastocysts and escaped blastocysts).
  • a blastocyst means an embryo composed of external cells with the potential to form the placenta and internal cell masses with the potential to form embryos.
  • ES cells refer to undifferentiated pluripotent or totipotent cells obtained from the inner cell mass of a blastocyst.
  • An iPS cell refers to a cell having a pluripotency similar to that of an ES cell by introducing several types of genes (transcription factors) into somatic cells (mainly fibroblasts).
  • the cell handling container 1 of the present embodiment is preferably suitable for culturing mammalian and avian cells, particularly mammalian cells.
  • Mammals refer to warm-blooded vertebrates, for example, primates such as humans and monkeys, rodents such as mice, rats and rabbits, pets such as dogs and cats, and domestic animals such as cattle, horses and pigs. Is mentioned.
  • the cell handling container 1 of this embodiment is suitable for culturing human and bovine fertilized eggs, and particularly suitable for culturing human fertilized eggs.
  • the cell handling container 1 includes a bottomed cylindrical container body 1a that opens upward, and a lid 1b that is detachably installed on the top of the container body 1a.
  • the lid 1b includes a disk-shaped top plate portion 51 that closes the opening of the container body 1a, and a cylindrical peripheral wall portion 52 that extends downward from the periphery of the top plate portion 51.
  • the container main body 1a is of a dish type having an inner diameter of 35 mm or 60 mm, for example, and has a substantially disk-shaped container bottom 10 having a top surface 10a and a bottom surface 10b arranged in parallel to each other, and a peripheral edge of the container bottom 10 A cylindrical side wall portion 11 erected from the bottom, and a cell accommodation area 12 provided at a central position of the container bottom portion 10.
  • the cell storage area 12 includes a plurality of microwells 13 for storing and culturing cells such as fertilized eggs, and a cylindrical culture solution storage unit (liquid storage unit) 14 that surrounds the microwells 13 and stores a culture solution.
  • a cylindrical culture solution storage unit (liquid storage unit) 14 that surrounds the microwells 13 and stores a culture solution.
  • Each of the microwells 13 has a U-shaped cross section, and is arranged close to each other at regular intervals in the front-rear and left-right directions.
  • the cross-section of the microwell 13 does not necessarily have a U-shape that opens upward.
  • the micro-well 13 may have a truncated cone shape with a cross-section opening upward, a U-shaped cross-section having an arc portion, or other shapes. good.
  • the bottom surface of the microwell 13 may be a flat surface or a tapered surface inclined from the outside toward the center.
  • a plurality (four in this embodiment) of drop forming portions 15 are provided on the upper surface 10a of the container bottom 10 so as to protrude upward from the upper surface 10a and form a liquid drop such as a culture solution.
  • These drop forming portions 15 each have a columnar shape, are formed integrally with the container bottom 10, and are arranged around the cell accommodation area 12 at equal intervals so as to surround the cell accommodation area 12.
  • the drop forming portion 15 has an upper surface 15a that is a drop forming region, and a side surface 15b that extends downward along the entire circumference of the upper surface 15a.
  • the upper surface 15a is a flat surface. Since the upper surface 15a is thus flat, for example, when a cell such as a fertilized egg placed on the upper surface 15a of the drop forming unit 15 is observed through a microscope, light reflection and scattering are less likely to occur, and the cell is clear. Can be obtained.
  • the flat surface here is, of course, a completely smooth horizontal surface, but may be substantially a horizontal surface, such as a difference of about several ⁇ m like a flat portion of a general injection molded product, This means that there may be a surface roughness of about 1% or less, which is within the standard tolerance in JIS B 0419-1991.
  • the side surface 15b is provided over the entire circumference along the circumferential direction of the upper surface 15a, and is disposed at a right angle to the upper surface 15a. Accordingly, the angle ⁇ formed by the upper surface 15a and the side surface 15b is 90 ° at the edge P1 of the upper surface.
  • the side surface 15b extends from the edge P1 of the upper surface 15a to a range of 5 ⁇ m or more.
  • the extending range in the vertical direction of the side surface 15b is relative to the upper surface 10a of the container bottom 10. This is the same as the height t of the upper surface 15a of the drop forming portion 15.
  • the height t of the upper surface 15a of the drop forming portion 15 with respect to the upper surface 10a of the container bottom 10 is 5 ⁇ m or more.
  • the lower limit of the height t, 5 ⁇ m is set based on the conditions under which the drop can be formed with the surface tension of the culture solution when the drop is formed using the fertilized egg culture solution.
  • the height t of the upper surface 15a of the drop forming portion 15 with respect to the upper surface 10a of the container bottom 10 is 10 ⁇ m or more, more preferably 50 ⁇ m or more, still more preferably 100 ⁇ m or more, and even more preferably 250 ⁇ m or more. It is.
  • the drop formed is easier to hold as the height t of the upper surface 15a of the drop forming part 15 with respect to the upper surface 10a of the container bottom 10 becomes higher, and sufficient retention is obtained when the drop reaches a certain level. is there.
  • the above-mentioned height t is 50 ⁇ m or more, a thick drop can be created even if the amount of liquid is increased, and when it is 100 ⁇ m or more, a thick drop can be stably formed even if the amount of liquid is increased. Further, when the thickness is 250 ⁇ m or more, a thick drop can be formed more stably even if the amount of liquid is increased.
  • the upper limit value of the height t is not particularly limited as long as the upper surface 15a of the drop forming portion 15 does not interfere with the top plate portion 51 of the lid 1b when the lid 1b is closed.
  • the width of the upper surface 15a (here, the diameter D of the upper surface 15a) is not less than 0.5 mm and not more than 10 mm.
  • the diameter D of the upper surface 15a is 3 mm or more and 6 mm or less.
  • a fertilized egg may grow to a diameter of about 250 ⁇ m in a blastocyst and to a diameter of about 500 ⁇ m including a zona pellucida.
  • the diameter D of the upper surface 15a is set to be not less than 0.5 mm, which is the maximum diameter of the fertilized egg, in order to facilitate the culture or washing operation.
  • a cell such as a fertilized egg may have a spherical shape or an elliptical shape depending on growth.
  • the diameter of the cell in this embodiment means the diameter when it is spherical, and the maximum width when it is not spherical.
  • the width of the upper surface 15a (that is, the diameter D of the upper surface 15a) may be set to 3.4 mm or more and 7.2 mm or less, and may be set to 3.8 mm or more and 5.4 mm or less. preferable.
  • the boundary point between the drop S and the upper surface 15a of the drop forming unit 15 here, an upper surface 15a described later.
  • h ′ exceeds r ′, the drop tends to collapse, so h ′ ⁇ r ′, V represents the amount of the culture solution [ ⁇ L], and ⁇ represents the circumference.
  • the culture solution volume V is preferably 10 ⁇ L or more and 100 ⁇ L or less, and more preferably 15 ⁇ L or more and 40 ⁇ L or less when cells such as fertilized eggs are cultured or washed.
  • the culture solution volume V is 10 ⁇ L.
  • the diameter D is in the range of 3.4 mm or more and 7.2 mm or less, and when the culture medium volume V is 15 ⁇ L or more and 40 ⁇ L or less, the diameter D is 3.8 mm or more and 5.4 mm or less. Become a range.
  • the diameter D is 2 mm or more.
  • the upper surface 15a of the drop formation part 15 is circular shape by planar view, polygonal shapes, such as a rectangle, an ellipse shape, a rounded square shape, etc. may be sufficient. However, considering the ease of holding the formed drop, the upper surface 15a is preferably circular.
  • the diameter D of the upper surface 15a exceeds 10 mm, the drop formed with respect to the size of the fertilized egg is too large, and it becomes difficult not only to find the fertilized egg during washing or culturing of the fertilized egg, but also limited.
  • the number of drop forming portions that can be installed in the space in the container is also reduced.
  • the diameter D of the upper surface 15a may be 10 mm or less. preferable.
  • drop forming portions 15 are provided, but the upper surfaces 15a of these drop forming portions 15 are all flush with each other.
  • the material of the cell handling container 1 is not particularly limited. Specifically, inorganic materials such as metal, glass, and silicon, plastics such as polystyrene resin, polyester resin, polyethylene resin, polypropylene resin, ABS resin, nylon, acrylic resin, fluororesin, polycarbonate resin, polyurethane resin, methyl resin Examples thereof include organic materials represented by pentene resin, phenol resin, melamine resin, epoxy resin, and vinyl chloride resin.
  • this cell handling container 1 can be manufactured by a method well-known to those skilled in the art. For example, when manufacturing using a plastic material, it can manufacture by injection molding.
  • the drop forming portion 15 protrudes upward from the upper surface 10a of the container bottom portion 10, and the upper surface 15a of the drop forming portion 15 is not directly connected to the upper surface 10a of the container bottom portion 10.
  • the upper surface 15a is a flat surface, and the height t of the container bottom 10 with respect to the upper surface 10a is 5 ⁇ m or more. For this reason, when the drop of the culture solution is formed on the upper surface 15a, the drop can be easily formed by the surface tension of the culture solution.
  • the upper surface 15a of the drop forming portion 15 is a hydrophilic surface, a thick drop S can be easily formed on the upper surface 15a as shown by the two-dot chain line in FIG. 1B. Then, cells C such as fertilized eggs can be arranged in the drop S.
  • the tip of the instrument such as a pipette is at the edge or side wall of the well as compared with the case where a well for forming a drop is provided as in the prior art. It is possible to improve workability without worrying about contact and damage.
  • the surface of the microwell 13 and the surface of the cell storage area 12 are further hydrophilic in order to easily remove air bubbles in the microwell 13 and improve workability when the culture solution is added. It may be the surface.
  • hydrophilic materials may be used for the entire cell-handling container 1 including the cell storage area 12 and the drop forming portion 15, or these surfaces may be subjected to a hydrophilic treatment. May be. Although the hydrophilic treatment can be patterned using a mask, the entire surface is preferable from the viewpoint of production cost and quality control.
  • the hydrophilization treatment can be performed by a method usually used in this technical field, and is not particularly limited. For example, plasma treatment, coating treatment, UV irradiation treatment, EB irradiation treatment, graft polymerization treatment of hydrophilic polymer on the surface, etc. are mentioned, but the shape of the treatment object has a three-dimensional fine and complicated structure. However, plasma treatment is preferable from the viewpoint of uniformly treating the whole.
  • the hydrophilic treatment is preferably performed after the cell handling container 1 is formed. In addition, it is preferable that water contact angle (theta) of a hydrophilic surface is less than 80 degrees (refer FIG. 1B).
  • the water contact angle ⁇ is less than 80 °, attempting to form a drop on a flat surface will cause the drop to collapse, and even if a drop can be formed, it is difficult to operate a pipette or other instrument.
  • cells such as eggs cannot be washed or cultured.
  • the water contact angle ⁇ is 60 ° or less, the above-described problem becomes more significant.
  • the thickness of the drop formed is used from the viewpoint of ensuring good operability of instruments such as pipettes when the amount of the culture solution is 15 ⁇ L or more and 30 ⁇ L or less. 1 mm or more is preferable, and 1.5 mm or more is more preferable.
  • the drop thickness is 1.5 mm or less, making it difficult to operate the instrument.
  • the thickness of the drop is 1.0 mm or less, which makes it very difficult to operate the instrument.
  • a thick drop can be easily formed even if the water contact angle ⁇ is less than 80 °, and a thick drop can be formed even if it is 60 ° or less. It can be easily formed, and even when the angle is 10 ° or less, a thick drop can be easily formed.
  • the side surface 15c of the drop forming portion 15 is an inclined surface that inclines outward as it goes downward in a sectional view.
  • the cross section of the drop forming portion 15 has a truncated cone shape.
  • the angle ⁇ formed by the upper surface 15a and the side surface 15c is an obtuse angle at the edge P1 of the upper surface 15a.
  • the angle ⁇ is preferably 136 ° or less.
  • a smaller angle ⁇ facilitates the formation of thicker drops, and the formed drops are easier to hold, so it is more preferably 100 ° or less, more preferably 95 ° or less, more preferably 93 ° or less, More preferably, it is 92 ° or less, and still more preferably 91 ° or less.
  • the angle ⁇ here includes a case where the tip is not sharp due to a manufacturing problem.
  • the water contact angle of the drop forming portion at the edge P1 is ⁇ ′ ⁇ ⁇ 90 ° (where ⁇ ⁇ 90 °)
  • the water contact angle of the drop forming portion at the edge P1 If ⁇ is in the range of ⁇ ⁇ 90 °, a drop can be formed.
  • ⁇ 120 °
  • the water contact angle ⁇ ′ of the entire inner surface of the container excluding the edge P1 is ⁇ ′ ⁇ 30 °
  • a drop is formed if ⁇ ⁇ 90 °. Is possible.
  • the side surface 15d is a stepped surface having a vertical portion and an inclined portion in order from above in a sectional view.
  • the vertical portion is a portion extending at a right angle to the upper surface 15a
  • the inclined portion is a portion inclined outward as it goes downward.
  • the angle ⁇ formed by the upper surface 15a and the vertical portion of the side surface 15d is 90 °.
  • the corners between the side surface 15e of the drop forming portion 15 and the upper surface 10a of the container bottom portion 10 are processed so as to have a curved portion in a cross-sectional view. Therefore, the side surface 15e has a vertical portion and a curved portion in order from above. At this time, at the edge P1 of the upper surface 15a, the angle ⁇ formed by the upper surface 15a and the vertical portion of the side surface 15e is 90 °.
  • FIG. 3A is a plan view showing the cell handling container of the second embodiment
  • FIG. 3B is a cross-sectional view taken along line BB of FIG. 3A.
  • the difference between the cell handling container 2 of the present embodiment and the first embodiment is that a groove 17 is provided between the drop forming portion 16 and the container bottom 10, and the top surface 16 a of the drop forming portion 16 is the same as that of the container bottom 10. It is flush with the upper surface 10a. Since other configurations are the same as those of the first embodiment, a duplicate description is omitted.
  • the container bottom 10 has a plurality of drop forming portions 16 (here, a plurality of drop forming portions 16 each having a top surface 16a that is a drop forming region and a side surface 16b that is provided continuously along the circumferential direction of the top surface 16a and extends downward). Then, four) are provided.
  • the drop forming part 16 is formed in a columnar shape, similar to the drop forming part 15 of the first embodiment.
  • the upper surface 16a of the drop forming portion 16 is a flat surface and is flush with the upper surface 10a of the container bottom 10. Further, as shown in FIG. 3B, the upper surface 16 a of the drop forming unit 16 is flush with the bottom surface 14 a of the culture solution storage unit 14 in the cell storage area 12.
  • the side surface 16b is provided over the entire circumference of the upper surface 16a, is disposed at a right angle to the upper surface 16a, and extends downward from the edge P1 of the upper surface 16a to a range of 5 ⁇ m or more.
  • the height t of the upper surface 16a of the drop forming portion 16 with respect to the bottom surface of the groove portion 17 (described later) is 5 ⁇ m or more.
  • the angle ⁇ formed by the upper surface 16a and the side surface 16b is 90 ° at the edge P1 of the upper surface 16a.
  • the height t of the upper surface 16a of the drop forming portion 16 with respect to the bottom surface of the groove portion 17 is 10 ⁇ m or more, more preferably 50 ⁇ m or more, still more preferably 100 ⁇ m or more, and even more preferably 250 ⁇ m or more.
  • these lower limit values are easier to hold the formed drop and increase to a certain level. This is because sufficient retention can be obtained.
  • the above-mentioned height t is 50 ⁇ m or more, a thick drop can be created even if the amount of liquid is increased, and when it is 100 ⁇ m or more, a thick drop can be stably formed even if the amount of liquid is increased. Further, when the thickness is 250 ⁇ m or more, a thick drop can be formed more stably even if the amount of liquid is increased.
  • the thickness of the bottom surface of a general cell culture container is as large as 1 mm.
  • the height t (groove depth) is 1 mm, the bottom surface of the container penetrates.
  • the height t (groove depth) is preferably less than 1 mm, more preferably 500 ⁇ m or less, more preferably 350 ⁇ m or less, even more preferably 250 ⁇ m or less, even more preferably 100 ⁇ m or less from the viewpoint of ease of production by injection molding. Is even more preferable.
  • the height t (depth of the groove portion) is 500 ⁇ m or less, the possibility that the bottom surface of the container is penetrated or broken due to physical damage is extremely low.
  • annular groove part 17 surrounding the drop forming part 16 is provided for each drop forming part 16.
  • the groove portion 17 is formed in a U shape whose cross section opens upward.
  • the side surface 16 b of the drop forming portion 16 constitutes one side surface of the groove portion 17.
  • the distance d from the edge P1 of the upper surface 16a of the drop forming part 16 to the edge P2 of the upper surface 10a of the adjacent container bottom 10 via the groove 17 is 0.01 mm or more, preferably 0.05 mm or more. More preferably, it is 0.1 mm or more, More preferably, it is 0.4 mm or more. This is because as the distance d is increased, the formed drop is more easily held, and when the distance d is increased to a certain level, sufficient holding can be obtained. On the other hand, if the distance d is too large, the number of drop forming portions that can be installed in a limited space is reduced. In the present embodiment, the distance d is 0.5 mm.
  • the upper surface 16a of the drop forming portion 16 is isolated from the upper surface 10a of the container bottom 10 by the groove portion 17, the upper surface 16a is a flat surface, and the side surface 16b is 5 ⁇ m or more from the upper surface 16a. It extends downward to the range. Therefore, similarly to the first embodiment described above, even when the upper surface 16a is a hydrophilic surface, the drop S of the culture solution can be easily formed on the upper surface 16a. Furthermore, since there is no fear of the tip of the instrument coming into contact with the edge or side wall of the well and being damaged as in the prior art, workability can be improved.
  • the upper surface 16a of the drop forming portion 16 is flush with the upper surface 10a of the container bottom portion 10, for example, when the fertilized egg is washed a plurality of times using an instrument such as a pipette, the instrument can be slid and moved. Therefore, workability can be further improved.
  • the bottom surface of the microwell 13 may be flush with the top surface 16a of the drop forming portion 16.
  • the groove portion 17 according to the present embodiment has an annular shape surrounding the drop forming portion 16.
  • the annular portion 17 has an annular shape. May be cut off and may be substantially C-shaped when viewed from above.
  • the side surface 16c of the drop forming portion 16 is an inclined surface that inclines outward as it goes downward in a sectional view.
  • the cross section of the drop forming portion 16 has a truncated cone shape.
  • the angle ⁇ formed by the upper surface 16a and the side surface 16c is an obtuse angle, but is preferably 136 ° or less for the reason described above.
  • a smaller angle ⁇ facilitates the formation of thicker drops, and the formed drops are easier to hold, so it is more preferably 100 ° or less, more preferably 95 ° or less, more preferably 93 ° or less, More preferably, it is 92 ° or less, and still more preferably 91 ° or less.
  • the side surface 16d of the drop forming portion 16 is an inclined surface that is inclined outward as it goes downward.
  • the cross section of the drop forming portion 16 has a truncated cone shape.
  • the angle ⁇ formed by the upper surface 16a and the side surface 16d is an obtuse angle, but is preferably 136 ° or less, more preferably 100 ° or less, still more preferably 95 ° or less, and 93 ° or less. More preferably, it is more preferably 92 ° or less, and still more preferably 91 ° or less.
  • the cross section of the groove 19 is V-shaped.
  • the side surface 16e is a stepped surface having a vertical portion and an inclined portion in order from above in a cross-sectional view.
  • the vertical portion is a portion that extends at a right angle to the upper surface 16a
  • the inclined portion is a portion that is inclined outward as it goes downward.
  • the angle ⁇ formed by the upper surface 16a and the vertical portion of the side surface 16e is 90 °.
  • the cross section of the groove part 20 is the shape combined by the upper side rectangular shape and the lower side inverted triangle.
  • the corners of the side surface 16f of the drop forming portion 16 and the container bottom portion 10 are processed so as to have a curved portion in a cross-sectional view.
  • the side surface 16f has a vertical portion and a curved portion in order from above.
  • the angle ⁇ formed by the upper surface 16a and the vertical portion of the side surface 16f is 90 °.
  • the cross section of the groove part 21 is U-shaped.
  • the side surface 16g of the drop forming portion 16 is an inclined surface that is inclined inward as it goes downward in a sectional view.
  • the cross section of the drop forming portion 16 has an inverted truncated cone shape.
  • the angle ⁇ formed by the upper surface 16a and the side surface 16g becomes an acute angle.
  • the cross section of the groove 22 has a truncated cone shape that opens upward.
  • the side surface 16h of the drop forming portion 16 is a stepped surface having an inclined portion and a vertical portion in order from above in a cross-sectional view.
  • the inclined portion is a portion inclined inward as going downward, and the vertical portion is a portion extending at a right angle to the upper surface 16a.
  • the angle ⁇ formed by the upper surface 16a and the inclined portion of the side surface 16h is an acute angle.
  • the cross section of the groove part 23 is the shape combined by the upper truncated cone shape and the lower rectangular shape.
  • the upper surface 16a of the drop forming portion 16 is flush with the upper surface 10a of the container bottom portion 10, but the upper surface 16a of the drop forming portion 16 is higher than the upper surface 10a of the container bottom portion 10 as shown in FIG. 6A. It may be arranged. In this case, a part of the side surface 16 i of the drop forming portion 16 (that is, a portion of the side surface 16 i below the upper surface 10 a of the container bottom 10) constitutes one side surface of the groove portion 24.
  • the upper surface 16a of the drop forming portion 16 may be disposed lower than the upper surface 10a of the container bottom portion 10.
  • the side surface 16j of the drop forming portion 16 constitutes one side surface of the groove portion 25.
  • FIG. 7A is a partial plan view showing the cell handling container of the third embodiment
  • FIGS. 7B and 7C are sectional views taken along the line CC of FIG. 7A.
  • the difference between the cell handling container 3 of this embodiment and the second embodiment is that it further includes a surrounding wall portion 26 surrounding the drop forming portion 16 and a groove portion 27 is provided between the container bottom portion 10 and the surrounding wall portion 26. It is. Since other configurations are the same as those of the second embodiment, a duplicate description is omitted.
  • annular surrounding wall portion 26 is provided on the outer periphery of the drop forming portion 16 so as to surround the drop forming portion 16.
  • the surrounding wall portion 26 is disposed at a predetermined distance from the side surface 16b of the drop forming portion 16.
  • the upper end surface 26 a of the surrounding wall portion 26 is flush with the upper surface 16 a of the drop forming portion 16 and the upper surface 10 a of the container bottom portion 10.
  • the distance c from the edge P1 of the upper surface 16a of the drop forming portion 16 to the outer edge P3 of the upper end surface 26a of the surrounding wall portion 26 is 1 mm or less, preferably 0.8 mm or less, preferably 0.5 mm or less. It is more preferable that
  • the groove portion 27 of the present embodiment is formed in a U shape whose cross section opens upward, like the groove portion 17 of the second embodiment described above.
  • One of the side surfaces of the groove portion 27 is constituted by the side surface of the surrounding wall portion 26.
  • the modification of the groove of the second embodiment is also applied.
  • the distance d2 to the edge P2 of the upper surface 10a of the adjacent container bottom 10 is 0.01 mm or more, preferably 0.05 mm or more, more preferably 0.1 mm or more, and still more preferably 0.4 mm or more. This is because the drop formed is easier to hold as the distances d1 and d2 are increased, and sufficient holding is obtained when the drop is increased to a certain level. On the other hand, if the distances d1 and d2 are too large, the number of drop forming portions that can be installed in a limited space is reduced.
  • the upper surface 16a of the drop forming part 16 is isolated from the surroundings by the space between the surrounding wall part 26, the upper surface 16a is similar to the above-described second embodiment. Even if it is a hydrophilic surface, the culture medium drop S can be easily formed on the upper surface 16a, and the tip of the instrument does not come into contact with the edge or side wall of the well as in the conventional case, Workability can be improved.
  • a surrounding wall portion 26 surrounding the drop forming portion 16 is provided, a distance c from the edge P1 of the upper surface 16a to the outer edge P3 of the upper end surface 26a of the surrounding wall portion 26 is 1 mm or less, and the surrounding wall portion 26 and the container bottom 10 Since the groove 27 is provided between the drop S and the drop S formed on the upper surface 16a of the drop forming part 16 spreads to the surroundings due to vibration or the like, the drop S extends to the surrounding wall 26 and It is possible to stay at the outer edge P3 (see FIG. 7C). As a result, the collapse of the drop S can be prevented. In this case, the thickness of the drop S is reduced by spreading to the outer edge P3, but it does not affect the culture and washing of the fertilized egg.
  • the upper end surface 26a of the surrounding wall portion 26 and the upper surface 16a of the drop forming portion 16 are flush with each other, but may not be flush with each other.
  • FIG. 8A is a partial plan view showing the cell handling container of the fourth embodiment
  • FIGS. 8B and 8C are cross-sectional views taken along the line DD of FIG. 8A.
  • the difference between the cell handling container 4 of this embodiment and the second embodiment is that the side surface 16k of the drop forming portion 16 is formed by two or more step surfaces. Since other configurations are the same as those of the second embodiment, a duplicate description is omitted.
  • the side surface 16k extends downward along the entire circumference of the upper surface 16a and is formed in a step shape (here, three steps) so as to expand from the inside to the outside.
  • the side surface 16k includes, in order from the top, a first portion 16k1 disposed at a right angle to the upper surface 16a, a second portion 16k2 having a horizontal portion and a vertical portion connected to the first portion 16k1, and a second portion 16k2. And a third portion 16k3 having a horizontal portion and a vertical portion.
  • the distance t1 from the edge P1 of the upper surface 16a of the side surface 16k to the horizontal portion of the second portion 16k2 (that is, the height of the first portion 16k1) t1, the horizontal portion of the second portion 16k2 to the horizontal portion of the third portion 16k3 Distance to the portion (ie, the height of the second portion 16k2) t2, and distance from the horizontal portion of the third portion 16k3 to the bottom surface of the groove portion 28 (described later) (ie, the height of the third portion 16k3) t3.
  • the above-described heights t1, t2, and t3 are each 10 ⁇ m or more, more preferably 50 ⁇ m or more, still more preferably 100 ⁇ m or more, and further preferably 250 ⁇ m or more.
  • the angle ⁇ formed by the upper surface 16a and the first portion 16k1 of the side surface 16k at the edge P1 of the upper surface 16a is 90 °. is there.
  • annular groove portion 28 surrounding the drop forming portion 16 is provided between the drop forming portion 16 and the container bottom 10.
  • the groove portion 28 opens upward, and one of the side surfaces of the groove portion 28 is formed in a step shape by the side surface 16k of the drop forming portion 16, and the other is a vertical surface.
  • the distance y1 from the edge P1 of the upper surface 16a of the drop forming portion 16 to the vertical portion of the second portion 16k2 (that is, the width of the second portion 16k2), and the vertical portion to the third portion of the second portion 16k2 is 1 mm or less, preferably 0.8 mm or less, and more preferably 0.5 mm or less.
  • the distance X from the vertical portion of the third portion 16k3 to the edge P2 of the upper surface 10a of the adjacent container bottom portion 10 via the groove portion 28 is 0.01 mm or more, preferably 0.05 mm or more. Yes, more preferably 0.1 mm or more, and still more preferably 0.4 mm or more. This is because as the distance X is increased, the formed drop is easily held, and when the distance X is increased to a certain level, sufficient holding is obtained. On the other hand, if the distance X is too large, the number of drop forming portions that can be installed in a limited space is reduced, and therefore it is preferably 4 mm or less.
  • the cell handling container 4 configured as described above can obtain the same effects as those of the third embodiment described above, and further has the following effects. That is, since the side surface 16k of the drop forming portion 16 is formed in a stepped shape, if the drop S formed on the upper surface 16a of the drop forming portion 16 spreads around due to vibration or the like, the drop S becomes the second side of the side surface 16k. It is possible to extend to the edge of the horizontal portion in the portion 16k2 or the edge of the horizontal portion in the third portion 16k3 and to stay there (see FIG. 8C). As a result, the collapse of the drop S can be prevented. In this case, the thickness of the drop S is reduced by the spread of the drop S, but it does not affect the culture and washing of the fertilized egg.
  • the side surface 16k of the drop forming portion 16 is a three-level step surface, but may be two or four or more levels.
  • FIG. 9A is a plan view showing the cell handling container of the fifth embodiment
  • FIG. 9B is a cross-sectional view taken along line EE of FIG. 9A.
  • the difference between the cell handling container 5 of the present embodiment and the first embodiment is the shape of the drop forming unit 30. Since other configurations are the same as those of the first embodiment, a duplicate description is omitted.
  • a plurality (four in this embodiment) of drop forming portions 30 for forming a liquid drop of a culture solution or the like are provided on the upper surface 10a of the container bottom 10.
  • These drop forming portions 30 have a cylindrical shape, are erected from the upper surface 10 a of the container bottom portion 10, and are arranged around the cell storage area 12 at equal intervals.
  • the drop forming portion 30 is formed by a concave portion 31 that opens upward and a peripheral wall portion 32 that forms the concave portion 31.
  • the recess 31 has a rectangular internal space in cross-sectional view, and its bottom surface 31a is formed flat.
  • the peripheral wall portion 32 has an inner wall surface 32a that forms the internal space of the recess 31, an outer wall surface 32b that is located outside the inner wall surface 32a, and an upper end surface 32c that is located above.
  • the upper end surface 32c corresponds to the “upper end portion of the peripheral wall portion” recited in the claims, and is a flat surface.
  • the upper end surface 32 c is a liquid drop formation region together with the recess 31. In the present embodiment, the upper end surfaces 32c of the peripheral wall portions 32 of the four drop forming portions 30 are all flush.
  • the angle ⁇ formed by the upper end surface 32c and the outer wall surface 32b is 90 ° at the edge P1 of the upper end surface 32c.
  • the outer wall surface 32b extends from the edge P1 of the upper end surface 32c to a range of 5 ⁇ m or more in the vertical direction.
  • the distance from the upper end surface 32c of the peripheral wall portion 32 to the upper surface 10a of the container bottom portion 10 is h1
  • the distance from the upper end surface 32c of the peripheral wall portion 32 to the bottom surface 31a (the deepest portion) of the recess 31 is h2
  • the sum of h1 and h2 is less than 0.2 mm (that is, h1 + h2 ⁇ 0.2 mm).
  • the sum of h1 and h2 is 0.1 mm or less, and more preferably the sum of h1 and h2 is 0.02 mm or less.
  • symbol f shown to FIG. 9B is the thickness (namely, distance from the inner wall surface of an outer peripheral wall part to an outer wall surface) in the horizontal direction.
  • the upper end surface 32c of the peripheral wall portion 32 and the concave portion 31 are used as a liquid drop forming region, and the outer wall surface 32b is in the range of 5 ⁇ m or more from the edge P1 of the upper end surface 32c in the vertical direction. It is extended to. Therefore, similarly to the first embodiment, even when the upper end surface 32 c is a hydrophilic surface, the culture solution drop S can be easily formed by the upper end surface 32 c and the recess 31.
  • the inner wall surface 32 d of the peripheral wall portion 32 is an inclined surface that is inclined inward as it goes downward in a cross-sectional view.
  • the cross section of the recess 31 has an inverted truncated cone shape.
  • the angle ⁇ formed by the upper end surface 32c and the outer wall surface 32b is 90 °.
  • the inner wall surface 32e of the peripheral wall portion 32 is an inclined surface that inclines inward and the concave portion 31 is V-shaped (that is, an inverted triangle that opens upward). It has become.
  • the angle ⁇ formed by the upper end surface 32c and the outer wall surface 32b is 90 °.
  • the inner wall surface 32f of the peripheral wall portion 32 is a stepped surface having a vertical portion and an inclined portion in order from above in a cross-sectional view.
  • the vertical portion is a portion extending at a right angle to the upper end surface 32c
  • the inclined portion is a portion inclined inward as going downward.
  • the recess 31 has a shape formed by combining the upper rectangular shape and the lower inverted triangle in a cross-sectional view.
  • the angle ⁇ formed by the upper end surface 32c and the outer wall surface 32b is 90 °.
  • the inner wall surface 32g of the peripheral wall portion 32 is formed so as to have a vertical portion and a curved portion in order from above in a sectional view.
  • the vertical portion is a portion that extends at a right angle to the upper end surface 32c
  • the curved portion is a portion that curves so as to protrude downward.
  • the angle ⁇ formed by the upper end surface 32c and the outer wall surface 32b is 90 °.
  • the inner wall surface 32 e of the peripheral wall portion 32 becomes an inclined surface that is inclined inward as it goes downward, and the concave portion 31 is V-shaped (that is, an inverted triangle that opens upward). It has become.
  • the outer wall surface 32b and the inner wall surface 32e intersect to form a ridge line portion 32h.
  • This ridge line portion 32h corresponds to an “upper end portion of the peripheral wall portion” recited in the claims.
  • the angle ⁇ formed by the outer wall surface 32b and the inner wall surface 32e is an acute angle. A sharper angle ⁇ can form a stable drop with the surface tension of the culture solution.
  • the inner wall surface 32 e of the peripheral wall portion 32 becomes an inclined surface that is inclined inward as it goes down, and the concave portion 31 has a V shape (an inverted triangle that opens upward). Yes.
  • the outer wall surface 32i has an inclined portion that inclines outward from the top and a vertical portion that extends in the vertical direction.
  • the inclined portion of the outer wall surface 32i intersects with the inner wall surface 32e to form a ridge line portion 32h.
  • This ridge line portion 32h corresponds to an “upper end portion of the peripheral wall portion” recited in the claims.
  • the angle ⁇ formed by the inclined portion of the outer wall surface 32i and the inner wall surface 32e is an acute angle.
  • the inner wall surface 32 e of the peripheral wall portion 32 becomes an inclined surface that is inclined inward as it goes downward, and the concave portion 31 is V-shaped (an inverted triangle that opens upward).
  • the outer wall surface 32j is an inclined surface that is inclined outward as it goes downward.
  • the outer wall surface 32j and the inner wall surface 32e intersect to form a ridge line portion 32h.
  • This ridge line portion 32h corresponds to an “upper end portion of the peripheral wall portion” recited in the claims.
  • the angle ⁇ formed by the outer wall surface 32j and the inner wall surface 32e is an acute angle.
  • the outer wall surface 32k and the inner wall surface 32l of the peripheral wall portion 32 have an inclined portion that inclines outward as it goes downward, and a vertical portion that extends in the vertical direction. Are formed respectively.
  • the upper end portion of the peripheral wall portion 32 is formed as a pointed portion when the inclined portion of the outer wall surface 32k and the inclined portion of the inner wall surface 32l intersect in an inverted V shape. For this reason, a ridge line portion 32 h is formed at the upper end of the peripheral wall portion 32. This ridge line portion 32h corresponds to an “upper end portion of the peripheral wall portion” recited in the claims.
  • the angle ⁇ formed by the outer wall surface 32k and the inner wall surface 32l is an acute angle.
  • the symbol g shown in FIG. 13B is the distance of the sharp portion in the vertical direction (that is, the distance from the ridgeline portion 32h to the boundary portion between the inclined portion and the vertical portion in the outer wall surface 32k (or the inner wall surface 32l)).
  • the bottom surface of the recess 31 is flush with the upper surface 10a of the container bottom 10.
  • the bottom surface 31a of the recess 31 is disposed higher than the upper surface 10a of the container bottom 10 as shown in FIG. 14A.
  • the bottom surface 31 a of the recess 31 may be disposed lower than the top surface 10 a of the container bottom 10.
  • FIG. 15A is a plan view showing a cell handling container of the sixth embodiment
  • FIG. 15B is a cross-sectional view taken along the line FF of FIG. 15A.
  • the difference between the cell handling container 6 of the present embodiment and the first embodiment is the shape of the drop forming unit 40. Since other configurations are the same as those of the first embodiment, a duplicate description is omitted.
  • a plurality (four in this embodiment) of drop forming portions 40 for forming a liquid drop such as a culture solution are provided on the upper surface 10a of the container bottom 10.
  • Each of these drop forming portions 40 has a cylindrical shape, is formed so as to be embedded in the container bottom 10, and is arranged around the cell storage area 12 at equal intervals.
  • the drop forming portion 40 is formed by a concave portion 41 that opens upward and a peripheral wall portion 42 that forms the concave portion 41.
  • a groove 43 that surrounds the peripheral wall 42 is provided for each drop forming portion 40 between the container bottom 10 and the peripheral wall 42.
  • the recess 41 has a rectangular internal space in cross-sectional view, and its bottom surface 41a is formed flat.
  • the peripheral wall portion 42 has an inner wall surface 42a that forms the inner space of the recess 41, an outer wall surface 42b that is positioned outside the inner wall surface 42a, and an upper end surface 42c that is positioned above.
  • the upper end surface 42c corresponds to an “upper end portion of the peripheral wall portion” described in the claims, is a flat surface, and is flush with the upper surface 10a of the container bottom portion 10.
  • the upper end surface 42 c is a liquid drop formation region together with the recess 41.
  • the upper end surfaces 42c of the peripheral wall portions 42 of the four drop forming portions 40 are all flush with each other and flush with the bottom surface 14a of the culture solution accommodating portion 14 in the cell accommodating area 12.
  • the outer wall surface 42b is disposed at a right angle to the upper end surface 42c, the angle ⁇ between the upper end surface 42c and the outer wall surface 42b is 90 ° at the edge P1 of the upper end surface 42c.
  • the outer wall surface 42b extends from the edge P1 of the upper end surface 42c to a range of 5 ⁇ m or more in the vertical direction, and constitutes one side surface of the groove 43.
  • the distance d from the edge P1 of the upper end surface 42c of the drop forming portion 40 to the edge P2 of the upper surface 10a of the adjacent container bottom 10 through the groove 43 is 0.01 mm or more, preferably 0.05 mm. It is above, More preferably, it is 0.1 mm or more, More preferably, it is 0.4 mm or more. This is because as the distance d is increased, the formed drop is more easily held, and when the distance d is increased to a certain level, sufficient holding can be obtained. On the other hand, if the distance d is too large, the number of drop forming portions that can be installed in a limited space is reduced. In the present embodiment, the distance d is 0.5 mm.
  • the distance from the upper end surface 42c of the peripheral wall portion 42 to the upper surface 10a of the container bottom portion 10 is h3 (see FIG. 18), and the upper end surface 42c of the peripheral wall portion 42 is the bottom surface 41a (the deepest).
  • the sum of h3 and h4 is 1 mm or less (that is, h3 + h4 ⁇ 1 mm).
  • the sum of h3 and h4 is 0.5 mm or less, more preferably the sum of h3 and h4 is 0.2 mm or less, more preferably the sum of h3 and h4 is 0.1 mm or less, More preferably, the sum of h3 and h4 is 0.02 mm or less.
  • h3 0.
  • the upper end surface 42c and the concave portion 41 of the peripheral wall portion 42 are liquid drop forming regions, and the outer wall surface 42b is in the range of 5 ⁇ m or more from the edge P1 of the upper end surface 42c in the vertical direction. It is extended to. Therefore, similarly to the first embodiment described above, even when the upper end surface 42c is a hydrophilic surface, the culture solution drop S can be easily formed by the upper end surface 42c and the recess 41.
  • the bottom surface of the microwell 13 may be flush with the bottom surface 41a of the recess 41 of the drop forming unit 40. In this way, for example, when a cell C such as a fertilized egg is moved between the microwell 13 and the drop forming unit 40, or when the cell C is placed in both places, the observation is performed at the same focus position in the microscopic observation. The workability can be further improved.
  • the inner wall surface 42d of the peripheral wall portion 42 is an inclined surface that inclines inward as viewed in cross section.
  • the bottom surface 41b of the recess 41 is a flat surface.
  • the cross section of the recess 41 has an inverted truncated cone shape that opens upward.
  • the angle ⁇ formed by the upper end surface 42c and the outer wall surface 42b is 90 °.
  • the inner wall surface 42e of the peripheral wall portion 42 is an inclined surface that inclines inward as viewed in cross section, and the concave portion 41 has a V shape (an inverted triangle that opens upward). It has become. For this reason, the bottom 41c of the recess 41 is the apex of an inverted triangle. At this time, at the edge P1 of the upper end surface 42c, the angle ⁇ formed by the upper end surface 42c and the outer wall surface 42b is 90 °.
  • the inner wall surface 42f of the peripheral wall portion 42 is a stepped surface having a vertical portion and an inclined portion in order from the top in a sectional view.
  • the vertical portion is a portion extending at a right angle to the upper end surface 42c
  • the inclined portion is a portion inclined inward as going downward.
  • the concave portion 41 has a shape formed by combining the upper rectangular shape and the lower inverted triangle in a sectional view. For this reason, the bottom 41d of the recess 41 is the vertex of an inverted triangle.
  • the angle ⁇ formed by the upper end surface 42c and the outer wall surface 42b is 90 °.
  • the inner wall surface 42g of the peripheral wall portion 42 is a curved surface that protrudes downward in a sectional view.
  • the bottom 41e of the recess 31 is a convex curved surface that protrudes downward.
  • the angle ⁇ formed by the upper end surface 42c and the outer wall surface 42b is 90 °.
  • the upper end surface 42 c of the peripheral wall portion 42 is flush with the upper surface 10 a of the container bottom portion 10.
  • the upper end surface 42 c of the peripheral wall portion 42 is the upper surface of the container bottom portion 10.
  • the upper end surface 42c of the peripheral wall portion 42 may be disposed lower than the upper surface 10a of the container bottom portion 10 as shown in FIG. 18B.
  • FIG. 19A is a partial plan view showing a cell handling container of a seventh embodiment
  • FIGS. 19B and 19C are cross-sectional views taken along the line GG of FIG. 19A.
  • the difference between the cell handling container 7 of the present embodiment and the sixth embodiment is that an enclosure wall 44 that surrounds the drop forming section 40 is further provided, and a groove 45 is provided between the container bottom 10 and the enclosure wall 44. It is.
  • the other configuration is the same as that of the sixth embodiment, and a duplicate description is omitted.
  • annular surrounding wall portion 44 is provided on the outer periphery of the peripheral wall portion 42 of the drop forming portion 40 so as to surround the peripheral wall portion 42.
  • the surrounding wall portion 44 is disposed away from the outer wall surface 42b of the peripheral wall portion 42 by a predetermined distance.
  • the upper end surface 44 a of the surrounding wall portion 44 is flush with the upper end surface 42 c of the peripheral wall portion 42 and the upper surface 10 a of the container bottom portion 10.
  • the distance c from the edge P1 of the upper end surface 42c of the peripheral wall portion 42 to the outer edge P3 of the upper end surface 44a of the surrounding wall portion 44 is 1 mm or less, preferably 0.8 mm or less, preferably 0.5 mm or less. It is more preferable that
  • a groove 45 surrounding the surrounding wall 44 is provided between the surrounding wall 44 and the container bottom 10.
  • the groove portion 45 is formed in a U shape whose cross section opens upward.
  • One side surface of the groove portion 45 is constituted by the side surface of the surrounding wall portion 44.
  • the distance d2 to the edge P2 of the upper surface 10a of the adjacent container bottom 10 is 0.01 mm or more, preferably 0.05 mm or more, more preferably 0.1 mm or more, and still more preferably 0.4 mm or more. This is because the drop is easily held when the distances d1 and d2 described above become large to some extent. On the other hand, if the distances d1 and d2 are too large, the number of drop forming portions that can be installed in a limited space is reduced.
  • the cell handling container 7 configured as described above can obtain the same effects as those of the above-described sixth embodiment. Further, since the surrounding wall portion 44 surrounding the drop forming portion 40 is provided and the groove portion 45 is provided between the surrounding wall portion 44 and the container bottom portion 10, the drop S formed by the concave portion 41 and the upper end surface 42c of the peripheral wall portion 42 is assumed. Can spread to the surrounding wall portion 44 and stay on the outer edge P3 of the surrounding wall portion 44 (see FIG. 19C). As a result, the collapse of the drop S can be prevented. In this case, the thickness of the drop S is reduced by the spread of the drop S, but it does not affect the culture and washing of the cells C such as fertilized eggs.
  • FIGS. 20B and 20C are cross-sectional views taken along the line HH of FIG. 20A.
  • the difference between the cell handling container 8 of the present embodiment and the sixth embodiment is that the outer wall surface 42h of the peripheral wall portion 42 is formed by two or more step surfaces.
  • the other configuration is the same as that of the sixth embodiment, and a duplicate description is omitted.
  • the outer wall surface 42h of the peripheral wall portion 42 extends downward along the entire circumference of the upper end surface 42c and is formed in a stepped shape (here, three steps) so as to expand from the inside to the outside. Yes.
  • the outer wall surface 42h includes, in order from the top, a first portion 42h1 disposed at a right angle to the upper end surface 42c, a second portion 42h2 connected to the first portion 42h1, and having a horizontal portion and a vertical portion, and a second portion 42h1.
  • the third portion 42h3 is connected to the portion 42h2 and has a horizontal portion and a vertical portion.
  • the distance from the edge P1 of the upper end surface 42c of the peripheral wall portion 42 to the horizontal portion of the second portion 42h2 of the outer wall surface 42h (that is, the height of the first portion 42h1) t1, from the horizontal portion of the second portion 42h2
  • the distance to the horizontal portion of the third portion 42h3 (ie, the height of the second portion 42h2) t2, and the distance from the horizontal portion of the third portion 42h3 to the bottom surface of the groove 46 (described later) (ie, the third portion) 42h3 height) t3 is 5 ⁇ m or more, respectively.
  • the above-described heights t1, t2, and t3 are each 10 ⁇ m or more, more preferably 50 ⁇ m or more, still more preferably 100 ⁇ m or more, and further preferably 250 ⁇ m or more.
  • An annular groove 46 surrounding the peripheral wall 42 is provided between the peripheral wall 42 and the container bottom 10.
  • the groove portion 46 opens upward, and one of the side surfaces of the groove portion 46 is formed in a stepped shape by the outer wall surface 42h of the peripheral wall portion 42, and the other is a vertical surface.
  • the distance to the vertical portion of 42h3 (that is, the width of the third portion 42h3) y2 is 1 mm or less, preferably 0.8 mm or less, and more preferably 0.5 mm or less.
  • the distance X from the vertical portion of the third portion 42h3 to the edge P2 of the upper surface 10a of the adjacent container bottom 10 through the groove 46 is 0.01 mm or more, preferably 0.05 mm or more. Yes, more preferably 0.1 mm or more, and still more preferably 0.4 mm or more. This is because as the distance X is increased, the formed drop is easily held, and when the distance X is increased to a certain level, sufficient holding is obtained. On the other hand, if the distance X is too large, the number of drop forming portions that can be installed in a limited space is reduced, and therefore it is preferably 4 mm or less.
  • the cell handling container 8 configured as described above can obtain the same effects as those of the above-described sixth embodiment, and further exhibits the following effects. That is, since the outer wall surface 42h of the peripheral wall portion 42 is formed in a staircase shape, when the drop S formed on the concave portion 41 and the upper end surface 42c of the peripheral wall portion 42 spreads around due to vibration or the like, the drop S is formed on the outer wall surface. It is possible to extend to the edge of the horizontal part in the second part 42h2 of 42h or the edge of the horizontal part in the third part 42h3 and remain there (see FIG. 20C). As a result, the collapse of the drop S can be prevented.
  • the thickness of the drop S is reduced by the spread of the drop S, but it does not affect the culture and washing of the cells C such as fertilized eggs.
  • the outer wall surface 42h of the peripheral wall portion 42 is a three-step step surface, but may be two steps or four or more steps.
  • FIG. 21A is a plan view showing a cell handling container (with the lid removed) of the ninth embodiment
  • FIG. 21B is a cross-sectional view taken along the line II of FIG. 21A
  • FIG. FIG. 21D is a sectional view taken along line KK in FIG. 21A
  • FIG. 21E is a sectional view taken along line LL in FIG. 21A.
  • the difference between the cell handling container 9 of the present embodiment and the above-described embodiment is a structure in which a plurality of types (here, four types) of drop forming units are combined. Since other configurations are the same as those of the above-described embodiment, a duplicate description is omitted.
  • one drop forming portion having a different structure is provided around the cell accommodation area 12.
  • the structure (see FIG. 21C) and the structure (see FIG. 21D) having the drop forming unit 30 of the fifth embodiment are arranged at the location M3.
  • the structure provided at the place M4 is provided with the surrounding wall portion 33 surrounding the drop forming portion 30 in addition to the drop forming portion 30 of the fifth embodiment.
  • the upper end surface 33a of the surrounding wall portion 33 is flush with the upper end surface 32c of the peripheral wall portion 32 (see FIG. 21E).
  • each drop forming part it is possible to easily form a drop even on a hydrophilic surface.
  • the forming part it is possible to meet various needs with one cell handling container 9 by properly using these drop forming parts as required, and there is an effect of increasing the versatility of the cell handling container 9.
  • FIG. 22A is a plan view showing the cell handling container of the tenth embodiment
  • FIG. 22B is a cross-sectional view taken along the line MM in FIG. 22A.
  • the difference between the cell handling container 53 of the present embodiment and the second embodiment is that the cell storage area 12 is not provided, but a drop forming part 54 is provided in the center of the container bottom 10 instead. Since other configurations are the same as those of the second embodiment, a duplicate description is omitted.
  • a drop forming portion 54 having an upper surface 54a serving as a drop forming region and a side surface 54b extending downward along the entire circumference of the upper surface 54a is provided at the center of the container bottom 10.
  • the drop forming part 54 is formed in a cylindrical shape like the four drop forming parts 16 arranged around it, but is formed wider than the drop forming part 16.
  • the upper surface 54 a of the drop forming portion 54 is a substantially flat surface and is flush with the upper surface 16 a of the drop forming portion 16 and the upper surface 10 a of the container bottom portion 10.
  • a plurality of microwells 55 are formed which are recessed from the upper surface 54a.
  • These microwells 55 each have a substantially cylindrical shape so as to be suitable for accommodating and culturing cells C such as fertilized eggs, and the bottom surface thereof is a conical surface inclined from the outside toward the center.
  • the microwell 55 may have a U shape with a cross section opened upward, a U shape with a circular arc portion, or other shapes, similar to the microwell 13 of the first embodiment.
  • the side surface 54b of the drop forming portion 54 is disposed at a right angle to the upper surface 54a.
  • annular groove 56 surrounding the drop forming portion 54 is provided between the drop forming portion 54 and the container bottom 10.
  • the groove portion 56 is formed in a U shape whose cross section opens upward.
  • the side surface 54 b of the drop forming portion 54 constitutes one side surface of the groove portion 56. Note that the height of the upper surface 54 a of the drop forming portion 54 with respect to the bottom surface of the groove portion 56 is the same as the height t of the upper surface 16 a of the drop forming portion 16 with respect to the bottom surface of the groove portion 17.
  • the distance from the edge of the upper surface 54a of the drop forming part 54 to the edge of the upper surface 10a of the adjacent container bottom 10 via the groove 56 is adjacent to the edge P1 of the upper surface 16a of the drop forming part 16 via the groove 17. It is the same as the distance d to the edge P2 of the upper surface 10a of the container bottom 10.
  • the cell handling container 53 configured as described above can obtain the same effects as those of the second embodiment described above, and further has the following effects. That is, since the drop forming portion 54 is provided, the number of drop formation locations can be increased. Further, since the upper surface 54a of the drop forming portion 54, the upper surface 16a of the drop forming portion 16, and the upper surface 10a of the container bottom portion 10 are flush with each other and there is no barrier, it is possible to slide and move an instrument such as a pipette. The workability can be further improved.
  • FIG. 23A is a plan view showing the cell handling container of the eleventh embodiment
  • FIG. 23B is a cross-sectional view taken along line NN in FIG. 23A.
  • the difference between the cell handling container 57 of the present embodiment and the tenth embodiment is that it further includes a surrounding wall 58 that surrounds the drop forming portion 54, and a groove 59 is provided between the container bottom 10 and the surrounding wall 58. It is. Since other configurations are the same as those of the tenth embodiment, a duplicate description is omitted.
  • annular surrounding wall portion 58 is provided on the outer periphery of the drop forming portion 54 so as to surround the drop forming portion 54.
  • the surrounding wall portion 58 is disposed at a predetermined distance from the side surface 54 b of the drop forming portion 54.
  • An upper end surface 58 a of the surrounding wall portion 58 is flush with the upper surface 54 a of the drop forming portion 54 and the upper surface 10 a of the container bottom portion 10.
  • the cell handling container 57 configured as described above can obtain the same effects as those of the tenth embodiment.
  • FIG. 24 is a perspective view showing a cell handling container (with the lid removed) of the twelfth embodiment.
  • the cell handling container 60 of the present embodiment includes a bottomed cylindrical container body 1a that opens upward, and a lid (not shown) that is detachably installed on top of the container body 1a.
  • the side wall 11 of the container main body 1a extends from the upper surface 10a of the container bottom 10 toward the opening, is formed to be relatively thick, and is positioned above the thick part 11a and the thick wall 11a. It is comprised from the thin part 11b formed thinner than the part 11a.
  • a step portion 11c is formed at a substantially intermediate position of the side wall portion 11 due to a change in thickness between the thick portion 11a and the thin portion 11b.
  • the step portion 11c is a portion that comes into contact with the edge of the lid when the lid is placed on the container body 1a.
  • a cell accommodation area 12 is provided at the center of the container bottom 10.
  • the container bottom 10 has a structure including a drop forming portion 16, an enclosing wall portion 26, and a groove portion 27 (hereinafter referred to as “with an enclosing wall portion and a groove portion in order to avoid complicated explanations) as in the third embodiment.
  • There are six “structures”. These surrounding wall portions and grooved structures are regularly arranged in three pieces on one side with respect to the cell containing area 12 (three on the upper side and three on the lower side in FIG. 24).
  • FIG. 25 is a plan view showing the container body of the cell handling container.
  • the shortest distance L1 from the groove part 27 to the side wall part 11 of the surrounding wall part and the grooved structure is somewhat. I need to leave.
  • the shortest distance between adjacent surrounding wall portions and grooved structures that is, the shortest distance between adjacent groove portions 27
  • L2 the shortest distance between adjacent groove portions 27
  • the shortest distance L3 to the outer wall is preferably 0.5 mm or more, and more preferably 1.0 mm or more.
  • the container bottom 10 the space excluding the surrounding wall and grooved structure, the cell storage area 12, and the outer wall well 61 becomes a plurality of free spaces W. Since the operator can use these free spaces W as necessary, the versatility of the cell handling container 60 can be enhanced.
  • the container bottom portion 10 is provided with a curved wall 62 that rises from the upper surface 10a and is curved in a C shape on the side wall portion 11 side.
  • the left and right ends of the curved wall 62 extend to the side wall portion 11 so as to contact the inside of the side wall portion 11.
  • a space surrounded by the side wall 11 and the curved wall 62 constitutes an outer wall well 61.
  • the curved wall 62 has an arc shape and is configured by a part such as a circle or an ellipse.
  • the curved wall 62 is formed of a part of an ellipse having a major axis of 12 mm and a minor axis of 6 mm.
  • FIG. 26 is a cross-sectional view taken along the line PP in FIG.
  • the curved wall 62 has a uniform thickness and height, and its cross section has a trapezoidal shape whose width decreases from the lower side to the upper side.
  • the container body 1a has an inner diameter of 35 mm
  • the height of the curved wall 62 is 0.5 mm
  • the lower base is 0.5 mm
  • the angle ⁇ ′′ of the trapezoidal side surface with respect to the vertical direction (that is, the vertical direction) is 3 °.
  • a recess 63 is provided on the bottom surface 10 b of the container bottom 10.
  • the depression 63 is recessed inward of the container bottom 10 in a bottom view and has a substantially U shape.
  • the hollow part 63 consists of a part of ellipse whose major axis is 6 mm and whose minor axis is 3 mm, for example.
  • the recess 63 is recessed 3 mm inside the container bottom 10 and extends up to a height of 5 mm upward from the bottom surface 10b.
  • the outer wall well 61 is provided in the container bottom 10
  • a liquid such as a culture solution is placed in the outer wall well 61 to clean the tip of an instrument such as a pipette (that is, Washing and culturing can be performed by placing cells C such as fertilized eggs with a pipette or the like.
  • the amount of liquid for example, 10 ⁇ L or more and 30 ⁇ L or less
  • the amount of liquid for example, 10 ⁇ L or more and 30 ⁇ L or less
  • the outer wall well 61 In addition to being held in the outer wall well 61, it is possible to realize a liquid height (for example, 1.0 mm or more) that is easy to operate when operating an instrument such as a pipette. Furthermore, since the outer wall well 61 is disposed on one side of the container bottom 10 as shown in FIG. 25, the vacant space including the free space W can be utilized to the maximum, and instruments such as pipettes are operated in order from the top. Thus, the work can be carried out efficiently.
  • a liquid height for example, 1.0 mm or more
  • the hollow part 63 is provided in the bottom face 10b of the container bottom part 10, this hollow part 63 can be utilized as marking which identifies visually. Therefore, for example, when observing under a microscope, the direction and position of the cell handling container 60 can be easily grasped using the depression 63 as a mark. In addition, since the operator can grasp the direction and position of the cell handling container 60 by touching the depression 63 with his / her finger, it is possible to perform the work with a tactile sensation without visual observation, thereby improving the work efficiency. it can. Furthermore, when providing the projection part etc. which can be engaged with the hollow part 63 in the mounting surface of a microscope, the position of the cell handling container 60 can be easily fixed by engaging the hollow part 63 and the protrusion part. The risk of shifting the observation position can be reduced.
  • FIG. 28A is a plan view showing a cell handling container of a thirteenth embodiment
  • FIG. 28B is a cross-sectional view taken along the line RR in FIG. 28A
  • FIG. 29A is an enlarged cross-sectional view of a drop forming portion.
  • the difference between the cell handling container 65 of the present embodiment and the second embodiment is that the upper surface 64a and the side surface 64c of the drop forming portion 64 are not directly connected but connected via a curved surface portion 64b having a predetermined radius of curvature R. It is to be done. Since other configurations are the same as those of the second embodiment, a duplicate description is omitted.
  • the drop forming portion 64 has a flat upper surface 64a that is a liquid drop forming region, and a side surface 64c that is connected to the upper surface 64a via a curved surface portion 64b and extends downward.
  • P6 indicates a boundary point between the upper surface 64a and the curved surface portion 64b (that is, the beginning of the curved surface portion 64b)
  • P7 indicates a boundary point between the side surface 64c and the curved surface portion 64b (that is, the end of the curved surface portion 64b).
  • the drop S formed on the upper surface 64a is disposed within a range surrounded by the boundary point P6.
  • P5 is an intersection of a straight line along the upper surface 64a and a straight line along the side surface 64c.
  • the angle ⁇ ′ is an angle formed by a straight line along the upper surface 64a and a straight line along the side surface 64c.
  • ⁇ ′ 90 °.
  • the angle ⁇ ′ becomes an obtuse angle.
  • the diameter D ′ which is the minimum width of the upper surface 64a, is preferably 0.5 mm or more and 10 mm or less, and more preferably 3 mm or more and 6 mm or less, as in the above-described embodiment.
  • the height t ′ of the upper surface 64a of the drop forming part 64 with respect to the bottom surface of the groove part 17 is 5 ⁇ m or more, preferably 10 ⁇ m or more, more preferably 50 ⁇ m or more, even more preferably 100 ⁇ m or more, even more preferably 250 ⁇ m or more, as in the second embodiment. Is most preferred.
  • the distance d ′ from the intersection P5 between the straight line along the upper surface 64a and the straight line along the side surface 64c to the edge P2 of the upper surface 10a of the adjacent container bottom 10 through the groove 17 is the same as that in the second embodiment. Similarly, it is 0.01 mm or more, preferably 0.05 mm or more, more preferably 0.1 mm or more, and further preferably 0.4 mm or more. Further, if the distance d ′ is too large, the number of drop forming portions 64 that can be installed in a limited space is reduced. Therefore, the distance d ′ is preferably 4 mm or less.
  • the radius of curvature R of the curved surface portion 64b is less than 250 ⁇ m. In this way, it is easy to hold the drop formed on the upper surface 64a.
  • the curvature radius R of the curved surface portion 64b is preferably 100 ⁇ m or less, more preferably 70 ⁇ m or less, further preferably 50 ⁇ m or less, and most preferably 20 ⁇ m or less.
  • the drop forming portion 64 having the curved surface portion 64b is also applied to the above-described first, third, fourth, tenth, eleventh and twelfth embodiments, and detailed description thereof is omitted.
  • the example in which the upper surface 64a of the drop forming portion 64 is a liquid drop forming region has been described, but the upper surface 64a and a part of the curved surface portion 64b may be a liquid drop forming region. good.
  • the drop S is formed so as to extend beyond the boundary point P6 to a part of the curved surface portion 64b.
  • FIG. 37A is a plan view showing the cell handling container of the fourteenth embodiment
  • FIG. 37B is a cross-sectional view taken along line BB of FIG. 37A.
  • the difference between the cell handling container 66 of the present embodiment and the second embodiment is that the side surface 16′b of the drop forming portion 16 ′ is provided intermittently along the circumferential direction of the upper surface 16′a. Since other configurations are the same as those of the second embodiment, a duplicate description is omitted.
  • the drop forming portion 16 ′ is disposed flat along the circumferential direction of the upper surface 16′a, which is a liquid drop forming region, and extends downward.
  • Side surface 16'b Due to the intermittent arrangement of the side surface 16'b, a plurality of recesses 17 'are formed in the circumferential direction of the upper surface 16'a. As shown in FIG. 37A, these recesses 17 'each have a substantially fan shape in plan view, and are arranged at equal intervals along the circumferential direction of the upper surface 16'a.
  • the intermittently provided side surface 16 ′ b extends downward from the upper surface 16 ′ a to a range of 5 ⁇ m or more and constitutes a part of the side surface of the recess 17 ′.
  • the side surface 16′b of the drop forming portion 16 ′ is provided intermittently along the circumferential direction of the upper surface 16′a. Since it extends downward from 'a to a range of 5 ⁇ m or more, even if the upper surface 16'a is a hydrophilic surface as in the second embodiment, it is easy to drop the culture medium S on the upper surface 16'a. Can be formed. Since other functions and effects are the same as those of the second embodiment, redundant description is omitted.
  • Example 1 In this example, 7.5 ⁇ L of water was dropped by using the nine types of samples (polystyrene material) shown in Table 1 after the hydrophilic treatment by plasma (water contact angle is 10 ° or less). It was verified whether it could be formed.
  • t is the height of the top surface of the drop forming portion
  • D is the diameter of the top surface of the drop forming portion
  • d is the bottom of the adjacent container bottom via the groove from the edge of the top surface of the drop forming portion.
  • H1 represents the distance from the upper end of the peripheral wall to the upper surface of the container bottom
  • f represents the thickness of the peripheral wall in the horizontal direction (that is, the distance from the inner wall to the outer wall of the peripheral wall).
  • FIG. 3 and no. 4 is a photograph showing the result of 4.
  • Example 2 In this example, No. 1 in Example 1 was used. Whether or not a drop can be formed by dripping water of each liquid amount after hydrophilization treatment with plasma (water contact angle is 10 ° or less) using 8 types of samples excluding 5 (material is polystyrene). Verified. Table 2-1 shows the results of samples (Nos. 1, 2, 6, and 7) corresponding to the drop formation part of the first embodiment, and Table 2-2 shows the samples (No. .3, 4, 8, 9) respectively.
  • the drop forming portion has a concave portion and a peripheral wall portion that forms the concave portion, and the bottom surface of the concave portion is the same as or higher than the top surface of the container bottom portion.
  • each sample shown in Table 3 was prepared by using a lower one. After raising the pipette along the outer wall surface of the peripheral wall portion for each sample, the operability of the pipette was verified according to the procedure of sliding inside the peripheral wall portion and further descending along the inner wall surface to the bottom surface of the recess.
  • the sum of the distance h1 from the upper end portion of the peripheral wall portion to the upper surface of the container bottom portion and the distance h2 from the upper end portion of the peripheral wall portion to the bottom surface of the recess is less than 0.2 mm ( That is, if h1 + h2 ⁇ 0.2 mm), the tip of the pipette hardly touched the peripheral wall (not in contact). It was confirmed that the operability of the pipette could be improved.
  • Example 4 In this example, a comparison was made between the calculated value and the actual measurement value regarding the thickness h ′ of the formed drop.
  • the measured value of the drop thickness h ′ was obtained by dropping water of each liquid amount after the hydrophilization treatment by plasma (water contact angle is 10 ° or less) using the sample shown in Table 4 (material is polystyrene). Thus, the formed drop is actually measured.
  • Example 5 In this example, when the upper surface of the drop forming portion is circular, the influence on the drop formation due to the difference in diameter D was evaluated. Specifically, each sample formed of polystyrene as shown in Table 5 was evaluated by dropping each amount of water after hydrophilization treatment with plasma (water contact angle is 10 ° or less). .
  • the volume of the culture solution is used in the range of 15 ⁇ L or more and 30 ⁇ L or less, and the thickness of the formed drop is 1 mm or more from the viewpoint of ensuring good operability of instruments such as pipettes. Is preferably 1.5 mm or more.
  • the drop thickness is 1.5 mm or less, making it difficult to operate the instrument.
  • the thickness of the drop is 1.0 mm or less, which makes it very difficult to operate the instrument.
  • the drop thickness according to the water contact angle when the drop was formed on a flat surface material: polystyrene
  • Example 6 the drop forming portion (see FIG. 3B) of the second embodiment described above was examined for drop thickness depending on the water contact angle at a liquid volume of 15 ⁇ L and 30 ⁇ L as in Comparative Example 2 (see Table 7).
  • t 250 ⁇ m
  • D 4.0 mm
  • d 0.5 mm
  • t 250 ⁇ m
  • D 5.0 mm
  • d 0.5 mm. there were.
  • Example 6 for example, in the range of 15 ⁇ L or more and 30 ⁇ L or less, even when the water contact angle is less than 80 °, further 60 ° or less, and further 10 ° or less, the drop thickness is 1. Since it was 5 mm or more, it was a result that instruments such as pipettes were easy to operate.
  • Example 7 the sample of the drop formation portion of the second embodiment shown in Table 8 (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less)) from the edge of the upper surface of the drop formation portion. The influence on the drop due to the difference in the distance d to the edge of the upper surface of the adjacent container bottom through the groove was evaluated.
  • Example 8 a sample (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less), liquid volume: 30 ⁇ L) of the drop forming portion (see FIGS. 7A to C) of the third embodiment is used. Then, vibration was applied after the drop was formed, and the drop change before and after the vibration was evaluated.
  • the sample dimensions are shown in Table 9.
  • FIG. 30A is a photograph showing the results of Example 8.
  • FIG. 30A shows photographs after the left side gives vibration and the right side before vibration. From the photograph, it was confirmed that even when vibration was applied, the drop did not collapse and stayed on the surrounding wall.
  • Example 9 a sample (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less), liquid amount: 15 ⁇ L) of the drop forming portion (see FIGS. 8A to C) of the fourth embodiment is used. It was verified whether a drop could be formed.
  • the sample dimensions are shown in Table 10.
  • h1 represents the distance from the upper end surface of the peripheral wall portion to the upper surface of the bottom of the container
  • h2 represents the distance from the upper end surface of the peripheral wall portion to the bottom surface of the recess
  • f represents the thickness of the peripheral wall portion in the horizontal direction. From the results shown in Table 10, when h1 was 50 ⁇ m or more, a drop (liquid amount 40 ⁇ L) could be formed in all samples. Further, when h1 was 250 ⁇ m or more, a drop having a liquid volume of 50 ⁇ L or more and 80 ⁇ L or less could be formed.
  • Example 11 In this example, the conditions described in Table 12 using a sample (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less)) of the drop forming portion (see FIG. 9B) of the fifth embodiment. Whether or not a drop can be formed was verified, and if a drop could be formed, the effect on the drop due to the difference in diameter D was evaluated.
  • h1 represents the distance from the upper end surface of the peripheral wall portion to the upper surface of the bottom of the container
  • h2 represents the distance from the upper end surface of the peripheral wall portion to the bottom surface of the recess
  • f represents the thickness of the peripheral wall portion in the horizontal direction.
  • Example 12 a sample of a drop forming portion (see FIG. 13B) according to a modification of the fifth embodiment (material: polystyrene, after hydrophilization treatment by plasma (water contact angle is 10 ° or less)) is used. It was verified whether or not a drop could be formed under the conditions described in.
  • g in Table 13 indicates the distance of the sharp portion in the vertical direction (that is, the distance from the upper end to the boundary portion between the inclined portion and the vertical portion on the outer wall surface).
  • Example 13 a sample (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less), liquid volume: 30 ⁇ L) of the drop forming portion (see FIGS. 19A to 19C) of the seventh embodiment is used. Then, vibration was applied after the drop was formed, and the drop change before and after the vibration was evaluated.
  • the sample dimensions are shown in Table 14.
  • FIG. 30B is a photograph showing the results of Example 13.
  • FIG. 30B shows photographs after the left side gives vibration and the right side before vibration. From the photograph, it was confirmed that even when vibration was applied, the drop did not collapse and stayed on the surrounding wall.
  • Example 14 a sample (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less), liquid amount: 15 ⁇ L) of the drop forming portion (see FIGS. 20A to C) of the eighth embodiment is used. It was verified whether a drop could be formed.
  • the sample dimensions are shown in Table 15.
  • Example 15 a sample (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° or less)) of a drop formation part (see FIG. 21E) provided at the place M4 of the ninth embodiment, the amount of liquid: 30 ⁇ L) was used to give a vibration after forming a drop, and the drop change before and after the vibration was evaluated.
  • the sample dimensions are shown in Table 16.
  • Example 16 In this example, using 7 types of samples shown in Table 17 (material is polystyrene), 15 ⁇ L of water can be dropped after plasma hydrophilic treatment (water contact angle is 10 ° or less) to form a drop. It was verified whether or not.
  • any of the samples shown in Table 17 could form a drop.
  • Example 17 In this example, a sample (material: polystyrene, after hydrophilization treatment by plasma) processed to have a curved surface portion between the upper surface and the side surface as shown in FIG. 33 with respect to the drop forming portion of the sixth embodiment described above. (Water contact angle is 10 ° or less)) was used to verify whether a drop could be formed. The amount of liquid used was 30 ⁇ L and 50 ⁇ L. Table 18 shows the sample dimensions. In Table 18, R is the radius of curvature of the curved surface portion, ⁇ ′ is the angle formed by the straight line along the top surface of the drop forming portion and the straight line along the side surface as described in the thirteenth embodiment, and d is the top surface. The distance from the intersection P5 between the straight line along the straight line and the straight line along the side surface to the edge P2 of the upper surface of the adjacent container bottom through the groove.
  • Example 18 In this example, as shown in FIG. 34, a sample (material: polystyrene, plasma treatment for hydrophilization) processed to have a curved surface portion between the upper surface and the side surface of the drop forming portion of the sixth embodiment. The later (water contact angle is 10 ° or less) was used to verify whether a drop could be formed. The liquid volume used was 15 ⁇ L.
  • Table 19 shows the sample dimensions. In Table 19, R is the radius of curvature of the curved surface portion, ⁇ ′ is the angle formed by the straight line along the top surface of the drop forming portion and the straight line along the side surface as described in the thirteenth embodiment, and d is the top surface. The distance from the intersection P5 between the straight line along the straight line and the straight line along the side surface to the edge P2 of the upper surface of the adjacent container bottom through the groove.
  • Example 19 based on each condition shown in FIG. 35, a sample of a drop forming portion in which the upper surface and the side surface are connected via a curved surface portion (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° The following)) was used to verify whether a water drop could be formed.
  • the liquid volume used was 10 microliters, 15 microliters, and 20 microliters.
  • R is a curvature radius of the curved surface portion
  • ⁇ ′ is an angle formed by a straight line along the upper surface and a straight line along the side surface in the thirteenth embodiment.
  • Example 20 based on each condition shown in FIG. 36, a sample of a drop forming portion in which the upper surface and the side surface are connected via a curved surface portion (material: polystyrene, after hydrophilization treatment with plasma (water contact angle is 10 ° The following)) was used to verify whether a culture drop could be formed.
  • the liquid volume used was 10 microliters, 15 microliters, and 20 microliters.
  • the culture solution used was Modified HTF Medium manufactured by Irvine Scientific.
  • R is a radius of curvature of the curved surface portion
  • ⁇ ′ is an angle formed by a straight line along the upper surface and a straight line along the side surface in the thirteenth embodiment.
  • FIG. 38A to C are photographs of the cell handling container used in Example 21, FIG. 38A is a cell handling container with a lid, FIG. 38B is a cell handling container without a lid, and FIG. 38C is a plan view of a cell handling container without a lid. It is a photograph which shows.
  • the cell handling container used in this example is processed so that the drop forming part and the surrounding wall part have curved parts with respect to the cell handling container 60 of the twelfth embodiment, and the drop forming part and the surrounding wall part
  • the side surfaces are processed so as to be inclined.
  • the specific shapes of the drop forming portion and the surrounding wall portion are as shown in FIG. In FIG.
  • R is the radius of curvature of the curved surface portion
  • ⁇ ′′ is the inclination angle of the side surface with respect to the vertical direction (ie, the vertical direction), and the angle ⁇ ′ described above (ie, as described in the thirteenth embodiment).
  • D ′ 5 mm
  • t ′ 250 ⁇ m.
  • Table 21 shows the evaluation results of this example. As can be seen from Table 21, in all types of cell handling containers, the drop was retained in the drop formation section.
  • Table 22 shows the evaluation results after vibration. As can be seen from Table 22, when vibration was applied, it was confirmed that the drop S (see FIG. 40A) spread to the surroundings due to the vibration, but remained in the surrounding wall portion without collapsing (see FIG. 40B). It was also confirmed that the thickness of the drop S was reduced by spreading.
  • the present invention is not limited to the above-described embodiments, and various designs can be made without departing from the spirit of the present invention described in the claims. It can be changed.
  • the cell handling container provided with the microwell has been described.
  • the present invention is also applied to the cell handling container not provided with the microwell.
  • the present invention is also applied to a cell handling container that does not include the lid 1b.
  • the circular drop forming portion is described in plan view, but the present invention is also applicable to a polygon such as a rectangle, an ellipse, etc. in plan view. Furthermore, the above-described embodiment and modification are only a part of the present invention, and the present invention is also applied to each combination of the above-described embodiment and modification.

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Abstract

La présente invention concerne un récipient de manipulation de cellules (1) comprenant une zone de stockage de cellules (12) ayant des micropuits (13), et une pluralité de parties de formation de gouttes (15) disposées autour de la zone de stockage de cellules (12). Chacune des parties de formation de gouttes (15) ayant une surface supérieure (15a) qui sert de zone de formation de gouttes liquides, et une surface latérale (15b) s'étendant vers le bas le long de la périphérie entière de la surface supérieure (15a). La surface supérieure (15a) de la partie de formation de goutte (15) est plate. Dans le sens vertical, la surface latérale (15b) de la partie de formation de goutte (15) s'étend vers le bas d'au moins 5 µm depuis le bord (P1) de la surface supérieure (15a).
PCT/JP2017/033209 2016-09-30 2017-09-14 Récipient de manipulation de cellules WO2018061795A1 (fr)

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Cited By (1)

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WO2019225633A1 (fr) * 2018-05-24 2019-11-28 国立研究開発法人量子科学技術研究開発機構 Micropuce d'échantillon biologique, couvercle, trousse de scellement d'échantillon biologique, et procédé

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JP6338037B1 (ja) * 2016-09-30 2018-06-06 大日本印刷株式会社 細胞取扱容器
US20220150285A1 (en) 2019-04-01 2022-05-12 Sumitomo Electric Industries, Ltd. Communication assistance system, communication assistance method, communication assistance program, and image control program

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US20070105185A1 (en) * 2003-05-16 2007-05-10 Cima Michael J Raised surface assay plate
JP2016082987A (ja) * 2016-02-17 2016-05-19 大日本印刷株式会社 細胞培養容器
JP2016106635A (ja) * 2013-09-11 2016-06-20 大日本印刷株式会社 細胞培養容器
JP2016129495A (ja) * 2015-01-13 2016-07-21 大日本印刷株式会社 細胞培養容器
JP2016144422A (ja) * 2015-02-06 2016-08-12 秋田県 液滴形成用シャーレ及びそれを用いた電界撹拌方法

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JP6338037B1 (ja) * 2016-09-30 2018-06-06 大日本印刷株式会社 細胞取扱容器

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US20070105185A1 (en) * 2003-05-16 2007-05-10 Cima Michael J Raised surface assay plate
JP2016106635A (ja) * 2013-09-11 2016-06-20 大日本印刷株式会社 細胞培養容器
JP2016129495A (ja) * 2015-01-13 2016-07-21 大日本印刷株式会社 細胞培養容器
JP2016144422A (ja) * 2015-02-06 2016-08-12 秋田県 液滴形成用シャーレ及びそれを用いた電界撹拌方法
JP2016082987A (ja) * 2016-02-17 2016-05-19 大日本印刷株式会社 細胞培養容器

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2019225633A1 (fr) * 2018-05-24 2019-11-28 国立研究開発法人量子科学技術研究開発機構 Micropuce d'échantillon biologique, couvercle, trousse de scellement d'échantillon biologique, et procédé

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JP2019000103A (ja) 2019-01-10
JP2019110923A (ja) 2019-07-11
JP6741112B2 (ja) 2020-08-19
JP6516041B2 (ja) 2019-05-22
JPWO2018061795A1 (ja) 2018-09-27

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