JP6892215B2 - Cell handling container - Google Patents

Description

The present invention relates to a cell handling container that can be used when handling cells such as fertilized eggs by the microdrop method or the like.

In vitro fertilization of sperms and eggs in a culture system to produce fertilized eggs (zygotes), and then the fertilized eggs are cleaved, morula, and blastocyst, and then the escaped scutellum hatched from the zona pellucida. It has become possible to culture up to the blastocyst stage. For this reason, assisted reproductive technology (ART), in which fertilized eggs at the stage of cleavage to blastocyst are transplanted into the uterus to obtain offspring, has been established not only in the domestic field but also in human infertility medicine.

However, the success rate of pregnancy by in vitro fertilization is not always high. For example, in humans, the pregnancy success rate is still around 25-35%. One of the causes is that there is not a high probability that a good quality fertilized egg suitable for transplantation into the uterus can be obtained in culture. The cultured fertilized eggs are individually observed by a specialist under a microscope to determine whether or not they are good quality fertilized eggs suitable for transplantation into the uterus. Therefore, there is a problem that the determination requires cost and time. Therefore, there is a demand for a technique for discriminating high-quality fertilized eggs suitable for transplantation into the uterus at low cost and in a short time.

Further, in recent years, research and development of cell therapy / regenerative medicine have progressed, and there is an increasing need for maintenance of a sterile environment during cell culture, quality assurance of cells, and / or rapid culture and discrimination means of a large amount of cells. Therefore, it is required to culture a plurality of fertilized eggs in parallel in one cell handling container.

For example, Patent Document 1 is a culture vessel having a bottom wall and a side wall for culturing cells that require individual management, and a cell accommodating portion having a recess is arranged on the bottom wall. Four or more recesses are close to each other, the opening of the recess is circular, the wall surface of the recess has an inclined surface that increases from the lowest position of the recess toward the outer edge of the recess, and is conical or conical. Described the culture vessel having a truncated cone shape, a pitch between adjacent recesses of 1 mm or less, and a maximum height Ry of less than 1.0 μm for surface roughness of an inclined surface.

Patent Document 2 is a cell culture container having a bottom and a side wall and provided with a culture system storage compartment having a predetermined capacity inside, and the culture system storage compartment has a bottom surface having a uniform thickness and a bottom surface. It is composed of a wall surface and a gentle curved surface connecting the bottom surface and the wall surface, and the wall surface of the culture system storage compartment is arranged so as to be inclined outward by 5 to 20 ° with respect to the vertical direction of the bottom surface. A cell culture container characterized by the above is described. The document describes that a plurality of culture system containment compartments may be arranged.

Patent Document 3 is a cell culture vessel having an open upper part, from a recess for accommodating cells and a culture solution at the bottom, and a wall portion formed so as to increase from the open end of the recess toward the outer edge. The side surface of the recess is formed so as to increase from the bottom surface of the recess toward the outer edge, and the angle θ1 between the bottom surface and the side surface of the recess is the wall surface of the culture solution storage portion. The cell culture vessel in which the angle between the cell and the bottom surface of the recess is larger than θ2 and θ1 is 75 ° or less is described. The document describes an embodiment in which a recess is formed on the bottom surface of the recess for positioning cells.

Japanese Patent No. 4724854 Japanese Unexamined Patent Publication No. 2006-280298 Japanese Unexamined Patent Publication No. 2014-207897

As described above, a container having a plurality of recesses such as microwells for accommodating cells is known. In these containers, cells such as fertilized eggs are usually housed in a recess and handled for culturing or the like. Such a container is suitable for classifying cells into a plurality of categories based on criteria such as developmental status, and culturing cells belonging to each category in parallel while individually managing them in one container. ..

When handling cells using a cell handling container having a recess such as a microwell, the presence of foreign matter such as dust or plastic pieces inside the recess is not preferable for observing and / or handling the cells. May affect. Therefore, in the manufacture of a cell handling container, it is usually shipped after inspecting that no foreign matter is present inside the recessed portion. However, if a foreign substance having a size capable of entering the inside of the recess is present in the internal space of the cell handling container, the foreign substance may enter the inside of the recess after shipment.

When the user discovers that a foreign substance is present inside the recess when using the cell handling container, the user either removes the foreign substance or discontinues the use of the recess containing the foreign substance and another depression. It is necessary to perform processing such as using a part. Such processing not only may reduce the work efficiency of the user, but may also cause mistakes.

Therefore, the present invention can substantially prevent problems caused by foreign substances that can invade the inside of the recessed portion that accommodates cells in a cell handling container that handles cells that require individual management such as fertilized eggs. The purpose is to provide a container.

The present inventors have studied various means for solving the above problems. In a cell handling container having a recessed portion, the present inventors set the number of foreign substances having a maximum length equal to or less than the opening width of the recessed portion at the bottom of the container to be less than the number of the recessed portions, thereby making the foreign substance into a foreign substance. We have found that the resulting problems can be substantially prevented. The present inventors have completed the present invention based on the above findings.

That is, the gist of the present invention is as follows.
(1) A cell handling container having a container bottom and a container side wall.
At the bottom of the container, n recesses having an opening width r are arranged, where n is an integer of 1 or more.
At the bottom of the container, there are m foreign substances with a maximum length p, where m is an integer greater than or equal to 0.
The opening width r of the recess and the maximum length p of the foreign matter satisfy r ≧ p.
A cell handling container in which the number n of dents and the number m of foreign matter satisfy n-1 ≧ m.
(2) The cell handling container according to (1) above, wherein the opening width r of the recess is in the range of 100 to 1000 μm.
(3) The cell handling container according to (1) or (2) above, wherein the opening width r of the recess and the maximum length p of the foreign substance satisfy r / 2 ≦ p.
(4) In any of the above (1) to (3), a storage compartment having a storage compartment bottom and a storage compartment side wall is arranged at the bottom of the container, and a recess is arranged at the bottom of the storage compartment. The cell handling container described.
(5) The number n of depressions existing at the bottom of the containment compartment and the number m'of foreign matter existing at the bottom of the containment compartment satisfy n-1 ≥ m', where m'is an integer greater than or equal to 0 and less than or equal to m. The cell handling container according to (4) above.
(6) The method for manufacturing a cell handling container according to any one of (1) to (5) above.
A container forming step of forming a cell handling container having a container bottom and a container side wall and having n recesses having an opening width r arranged at the container bottom;
A foreign matter removing step of reducing the number m of foreign matter having a maximum length p at the bottom of the container to n-1 or less by a foreign matter removing means using a gas flow, a liquid flow or a transfer member;
Including
The number n of the recesses is an integer greater than or equal to 1.
The number m of foreign matter is an integer greater than or equal to 0,
The opening width r of the recess and the maximum length p of the foreign matter satisfy r ≧ p.
The above method, wherein the number n of the recesses and the number m of the foreign matter satisfy n-1 ≧ m.
(7) In the container forming step, a cell handling container in which a storage compartment having a storage compartment bottom and a storage compartment side wall is arranged at the bottom of the container and a recess is arranged at the bottom of the storage compartment is formed. Including
The foreign matter removing step reduces the number m'of foreign matter having a maximum length p placed at the bottom of the storage compartment to n-1 or less by a foreign matter removing means using a gas flow, a liquid flow or a transfer member. Including
The number n of dents existing at the bottom of the containment compartment and the number m'of foreign matter existing at the bottom of the containment compartment satisfy n-1 ≥ m', where m'is an integer greater than or equal to 0 and less than or equal to m. The method according to (6) above.

INDUSTRIAL APPLICABILITY The present invention provides a cell handling container that can substantially prevent problems caused by foreign substances that can invade the inside of a recess that houses cells in a cell handling container that handles cells that require individual management such as fertilized eggs. It becomes possible to do.

It is a top view of one Embodiment of the cell handling container of this invention. FIG. 5 is a schematic vertical cross-sectional view taken along the line I-I'in FIG. 1 in one embodiment of the cell handling container of the present invention. FIG. 5 is a schematic vertical cross-sectional view taken along the line II-II'in FIG. 1 in one embodiment of the cell handling container of the present invention. FIG. 5 is a schematic vertical cross-sectional view taken along the line II-II'in FIG. 1 in one embodiment of the cell handling container of the present invention. FIG. 5 is a schematic vertical cross-sectional view taken along the line II-II'in FIG. 1 in one embodiment of the cell handling container of the present invention. It is a top view of one Embodiment of the cell handling container of this invention. FIG. 6 is a schematic vertical cross-sectional view taken along the line III-III'in FIG. 6 in one embodiment of the cell handling container of the present invention. It is a schematic diagram which shows the embodiment of the microdrop method using the cell handling container of this invention. In Example I-1, it is a photograph showing the relationship between the recessed portion at the bottom of a circular petri dish and the foreign matter existing on the inner surface of the bottom of the storage compartment. (A): Relationship between the foreign matter existing on the inner surface of the bottom of the storage compartment and the opening of the recess; (b): Foreign matter entering the inside of the recess. In Example I-1, it is a graph which shows the relationship between the distance between a circular petri dish and an air gun, and the wind speed of a gas flow. In Example I-1, it is a photograph which shows the foreign matter existing on the inner surface of the bottom of the accommodation section of a circular petri dish before and after the treatment of the foreign matter removing means using a gas flow. (A): Inner surface of the bottom of the petri dish before treatment and foreign matter present there; (b): Inner surface of the bottom of the petri dish after treatment with a gas flow of 6.2 m / s and present there Foreign matter; (c): Petri dish storage after treatment with a gas flow of 9.0 m / s The inner surface of the bottom of the compartment and foreign matter existing there; (d): Storage of the petri dish after treatment with a gas flow of 12.4 m / s Inner surface of the bottom of the compartment. In the figure, circles indicate foreign substances. In Example I-2, it is a photograph which shows the foreign matter existing on the inner surface of the accommodation section bottom of the circular petri dish before and after the treatment of the foreign matter removing means using a liquid flow. (A): Inner surface of the bottom of the petri dish before treatment and foreign matter present there; (b): Inner surface of the bottom of the petri dish after treatment with a liquid flow of 70 ml / min; (c): Inner surface of the bottom of the petri dish's containment compartment after treatment with a liquid flow of 600 ml / min. In Example I-3, it is a photograph which shows the foreign matter existing on the inner surface of the accommodation section bottom of the circular petri dish before and after the processing of the foreign matter removing means using a transfer member. (A): Inner surface of the bottom of the petri dish storage compartment before treatment and foreign matter present there; (b): Inside the bottom of the petri dish storage compartment after contacting the cleaning stick with a contact pressure of ^{0.51 to 2.04 kgf / cm 2.} Surface and foreign matter present therein; (c): Inner surface of the bottom of the petri dish's containment compartment after contacting the cleaning stick with a contact pressure of ^{2.04-6.12 kgf / cm 2.}

In the present specification, the features of the present invention will be described with reference to the drawings as appropriate. In the drawings, the dimensions and shapes of each part are exaggerated for clarification, and the actual dimensions and shapes are not accurately depicted. Therefore, the technical scope of the present invention is not limited to the dimensions and shapes of the parts shown in these drawings.

<1: Cell handling container>
As shown in FIG. 1, the cell handling container of the present invention is, for example, a cell handling container 1 having a container bottom portion 11 and a container side wall portion 12, and n recesses having an opening width r in the container bottom portion 11. It is necessary that the part 20 is arranged. However, n is an integer of 1 or more.

An embodiment of the cell handling container 1 of the present invention is shown in FIGS. 1 to 4. FIG. 2 is a schematic vertical cross-sectional view taken along the line I-I'in FIG. 3 and 4 are schematic vertical cross-sectional views taken along the line II-II'in FIG. As shown in FIG. 2, the cell handling container 1 of the present invention has a container bottom portion 11 and a container side wall portion 12 erected on the peripheral edge of the container bottom portion 11. The cell handling container 1 of the present invention is the surface of the container bottom 11 facing the space side on which the cells and / or the culture solution are arranged (hereinafter, “the inner surface of the container bottom 11 facing the space side on which the cells and / or the culture solution is arranged”). , "Inner surface of container bottom 11" or "Inner bottom surface of container"), and surface of container side wall 12 facing the space side where cells and / or culture solution is placed (hereinafter, "cells and / or culture"). An upwardly open bottomed internal space formed by the "inner surface of the side wall portion 12 of the container facing the space where the liquid is placed", "inner surface of the side wall portion 12 of the container" or "inner wall surface of the container"). Have 10. The internal space 10 is a space for arranging cells and / or the culture medium. The cell handling container 1 of the present invention is usually the surface of the container bottom 11 facing the external space side of the cell handling container 1 (hereinafter, "the outer surface of the container bottom 11 facing the external space side of the cell handling container 1", " (Also referred to as "outer surface of container bottom 11" or "outer bottom surface of container") is arranged and used in a horizontal state. In the present invention, the arrangement of the cell handling container of the present invention as described above may be described as "arrangement at the time of use". In the present invention, the "vertical direction" means a direction perpendicular to the outer bottom surface of the container in the arrangement during use. In this case, the direction from the outer bottom surface of the container to the opening formed by the peripheral edge of the side wall of the container may be described as upward, and the direction from the opening toward the outer bottom surface of the container may be described as downward. In the present invention, "height" means the vertical length of each part of the cell handling container of the present invention in the arrangement at the time of use. Further, in the present invention, the "horizontal direction" means a direction parallel to the outer bottom surface of the container in the arrangement at the time of use.

The shape and dimensions of the cell handling container 1 of the present invention are not particularly limited as long as n recesses 20 can be arranged on the bottom 11 of the container. As the shape of the cell handling container 1, for example, a figure formed by the peripheral edge of the inner surface of the container bottom 11 facing the space side where the cells and / or the culture solution are arranged is projected onto a horizontal plane in the arrangement at the time of use. , Circular (including perfect circle and oval), or polygonal shapes such as triangle, quadrangle, pentagon, hexagon or octagon. It is preferable that the figure formed by the peripheral edge of the inner surface of the bottom 11 of the container facing the space where the cells and / or the culture medium are arranged in use is circular when projected onto a horizontal plane. In this case, the cell handling container 1 of the present invention has the shape of a circular petri dish usually used in the art. The dimensions of the cell handling container 1 are typically in the range of 30-60 mm, for example 35-60 mm, in the top view of the arrangement in use, where the opening width of the side wall 12 of the container is in the range of 30-60 mm. The target is about 35 mm. In the present invention, the "opening width of the opening" means the length of the longest diameter of the figure formed by the outer edge of the opening. For example, if the outer edge of the opening is circular, the dimensions of the opening correspond to the diameter of the circle. In addition, the height of the side wall portion 12 of the container is usually in the range of 5 to 20 mm, and typically in the range of 7 to 15 mm. In a particularly suitable cell handling container 1, the opening width of the opening of the side wall portion 12 of the container is preferably in the range of 30 to 60 mm, more preferably in the range of 35 to 60 mm, and particularly preferably in the range of 35 to 60 mm in terms of top view in the arrangement during use. Is about 35 mm, the height of the container side wall 12 is preferably in the range of 5 to 20 mm, more preferably in the range of 7 to 15 mm, and the container bottom 11 has a circular shape. The cell handling container 1 having such dimensions and shapes has the same shape and dimensions as a circular petri dish usually used for culturing fertilized eggs and the like in the cell culture field. Therefore, the cell handling container of the present invention having the above-mentioned shape and dimensions can be easily adapted to various culture devices and the like usually used in the cell culture field.

The shape of the bottom 11 of the container is not particularly limited. The inner surface of the bottom 11 of the container facing the space side on which the cells and / or the culture medium are arranged may be formed so that substantially the entire surface is a horizontal plane in the arrangement at the time of use, and proceeds from the center to the outer edge in the arrangement at the time of use. It may be formed so as to be an inclined surface that becomes higher in the vertical direction according to the above. Various shapes of the container bottom of a cell handling container such as a cell culture container known in the art can be applied to the shape of the container bottom 11.

Similarly, the shape of the side wall portion 12 of the container is not particularly limited. The inner surface of the side wall portion 12 of the container facing the space side where the cells and / or the culture solution are arranged, and / or the surface of the side wall portion 12 of the container facing the outer space side of the cell handling container 1 (hereinafter, "outside of the cell handling container 1"). The "outer surface of the side wall portion 12 of the container facing the space side" or "the outer surface of the side wall portion 12 of the container") may be formed so that substantially the entire surface is a vertical surface in the arrangement at the time of use. In the arrangement of time, it may be formed so as to be an inclined surface that is inclined in the inner direction of the container side wall portion 12 with respect to the vertical surface as it advances from the connection portion with the container bottom portion 11 to the upper peripheral portion. Various shapes of the side wall portion of the cell handling container such as the cell culture container known in the art can be applied to the shape of the side wall portion 12 of the container. When the cell handling container 1 of the present invention is made of a material containing a plastic material and is manufactured by a conventional molding method such as injection molding, a container facing the space side where cells and / or a culture solution is placed. If the inner surface of the side wall portion 12 and / or the outer surface of the container side wall portion 12 facing the outer space side of the cell handling container 1 is formed to be an inclined surface as described above, it functions as a draft and is efficient. It is advantageous because it can be removed from the mold.

As shown in FIG. 3, n recesses 20 having an opening width r are arranged on the bottom 11 of the container. The number n of the recess 20 is usually an integer greater than or equal to 1. When culturing fertilized eggs using the cell handling container 1 of the present invention, by handling the fertilized eggs to be cultured at the same time in one cell handling container 1, it is possible to prevent work mistakes such as mistaking the container, and the container Since management is easy, the number n of the recessed portions 20 is preferably 1 or more. By managing a plurality of fertilized eggs as one group and culturing them at the same time, the management becomes easier. Therefore, the number n of the recesses 20 is more preferably 3 or more. In addition, when handling too many fertilized eggs in one cell handling container 1, the time required to observe individual fertilized eggs outside the incubator increases, and there is a risk of temperature changes and / or foreign matter contamination. The number n of the recesses 20 is preferably 50 or less, more preferably 30 or less, and even more preferably 25 or less, because it may increase. n is preferably in the range of 1 to 50, more preferably in the range of 3 to 30. The number n of the recesses 20 is the number that can form a symmetrical matrix-like arrangement having the same number of recesses 20 in the vertical and horizontal directions, for example, 25 (5 × 5 matrix-like arrangement), 16 (4 × 4). Matrix-like arrangement) or 9 (3 × 3 matrix-like arrangement) is preferable. In the case of the above embodiment, the number n of the recesses 20 is the number n that can form an arrangement in which the recess is not present in the central portion in a symmetric matrix, for example, 24 (5 × 5 matrix-like arrangement) or 8 (. 3 × 3 matrix arrangement) is more preferable. By arranging the cells inside the recess 20, the movement of the cells can be suppressed. As a result, even when a plurality of cells are handled in one cell handling container 1, it is possible to evaluate or determine a specific cell while distinguishing each cell.

The shape of the recess 20 is not particularly limited. As the shape of the recess 20, for example, the figure formed by the peripheral edge of the opening is a circle (including a perfect circle and an ellipse), or a polygon such as a triangle, a quadrangle, a pentagon, a hexagon, or an octagon. The shape can be mentioned. As shown in FIG. 1, it is preferable that the figure formed by the peripheral edge of the opening of the recess 20 is circular. The bottom portion 21 of the recess 20 may be formed so that substantially the entire surface is a horizontal plane in the arrangement during use, as shown in FIG. 3, for example, or as shown in FIG. 4, from the center in the arrangement during use. It may be formed so as to be an inclined surface that becomes higher in the vertical direction toward the outer edge. As shown in FIGS. 2 and 3, for example, the side wall portion 22 of the recess portion 20 is the surface of the side wall portion 22 facing the space side where the cells and / or the culture medium are arranged in the arrangement during use (hereinafter, “cells and / or”. Approximately the entire surface of the "inner surface of the side wall portion 22 facing the space side on which the culture solution is placed" or "inner surface of the side wall portion 22") may be formed to be a vertical surface, or during use. In the arrangement, the side wall portion 22 may be formed so as to be an inclined surface that is inclined inward with respect to the vertical surface as it advances from the connection portion with the bottom portion 21 to the upper peripheral portion. The recess 20 is the surface of the bottom 21 facing the space side where the cells and / or the culture medium are placed (hereinafter, “the inner surface of the bottom 21 facing the space side where the cells and / or the culture medium is placed” or “the bottom 21”. (Also referred to as "inner surface"), and the inner surface of the side wall portion 22 facing the space side where the cells and / or the culture medium are arranged advances from the lowest position in the vertical direction to the peripheral edge of the upper opening in the arrangement in use. It may have a curved surface that is continuously inclined so as to become higher according to. In the case of the present embodiment, the inner surface of the bottom portion 21 facing the space side where the cells and / or the culture medium is arranged and the inner surface of the side wall portion 22 facing the space side where the cells and / or the culture medium are arranged are combined. It is preferable to form a shape including a conical or truncated cone-shaped portion. In this case, the conical or truncated cone-shaped portion is oriented toward the bottom 11 side of the container (that is, the lower side in the arrangement at the time of use), with the apex of the cone or the upper surface and the lower surface of the truncated cone having the smaller area oriented. Is formed to do. In the present embodiment, the conical shape is not only a cone and an elliptical cone, but also a shape in which the apex of the cone or the elliptical cone is rounded, a shape in which the conical surface is bulging outward, and a conical surface in which the conical surface is recessed inward. Shapes similar to these, such as shapes, are also included. The truncated cone shape is not limited to the truncated cone and the truncated cone, but also has a rounded joint between the upper surface or the lower surface of the truncated cone or the truncated cone and the conical surface, and the conical surface bulges outward. It also includes shapes similar to these, such as shapes and shapes in which the conical surface is recessed inward. Further, the inner surface of the bottom 21 facing the space side where the cells and / or the culture solution of the recess 20 is arranged, and the inner surface of the side wall portion 22 facing the space side where the cells and / or the culture solution are arranged have the above-mentioned shape. Other shapes, such as, but not limited to, a polygonal pyramid or a polygonal frustum, can also be formed.

The size of the recess 20 is not particularly limited as long as it can accommodate at least one cell inside the recess 20. In the present invention, the "dimension of the recessed portion" is defined by the opening width r and the depth d of the recessed portion 20. As shown in FIGS. 3 and 4, the opening width r of the recess 20 means the length of the longest diameter of the figure formed by the peripheral edge of the opening. The depth d of the recess 20 means the vertical length from the lowest position in the vertical direction of the bottom portion 21 to the plane including the peripheral edge of the upper opening in the arrangement at the time of use. For example, when the peripheral edge of the opening of the recess 20 is circular, the opening width r corresponds to the diameter thereof. The opening width r and the depth d may be values equal to or larger than the outer diameter of the contained cells. For example, when a fertilized egg is cultured using the cell handling container 1 of the present invention, it is usually cultured up to the stage of blastocyst, particularly from the early stage of blastocyst to the stage of prolapsed blastocyst via expanded blastocyst. .. In this case, the opening width r and the depth d of the recess 20 are preferably values larger than the outer diameter of the cell at the stage of the blastocyst. For example, in the case of fertilized human eggs, the outer diameter of cells in the early blastocyst stage is usually 100 μm to 280 μm, and the outer diameter of cells in the prolapsed blastocyst stage that escapes from the transparent zone is usually. Is 200 μm to 600 μm, so that the opening width r and the depth d of the recess 20 are usually 100 μm or more, and 1000 μm or less. In non-human mammals, such as fertilized bovine eggs, the outer diameter of cells at the early blastocyst stage is typically 100 μm to 200 μm, at the stage of prolapsed blastocysts escaping from the zona pellucida. Since the outer diameter of cells is usually 300 μm to 600 μm, and in the case of fertilized mouse eggs, the outer diameter of cells at the early blastocyst stage is usually 60 μm to 100 μm. Even when culturing a non-human mammal to an escaped blastocyst, the opening width r and depth d of the recess 20 may usually be 1000 μm or less.

For example, when culturing fertilized eggs of humans or non-human mammals such as cows or mice using the cell handling container 1 of the present invention, if the opening width r of the recess 20 is too small, it is at the stage of early embryo. Since it may be difficult to culture for several hours, it is usually 100 μm or more, preferably 200 μm or more, and more preferably 250 μm or more. Further, if the opening width r of the recessed portion 20 is too large, it may be difficult to arrange the plurality of recessed portions 20 in the container bottom portion 11, and / or the inside of the container bottom portion 11 other than the recessed portion 20. Since the work space on the surface may be reduced and workability may be reduced, it is usually 1000 μm or less, preferably 800 μm or less, more preferably 500 μm or less, and 300 μm or less. Is more preferable. For example, the opening width r of the recess 20 is preferably in the range of 100 to 1000 μm, more preferably in the range of 250 to 500 μm. The depth d is usually 100 μm or more, preferably 200 μm or more, more preferably 250 μm or more, and usually 1000 μm or less, preferably 800 μm or less. It is preferably 500 μm or less, more preferably 300 μm or less. For example, the depth d of the recess 20 is preferably in the range of 50 to 1000 μm, more preferably in the range of 150 to 500 μm. When the depth d of the recess 20 is less than the lower limit, the cell may not fit in the recess 20 when the cell grows, and the upper part of the cell may be exposed from the opening of the recess 20. In such a case, especially when most of the cells are exposed from the opening of the recess 20, the cells are likely to come out of the recess 20, so it is necessary to perform various operations extremely carefully. When the depth d of the recess 20 is equal to or greater than the upper limit, it may be difficult to remove cells from the recess 20. Therefore, the depth d of the recess 20 is preferably within the above range. Alternatively, the opening width r and the depth d of the recess 20 can be defined as X + α (where X represents the outer diameter of the cell). Here, α is preferably 0.01 mm or more, and more preferably 0.02 mm or more. When the opening width r and the depth d of the recessed portion 20 are at least the above lower limit values, it is possible to secure a space for handling the cells inside the recessed portion 20 even at the stage when the cells (for example, fertilized egg) have grown. Further, when the opening width r and the depth d of the recessed portion 20 are equal to or less than the upper limit values, even if the plurality of recessed portions 20 are arranged, the entire plurality of recessed portions 20 can be included in one field of view of the microscope. The cells can be efficiently observed with a microscope.

The depth d of the recess 20 is preferably 0.2 times or more, more preferably 0.5 times or more, more preferably 2 times or less, and more preferably 1 time or less of the opening width r. preferable. When the opening width r and the depth d of the recess 20 satisfy the above relationship, the cells can be easily accommodated and taken out from the recess 20. Further, as described below, even when a foreign matter has entered the inside of the recessed portion 20, the foreign matter can be easily removed.

In the cell handling container 1 of the present invention, the bottom portion 11 of the container has m foreign substances 100 having a maximum length p. However, m is an integer of 0 or more. When m is 0, the number of foreign matter 100 having the maximum length p at the bottom 11 of the container is zero, which means that there is no foreign matter 100 having the maximum length p at the bottom 11 of the container. In the present invention, the "maximum length" means the maximum length of the longest diameter of a figure formed by the peripheral edge of an object (for example, a foreign object) when it is projected onto a plane from an arbitrary direction. The maximum length p of the foreign matter 100 can be determined, for example, by measuring the length of the foreign matter 100 from a plurality of directions using a microscope and obtaining the maximum length thereof.

In a conventional cell handling container having a recess for containing cells at the bottom of the container, it is usually shipped after inspecting the inside of the recess for foreign matter such as dust or plastic pieces. To. Therefore, normally, at the time of product shipment, there is no foreign matter inside the recessed portion of the cell handling container. In the cell handling container having such a recess, if foreign matter such as dust or plastic pieces is present at the bottom of the container, the present inventors will use the cell handling container until or after the start of use. It was found that foreign matter could enter the inside of the recess. In addition, the present inventors have found that various problems can occur due to the foreign matter that has entered the inside of the recess. For example, if the cells are handled with the foreign matter invading the inside of the recessed portion, the foreign matter may block the transmitted light, thereby hindering the observation of the cells with a microscope or the like. In addition, the cells may be damaged by the contact between the foreign matter and the cells. Alternatively, substances eluted from foreign substances can affect cell growth. When the user discovers that a foreign substance is present inside the recess when using the cell handling container, the user either removes the foreign substance or discontinues the use of the recess containing the foreign substance and another depression. It is necessary to perform processing such as using a part. Such processing not only may reduce the work efficiency of the user, but may also cause mistakes.

In the cell handling container 1 of the present invention, the opening width r of the recessed portion and the maximum length p of the foreign matter satisfy r ≧ p, and the number n of the recessed portion and the number m of the foreign matter are n-1 ≧ m. It is necessary to meet. That is, in the cell handling container 1 of the present invention, there is no foreign substance whose maximum length p exceeds the opening width r of the recessed portion. Further, the number m of the foreign matter having the maximum length p is never more than the number n of the recessed portions. FIG. 5 is a schematic vertical cross-sectional view taken along II-II'in FIG. 1, showing a case where a foreign substance 100 has invaded the inside of the recess 20 of the cell handling container 1 of the present invention. As shown in FIG. 5, when the above requirements are satisfied, the maximum length p of the foreign matter 100 is equal to or less than the opening width r of the recess 20. Therefore, m foreign substances 100 may invade the inside of n recesses 20. However, the number m of the foreign matter is one less than or less than the number n of the recess 20. Therefore, even if the foreign matter 100 invades the inside of the recess 20, the foreign matter 100 does not exist inside at least one of the recesses 20. In the recess 20 where the foreign matter 100 does not exist, the cells can be handled without being affected by the foreign matter 100. Therefore, by providing the above-mentioned features, the cell handling container 1 of the present invention can handle cells by substantially preventing problems caused by the foreign substance 100.

In the present invention, the "foreign substance" means an object other than a member constituting the shape of the cell handling container itself of the present invention, and dust, dust, fibers mixed during the manufacture and / or packaging of the cell handling container of the present invention. Pieces, plastic pieces, metal pieces, and all or part of an organism can be mentioned. The foreign matter 100 is usually dust such as dust, fiber pieces or plastic pieces. For example, when the cell handling container 1 of the present invention is made of a material containing a plastic material and is manufactured by a conventional molding method such as injection molding, it occurs due to friction when the molded product is removed from the mold. A plastic piece, a burr of a molded product, a cut portion of a gate, or the like may be mixed as a foreign substance 100. The shape and dimensions of the foreign matter 100 are not particularly limited as long as the above and the following requirements are satisfied. For example, as shown in FIG. 5, the foreign matter 100 is spherical (FIG. 5 (a)), needle-shaped (FIG. 5 (b)), plate-shaped (FIG. 5 (c)), or irregularly shaped (FIG. 5 (d)). )) Can have various shapes. For example, when the foreign matter 100 is spherical, the maximum length p corresponds to the major axis diameter. When the foreign matter 100 is needle-shaped, the maximum length p corresponds to the major axis length. When the foreign matter 100 is plate-shaped, the maximum length p corresponds to the diagonal length of the plate-shaped surface. When the foreign matter 100 has an indefinite shape, the maximum length p corresponds to the major axis length. By having the above-mentioned characteristics, the cell handling container 1 of the present invention can handle cells by substantially preventing problems caused by the foreign matter 100 regardless of the shape of the foreign matter 100.

The opening width r of the recess 20 and the maximum length p of the foreign matter 100 preferably satisfy 0.95 × r ≧ p, and more preferably 0.8 × r ≧ p. Further, it is preferable that the opening width r of the recess 20 and the maximum length p of the foreign matter 100 satisfy r / 2 ≦ p. If the maximum length p of the foreign matter 100 is 95% or less of the opening width r of the recess 20, even if the foreign matter 100 enters the inside of the recess 20, use an instrument such as tweezers, a pipette, or a glass capillary. The foreign matter 100 can be easily removed. If the maximum length p of the foreign matter 100 is less than 1/2 of the opening width r of the recess 20, two or more foreign matter 100 may enter the inside of one recess 20. In such a case, not only the problem caused by the foreign matter 100 becomes more prominent, but also it may be difficult to remove the foreign matter 100 using an instrument. By setting the opening width r of the recess 20 and the maximum length p of the foreign matter 100 within the above ranges, even if the foreign matter 100 enters the inside of the recess 20, the foreign matter 100 can be easily removed by using an instrument. be able to.

The depth d of the recess 20 and the maximum length p of the foreign matter 100 preferably satisfy 1.1 × d ≧ p, more preferably d ≧ p, and even more preferably 0.8 × d ≧ p. Further, it is preferable that the depth d of the recess 20 and the maximum length p of the foreign matter 100 satisfy 0.5 × d ≦ p. For example, as shown in FIG. 5 (e), when the maximum length p of the foreign matter 100 exceeds 1.1 times the depth d of the recess 20, a part of the foreign matter 100 protrudes greatly from the opening of the recess 20. there is a possibility. In such a case, not only the problem caused by the foreign matter 100 becomes more prominent, but also it may be difficult to remove the foreign matter 100 using an instrument. Further, when the maximum length p of the foreign matter 100 is less than 0.5 times the depth d of the recessed portion 20, it may be difficult to remove the foreign matter 100 that has entered the inside of the recessed portion 20. By setting the depth d of the recess 20 and the maximum length p of the foreign matter 100 within the above ranges, even if the foreign matter 100 enters the inside of the recess 20, the foreign matter 100 can be easily removed by using an instrument. be able to. In addition, the cells can be handled by substantially preventing problems caused by the foreign matter 100.

For example, in the case of the cell handling container 1 in which the opening width of the opening of the side wall portion 12 of the container is about 35 mm and the bottom portion 11 of the container has a circular shape in the top view in the arrangement at the time of use, the number n of the recessed portions 20. Is preferably 50 or less, more preferably 30 or less, still more preferably 25 or less; the opening width r is preferably in the range of 100 to 1000 μm, 250 to 500 μm. The maximum length p of the foreign matter 100 is preferably 1000 μm or less, more preferably 950 μm or less, further preferably 800 μm or less; the number of foreign matter 100. The number of m is preferably 24 or less, more preferably 5 or less, and further preferably 1 or less. When the number m of the foreign matter 100 is within the above range, the foreign matter 100 does not exist inside at least one of the recessed portions 20. Therefore, the cells can be handled without being affected by the foreign matter 100 in the recessed portion 20 in which the foreign matter 100 does not exist. In the case of the above embodiment, the maximum length p of the foreign matter 100 can be, for example, in the range of 50 to 950 μm, particularly in the range of 200 to 400 μm. When the maximum length p of the foreign matter 100 is within the above range, the foreign matter 100 can be easily removed by using an instrument even when the foreign matter 100 invades the inside of the recess 20. Alternatively, the maximum length p of the foreign matter 100 can be, for example, in the range of 1 to 50 μm. If the maximum length p of the foreign matter 100 is less than 1 μm, it may be difficult to visually recognize the foreign matter 100 using a microscope, and / or it may be difficult to remove the foreign matter 100 that has entered the recess 20. There is a possibility of becoming. Therefore, when the maximum length p of the foreign matter 100 is within the above range, the foreign matter 100 can be visually recognized and removed by using a microscope. The cell handling container 1 of the present invention that satisfies the above requirements can handle cells by substantially preventing problems caused by the foreign substance 100.

In one embodiment, in the cell handling container 1 of the present invention, a storage compartment 30 is arranged at the bottom 11 of the container, and a recess 20 is arranged in the internal space of the storage compartment 30. The number and position of the storage compartments 30 at the bottom 11 of the container are not particularly limited, and any number, that is, one or more storage compartments 30 can be arranged at any position. For example, as shown in FIG. 6, in the figure formed by the peripheral edge of the inner surface of the container bottom 11 facing the space side where the cells and / or the culture medium are arranged in the arrangement at the time of use, when projected onto a horizontal plane. One storage compartment 30 can be arranged in the area corresponding to the center of gravity of the figure. Alternatively, in a figure formed by the peripheral edge of the inner surface of the bottom 11 of the container facing the space side where the cells and / or the culture medium are placed in the arrangement at the time of use projected onto a horizontal plane, the center of gravity of the figure is opposite. Two containment compartments can be placed in the area to be occupied. Alternatively, in the figure formed by the peripheral edge of the inner surface of the container bottom 11 facing the space side where the cells and / or the culture medium are placed in the arrangement at the time of use projected onto a horizontal plane, a polygon inscribed in the figure ( For example, a plurality of storage compartments can be arranged in the regions corresponding to the respective corners of a triangle, a quadrangle, a pentagon, a hexagon, etc.). In this case, in addition to the same region as described above, one or more storage compartments may be further arranged in other regions (for example, a region corresponding to the center of gravity of the figure).

FIG. 7 is a schematic vertical cross-sectional view taken along the line III-III'in FIG. In the present embodiment, the containment compartment 30 has a containment compartment bottom 31 and a containment compartment side wall 32 erected on the periphery of the containment compartment bottom 31. The storage compartment 30 is the surface of the storage compartment bottom 31 facing the space side where the cells and / or the culture medium are arranged (hereinafter, “the inner surface of the storage compartment bottom 31 facing the space side where the cells and / or the culture medium are arranged” or Also referred to as "inner surface of the bottom 31 of the containment compartment") and the surface of the side wall of the containment compartment 32 facing the space side where the cells and / or the culture medium are arranged (hereinafter, "the space side where the cells and / or the culture medium are arranged"). It is a bottomed shape that opens upward and is formed by "the inner surface of the side wall portion 32 of the accommodation compartment 32 facing toward" or "the inner surface of the side wall portion 32 of the accommodation compartment").

N recesses 20 are arranged at the bottom 31 of the containment compartment. In the case of the embodiment having a plurality of storage compartments, the recess 20 may be arranged at the bottom portion 31 of the storage compartment 30 of at least one storage compartment 30. In this case, the total number of recesses 20 arranged in the bottom 31 of the storage compartments 30 of the plurality of storage compartments 30 may be n. In the case of the present embodiment, no recess is arranged at the bottom of the accommodation compartment of the remaining accommodation compartment. That is, in the case of the present embodiment, the cell handling container 1 of the present invention has at least one storage compartment 30 having n recesses 20 and, in some cases, at least one other storage compartment 30 having no recesses 20. And can have. By arranging the recess 20 in the bottom 31 of the storage compartment, it is possible to substantially prevent the foreign matter 100 existing on the outside of the side wall 32 of the storage compartment from entering the inside of the recess 20. In the case of the present embodiment, the storage compartment 30 in which the recess 20 is arranged is used as the main compartment for culturing cells in the microdrop method, and another storage compartment in which the depression 20 is not arranged is used as the depression. Each cell can be used as a compartment for culturing cells taken out from the containment compartment 30 in which 20 is arranged in parallel while individually managing them. Therefore, the cell handling container of the present embodiment can be advantageously used for cell culture by the microdrop method.

The shape and dimensions of the containment compartment 30 are not particularly limited. As the shape of the storage compartment 30, for example, a figure formed by the peripheral edge of the storage compartment 30 when the inner surface of the bottom 31 of the storage compartment facing the space side where the cells and / or the culture medium are arranged is projected onto a horizontal plane in the arrangement at the time of use. , Circular (including perfect circle and oval), or polygonal shapes such as triangle, quadrangle, pentagon, hexagon or octagon. It is preferable that the figure formed by the peripheral edge of the inner surface of the bottom 31 of the containment compartment facing the space where the cells and / or the culture medium are arranged in use is circular when projected onto a horizontal plane. For example, in the case of the cell handling container 1 in which the opening width of the opening of the side wall portion 12 of the container is about 35 mm and the bottom portion 11 of the container has a circular shape in the top view in the arrangement at the time of use, the dimension of the storage compartment 30 is Although it depends on the opening width r, the number n, and the arrangement thereof of the recessed portions 20, it is preferable that the opening width of the opening of the side wall portion 32 of the accommodation section is in the range of 3 to 20 mm in the top view of the arrangement at the time of use. More preferably, it is in the range of 5 to 15 mm. For example, in the case of the accommodation compartment 30 in which n recesses 20 are arranged in the bottom portion 31 of the accommodation compartment, the dimensions of the accommodation compartment 30 are the opening of the opening of the side wall portion 32 of the accommodation compartment in the top view of the arrangement in use. The width is preferably in the range of 3 to 20 mm, more preferably in the range of 5 to 15 mm. For example, in the case of another containment compartment in which the recess is not arranged at the bottom of the containment compartment, the dimensions of the other containment compartment are such that the opening width of the opening of the side wall of the containment compartment is 1.5 to 1.5 when viewed from above in the arrangement in use. The range is preferably in the range of 15 mm, more preferably in the range of 3-7 mm. When the dimension of the accommodation compartment 30 is equal to or larger than the lower limit value, a sufficient space for performing the work can be secured inside the accommodation compartment 30. Further, when the dimension of the accommodation compartment 30 is equal to or less than the upper limit value, a sufficient space for performing the work can be secured even outside the accommodation compartment 30. When the containment compartment 30 has the above shape and the size of the containment compartment 30 is within the above range, a small volume usually used for cell culture by the microdrop method is inside the containment compartment 30 in which the recess 20 is arranged. By accommodating a drop of culture medium, eg 10-100 μL, a drop of culture medium, typically 0.35 mm or more, eg 0.5 mm or more in height, is formed inside the containment compartment 30 and said. The drop can cover substantially the entire surface of the bottom 31 of the containment compartment. Therefore, the cell handling container of the present invention in which the storage compartment has the above-mentioned shape and dimensions can be advantageously used for cell culture by the microdrop method.

The shape of the bottom 31 of the containment compartment is not particularly limited. The inner surface of the bottom 31 of the containment compartment facing the space side on which the cells and / or the culture medium are arranged may be formed so that substantially the entire surface is a horizontal plane in the arrangement during use, from the center to the outer edge in the arrangement during use. It may be formed so as to be an inclined surface that becomes higher in the vertical direction as it advances. Various shapes of a compartment for accommodating cells and a culture solution in a cell handling container such as a cell culture vessel known in the art can be applied to the shape of the bottom 31 of the containment compartment.

The cell handling container 1 of the present invention is usually used for observing the cells contained therein using a microscope. Here, when the surface roughness of the container bottom 11 of the cell handling container 1, the bottom 21 of the recess 20 and the bottom 31 of the storage compartment 31 is large, a clear image may not be obtained due to the unevenness of the surface. is there. Therefore, it is preferable that the surfaces of the container bottom 11, the bottom 21 of the recess 20 and the bottom 31 of the storage compartment are smooth surfaces having a surface roughness value as small as possible. Specifically, the surface of the bottom portion 11 of the container, the bottom portion 21 of the recessed portion 20, and the bottom portion 31 of the storage compartment preferably has a maximum height Ry of less than 1.0 μm, more preferably less than 0.5 μm. In the present invention, the maximum height Ry means the distance between the peak line and the valley bottom line in the extracted portion by extracting only the reference length from the roughness curve in the direction of the average line. The surface roughness of the surfaces of the bottom 11 of the container, the bottom 21 of the recess 20 and the bottom 31 of the storage compartment can be adjusted by polishing the mold of the cell handling container 1 of the present invention to improve the processing accuracy of the mold. It can be made smaller by increasing it. Further, the surface roughness can be measured by using a normal cross-sectional shape measuring device. As the measurement conditions, for example, a double-scan high-precision laser measuring instrument (Keyence, LT-9010M) and a high-precision shape measurement system (Keyence, KS-1100) can be used to measure at a measurement interval of 5 μm. it can.

The shape of the side wall portion 32 of the accommodation compartment is not particularly limited. The inner surface of the containment compartment side wall 32 facing the space side where the cells and / or the culture medium are arranged, and / or the surface of the containment compartment side wall 32 facing the outer space side of the containment compartment 30 (hereinafter, "outside of the containment compartment 30"). The "outer surface of the side wall portion 32 of the accommodation compartment 32 facing the space side" or "the outer surface of the side wall portion 32 of the accommodation compartment") may be formed so that substantially the entire surface is a vertical surface in the arrangement during use. , In the arrangement at the time of use, it is formed so as to become an inclined surface that is inclined toward the inside of the accommodation compartment side wall portion 32 with respect to the vertical surface as it advances from the connection portion with the accommodation compartment bottom portion 31 to the upper peripheral portion. May be good. Various shapes of the partition side wall portion for accommodating cells and the culture solution in a cell handling container such as a cell culture container known in the art can be applied to the shape of the compartment side wall portion 32. When the cell handling container 1 of the present invention is made of a material containing a plastic material and is manufactured by a conventional molding method such as injection molding, the inner surface and / or outer surface of the side wall portion 32 of the storage compartment 32 is formed. If it is formed so as to have an inclined surface as described above, it functions as a draft and can be efficiently released from the mold, which is advantageous.

In the present invention, the height of the side wall of the storage compartment is the vertical direction from the connection portion between the side wall of the storage compartment and the bottom of the storage compartment to the upper end of the side wall of the storage compartment in the schematic vertical cross-sectional view of the cell handling container of the present invention. Means the length of. Further, in the present invention, the thickness of the side wall portion of the storage compartment is the width of the side wall portion of the storage compartment in the vertical cross-sectional schematic view of the cell handling container of the present invention, that is, the accommodation facing the space side where the cells and / or the culture medium are arranged. It means the distance between the inner surface of the side wall of the compartment and the outer surface of the side wall of the containment compartment facing the outer space side of the containment compartment. For example, when substantially the entire inner surface and outer surface of the side wall portion of the accommodation compartment are formed to be vertical surfaces in the arrangement at the time of use, the width of the side wall portion of the accommodation compartment is constant in the schematic view of the vertical cross section. Therefore, the width of any position can be adopted. On the other hand, when the inner surface and / or the outer surface of the side wall portion of the accommodation compartment is formed to be an inclined surface, the maximum value of the width of the side wall portion of the accommodation compartment in the schematic vertical cross-sectional view may be adopted. The height of the side wall portion 32 of the storage compartment 32 can be appropriately set within a range in which the upper end of the side wall portion 32 of the storage compartment does not protrude from the upper end of the side wall portion 12 of the container. For example, if the height of the container side wall 12 is typically in the range of 5 to 20 mm, typically 7 to 15 mm, then the height of the containment compartment side wall 32 is in the range of 0.5 to 18 mm. Is preferable. If the height of the side wall portion 32 of the containment compartment 32 is less than the lower limit value, it may be difficult to put a sufficient amount of the culture solution inside the containment compartment 30. Further, when the height of the side wall portion 32 of the storage compartment exceeds the upper limit value, when the cell handling container 1 has a lid, the side wall portion 32 of the storage compartment and the lid come into contact with each other to close the opening of the side wall portion 12 of the container. May interfere. When the height of the side wall portion 32 of the containment compartment is within the above range, a drop of the culture solution is formed inside the containment compartment 30, and an amount of oil exceeding the height of the side wall portion 32 of the containment compartment 32 is put into the internal space. , The entire surface of the culture solution can be coated with oil to substantially suppress the evaporation of the culture solution. In the case of the above embodiment, the height of the side wall portion 32 of the containment compartment 32 is 2 to 6 mm in order to allow a sufficient amount of the culture solution to be put inside the containment compartment 30 and to further suppress the evaporation of the culture solution. It is more preferable that the range is. The thickness of the side wall portion 32 of the accommodation compartment is preferably in the range of 0.3 to 3 mm, more preferably in the range of 0.5 to 1.5 mm. If the thickness of the side wall portion 32 of the storage compartment exceeds the upper limit value, the space for working inside the storage compartment 30 may be narrowed. Further, when the thickness of the side wall portion 32 of the storage compartment is less than the lower limit value, the strength of the side wall portion 32 of the storage compartment may decrease and / or molding may become difficult. Therefore, when the thickness of the side wall portion 32 of the accommodation compartment 32 is within the above range, a sufficient space for working inside the accommodation compartment 30 is secured, and a sufficient strength of the side wall portion 32 of the accommodation compartment is secured. Can be done.

The structure of the side wall portion 32 of the accommodation compartment is not particularly limited. The interior of the containment compartment side wall 32 can usually be substantially completely filled with the materials that make up the containment compartment side wall. However, the accommodation compartment side wall 32 may optionally have a void within it. By having the void portion inside the side wall portion of the accommodation compartment, the material constituting the side wall portion of the accommodation compartment can be reduced according to the proportion of the void portion. Thereby, the manufacturing cost of the cell handling container of the present invention can be reduced.

The optical characteristics of the side wall portion 32 of the accommodation compartment 32 are not particularly limited. The cell handling container 1 of the present invention is usually used for observing the cells contained therein using a microscope. For this reason, the bottom 11 of the cell handling container 1, the bottom 21 of the recess 20, and the bottom 31 of the containment compartment usually have optical properties that do not substantially impede microscopic observation, such as substantially transparent and colorless optics. Has characteristics. However, the containment compartment side wall 32 may optionally have different optical properties, such as different transparency and color combinations than the containment compartment bottom 31. Since the side wall portion 32 of the containment compartment is not located on the optical axis connecting the cell to be observed and the objective lens, even if the optical characteristic of the side wall portion 32 of the containment compartment 32 is an optical characteristic that can hinder the observation by a microscope, the cell It does not have a substantial effect on the observation. Optical properties such as the transparency and color combination of the containment compartment side wall 32 can be imparted when or after the cell handling container 1 or containment compartment 30 of the present invention is made. For example, when the cell handling container 1 and the storage compartment 30 of the present invention are made of a material containing plastic and are integrally manufactured by injection molding, a masterbatch method, a dry blend method, a kneading method or a surface coating method. By adding an additive such as a brewing agent or various pigments in a transparent plastic molded product by using a method such as the above, a desired transparency and color combination can be imparted.

The surface characteristics of the side wall portion 32 of the containment compartment 32 are not particularly limited. In the cell handling container 1 of the present invention, the surfaces of the container bottom 11 of the cell handling container, the bottom 21 of the recess 20 and the bottom 31 of the storage compartment are smooth surfaces having a surface roughness value as small as possible. preferable. However, since the side wall portion 32 of the containment compartment is not located on the optical axis connecting the cell to be observed and the objective lens, even if the surface roughness of the side wall portion 32 of the containment compartment 32 is a value that can hinder the observation by the microscope. It does not have a substantial effect on cell observation. Therefore, the containment compartment side wall 32 may optionally have a different surface roughness than the containment compartment bottom 31. In the case of the present embodiment, the surface roughness of the side wall portion of the storage compartment can be appropriately set from the preferable range described above. The surface roughness of the side wall portion of the containment compartment can be adjusted by the method described above.

In the cell handling container 1 of the present invention, the storage compartment 30 may be formed integrally with the cell handling container 1 and may be formed as a member separate from the cell handling container 1. In the latter embodiment, the storage compartment 30 formed as a separate member may be formed of the same material as the cell handling container 1, or may be formed of a material different from that of the cell handling container 1. ..

In the embodiment in which the recess 20 is arranged at the bottom 31 of the storage compartment, the number n of the recesses 20 existing at the bottom 31 of the storage compartment and the number m'of the foreign matter existing at the bottom 31 of the storage compartment are n-1 ≧ m. 'It is preferable to satisfy. However, m'is an integer greater than or equal to 0 and less than or equal to m. In the case of the present embodiment, of the m foreign substances 100 existing on the bottom 11 of the container, m'foreign substances 100 are present in the internal space of the storage compartment 30, and m-m'foreign substances 100 are present in the storage compartment 30. It exists on the outside. The foreign matter 100 existing outside the containment compartment 30 is substantially prevented from entering the inside of the recess 20 by the containment compartment side wall portion 32. Therefore, by providing the above-mentioned characteristics, it is possible to reduce the foreign matter 100 that invades the inside of the recessed portion 20.

The cell handling container 1 of the present invention can have a lid if desired. By having the lid, it is possible to close the opening of the side wall portion 12 of the container so as to substantially prevent the invasion of foreign matter from the outside into the internal space 10 while ensuring the air permeability of the internal space 10.

The material of the cell handling container 1 of the present invention is not particularly limited. The materials of the cell handling container 1 and the storage compartment 30 include, for example, inorganic materials such as metal, glass and silicon, and plastics (for example, polystyrene resin, polyester resin, polyethylene resin, polypropylene resin, ABS resin, nylon, acrylic resin, etc. Fluorescent resin, polycarbonate resin, polyurethane resin, methylpentene resin, phenol resin, melamine resin, epoxy resin and vinyl chloride resin) can be mentioned. The material of the cell handling container 1 and the storage compartment 30 preferably contains one or more kinds of plastic materials exemplified above, and more preferably consists of one or more kinds of plastic materials exemplified above. The plastic material is desired by adding an additive such as a bluing agent or various pigments in a transparent plastic molded product by using a method such as a masterbatch method, a dry blending method, a kneading method or a surface coating method. A combination of transparency and color can be imparted. Therefore, since the cell handling container 1 and the storage compartment 30 of the present invention are made of the above-mentioned materials, a clear image can be obtained when observing the cells housed in the recess 20 using an optical microscope. Can be done.

The cell handling container 1 of the present invention may be surface-treated or surface-coated so as to promote the growth of fertilized eggs. For example, when co-culturing with cells of other organs (for example, endometrial cells or oviductal epithelial cells) in order to promote the development of fertilized eggs, these co-cultured cells are preliminarily placed in the cell handling container 1 Must be adhered to the surface of the. In such a case, the surface of the cell handling container 1, for example, the inner surface of the bottom 11 of the container, the inner surface of the bottom 21 of the recess 20 and / or the inner surface of the bottom 31 of the storage compartment, is coated with a cell adhesive material. Is advantageous. As the cell adhesion material, for example, the temperature-responsive polymer described in JP-A-2008-220320 and JP-A-2008-263863, etc., an environmental response capable of changing the cell adhesion under predetermined conditions. Sex polymers can be mentioned. By applying the above-mentioned surface treatment or surface coating, the culture efficiency in co-culture can be improved.

The type of cells to be cultured and / or observed using the cell handling container 1 of the present invention is not particularly limited. Examples of the cells include fertilized eggs, egg cells, ES cells (embryonic stem cells) and iPS cells (induced pluripotent stem cells). In the present invention, "egg cell" means an unfertilized egg cell, and includes an immature oocyte and a mature oocyte. After fertilization, the number of fertilized eggs increases to 2-cell stage, 4-cell stage, and 8-cell stage by cleavage, and develops into blastocysts via morula. The fertilized egg includes early embryos such as 2-cell embryos, 4-cell embryos and 8-cell embryos, mulberry embryos, and blastocysts (including early blastocysts, expanded blastocysts and prolapsed blastocysts). Blastocyst means an embryo consisting of an external cell capable of forming a placenta and an inner cell mass capable of forming an embryo. ES cells mean undifferentiated pluripotent or totipotent cells obtained from the inner cell mass of blastocysts. iPS cells mean cells that have been given pluripotency similar to ES cells by introducing several types of genes (transcription factors) into somatic cells (mainly fibroblasts). That is, the target cell in the cell handling container 1 of the present invention includes not only a single cell but also an aggregate of a plurality of cells such as a fertilized egg and a blastocyst. The cell handling container 1 of the present invention is suitable for culturing mammalian and avian cells, particularly mammalian cells. Mammals mean warm-blooded vertebrates, for example, primates such as humans and monkeys, rodents such as mice, rats and rabbits, pet animals such as dogs and cats, and domestic animals such as cows, horses and pigs. Can be mentioned.

The cell handling container 1 of the present invention can be advantageously used for cell culture of the cells by the microdrop method. An embodiment of the microdrop method using the cell handling container 1 of the present invention is shown in FIG. As shown in FIG. 8, usually, the culture solution M is added to the inner surface of the bottom portion 11 of the container, for example, in the case of the embodiment having the storage compartment 30, and then the culture liquid M is covered. Add oil O. Then, in the culture solution M, the cells A are added to the inside of the recess 20. These operations are usually performed using instruments such as pipettes or glass capillaries. Since the cell handling container 1 of the present invention has a large opening at the top, these operations can be performed relatively easily. In the cell handling container 1 of the present invention, there is no foreign substance having a maximum length p exceeding the opening width r of the recess 20 and the foreign substance 100 having a maximum length p equal to or less than the opening width r of the recess 20. The number m is one less than or less than the number n of the recess 20. Therefore, even if the foreign matter 100 invades the inside of the recess 20, the foreign matter 100 does not exist inside at least one of the recesses 20. In the recess 20 where the foreign matter 100 does not exist, the cells can be handled without being affected by the foreign matter 100. Therefore, the cell handling container 1 of the present invention can handle cells by substantially preventing problems caused by the foreign substance 100.

For example, when the cell culture of the cells is carried out using the cell handling container 1 of the present invention, the cell culture usually contains the gas necessary for the growth and maintenance of the cultured cells in the cell handling container 1 of the present invention. It is carried out by placing it in an incubator that provides an environmental atmosphere and a constant environmental temperature. In the case of the present embodiment, examples of the required gas include water vapor, free oxygen (O ₂ ) and carbon dioxide (CO ₂ ). By adjusting the environmental temperature and CO ₂ content within an appropriate range, the pH of the culture solution can be stabilized within a certain period of time. By comparing the image of the cells being cultured with the image stored in advance by the image comparison program, the culture conditions such as temperature, gas and medium at the time of culturing can be adjusted.

For example, when a fertilized egg is cultured using the cell handling container 1 of the present invention, it is usually determined after the culture whether or not the fertilized egg is of good quality suitable for transplantation into the uterus. In the case of the present embodiment, the discrimination may be performed automatically or manually using an optical microscope or the like. When performing automatic discrimination of cultured cells, an image of cells inside the recess 20 acquired by an optical microscope is imaged using a detection device such as a CCD camera, and the obtained image is subjected to contour extraction processing. The quality of cells (for example, fertilized eggs) can be determined by extracting a portion corresponding to a cell in an image and analyzing the image of the extracted cell with an image analyzer. As the image contour extraction process, for example, the means described in Japanese Patent Application Laid-Open No. 2006-337110 can be adopted.

As described in detail above, the cell handling container of the present invention can handle cells by substantially preventing problems caused by foreign substances. Therefore, the cell handling container of the present invention can be advantageously used for cell culture by the microdrop method.

<2: Manufacturing method of cell handling container>
The present invention also relates to a method for producing a cell handling container of the present invention. According to the method of the present invention, the cell handling container of the present invention having the characteristics described above can be produced. The method of the present invention needs to include a container forming step and a foreign matter removing step. Hereinafter, each step will be further described.

[2-1: Container forming process]
This step is a step of forming a cell handling container 1 having a container bottom portion 11 and a container side wall portion 12. By this step, it is possible to form a cell handling container 1 having a container bottom portion 11 and a container side wall portion 12, and n recessed portions 20 having an opening width r are arranged in the container bottom portion 11. Here, r and n have the same meanings as described above.

In the case of the embodiment in which the cell handling container 1 of the present invention has a storage compartment 30, in this step, a storage compartment 30 having a storage compartment bottom 31 and a storage compartment side wall 32 is arranged at the container bottom 11, and the storage compartment 30 is arranged. It is preferable to include forming a cell handling container 1 in which the recess 20 is arranged at the bottom 31 of the compartment.

This step can be carried out by using various methods known to those skilled in the art based on the material of the cell handling container 1 of the present invention. For example, when the material of the cell handling container 1 of the present invention contains one or more of the plastic materials exemplified above, this step can be carried out by using a conventional molding method such as injection molding.

[2-2: Foreign matter removal process]
This step is a step of reducing the foreign matter having the maximum length p arranged on the bottom portion 11 of the container by the foreign matter removing means. By this step, the number m of foreign matter having the maximum length p arranged on the bottom 11 of the container is reduced to n-1 or less, and the opening width r of the recess and the maximum length p of the foreign matter are r ≧ p. The cell handling container 1 of the present invention can be obtained, wherein the number n of the recesses and the number m of the foreign matter satisfy n-1 ≧ m. Here, p, r, m and n have the same meanings as described above.

In the case of the embodiment in which the cell handling container 1 of the present invention has a storage compartment 30, the step includes reducing the foreign matter 100 having the maximum length p arranged at the bottom 31 of the storage compartment by means of removing foreign matter. Is preferable. By carrying out this step having the above-mentioned characteristics, the number m'of 100 foreign substances having a maximum length p arranged at the bottom 31 of the storage compartment is reduced to n-1 or less, and the foreign matter exists at the bottom 31 of the storage compartment. The cell handling container 1 of the present invention can be obtained in which the number n of the recessed portion 20 and the number m'of the foreign matter 100 existing in the bottom portion 31 of the storage compartment satisfy n-1 ≧ m'. Here, p, r, m'and n have the same meanings as described above.

As the foreign matter removing means used in this step, a gas flow, a liquid flow or a transfer member is usually used.

The foreign matter removing means using the gas flow can be carried out by bringing the gas flow into contact with the inner surface of the bottom of the cell handling container obtained in the container forming step and, in some cases, the inner surface of the bottom of the storage compartment. Examples of the embodiment in which the gas flow is brought into contact include, for example, a treatment of spraying the gas flow on the inner surface of the bottom of the container of the cell handling container and, in some cases, the inner surface of the bottom of the storage compartment, and the inner surface of the bottom of the container of the cell handling container and the case. Therefore, a process of sucking a gas flow near the inner surface of the bottom of the storage compartment can be mentioned. The treatments may be carried out individually or in combination as desired, and may be carried out continuously, for example, alternately.

In the foreign matter removing means using a gas flow, examples of the gas forming the gas flow include air, nitrogen and carbon dioxide, and a mixed gas thereof. From the viewpoint of cost and safety at the time of residue, it is preferable to use air or nitrogen as the gas forming the gas flow.

In the foreign matter removing means using the gas flow, the wind speed, contact time, etc. of the gas flow can be appropriately set based on the shape and dimensions of the cell handling container, the shape and dimensions of the existing foreign matter, and the like. For example, in a cell handling container, the opening width of the opening of the side wall of the container is in the range of 30 to 60 mm and the height of the side wall of the container is in the range of 5 to 20 mm in the top view in the arrangement at the time of use. In the case of a circular petri dish, which has a circular shape at the bottom of the container and is usually used for culturing fertilized eggs in the cell culture field, the wind velocity of the gas flow near the inner surface of the bottom of the container and, in some cases, the inner surface of the bottom of the storage compartment. However, it is preferably 9 m / s or more, more preferably 12 m / s or more, preferably 15 m / s or less, and more preferably 13 m / s or less. The contact time of the gas flow is preferably 0.5 seconds or more, more preferably 1 second or more, preferably 2 seconds or less, and more preferably 1 second or less. When the wind speed and contact time are equal to or higher than the lower limit, foreign matter existing at the bottom of the container and, in some cases, at the bottom of the storage compartment can be within a predetermined range. Further, when the wind speed and the contact time are equal to or less than the upper limit value, it is possible to substantially prevent the cell handling container from moving or being damaged by the gas flow.

When the present step is carried out by a foreign matter removing means using a gas stream, the present step can further include removing the charge of the cell handling container by the charge removing means. In this case, the treatment by the foreign matter removing means may be carried out at any one or more time points before, during and after the treatment by the foreign matter removing means, and at all time points before, during and after the treatment by the foreign matter removing means. It may be carried out at. As the charge removing means, various means known to those skilled in the art such as an ionizer can be used based on the material of the cell handling container. By carrying out the treatment by the charge removing means in this step, the removal of foreign matters can be made more reliable.

The foreign matter removing means using the liquid flow can be carried out by bringing the liquid flow into contact with the inner surface of the bottom of the container of the cell handling container obtained in the container forming step and, in some cases, the inner surface of the bottom of the storage compartment. Examples of the embodiment in which the liquid flow is brought into contact include, for example, circulating the liquid flow on the inner surface of the bottom of the cell handling container and, in some cases, the inner surface of the bottom of the storage compartment.

In the foreign matter removing means using a liquid flow, examples of the liquid forming the liquid flow include water, methanol, ethanol and isopropyl alcohol, and a mixed liquid thereof. From the viewpoint of cost and safety at the time of residue, it is preferable to use water as the liquid forming the liquid flow.

In the foreign matter removing means using the liquid flow, the flow velocity and contact time of the liquid flow can be appropriately set based on the shape and size of the cell handling container, the shape and size of the existing foreign matter, and the like. For example, in the cell handling container, the opening width of the opening of the side wall of the container is in the range of 30 to 60 mm and the height of the side wall of the container is in the range of 5 to 20 mm in the top view in the arrangement at the time of use. In the case of a circular petri dish, which has a circular shape at the bottom of the container and is usually used for culturing fertilized eggs in the cell culture field, the flow velocity of the liquid flow near the inner surface of the bottom of the container and, in some cases, the inner surface of the bottom of the storage compartment. However, it is preferably 10 ml / min or more, more preferably 50 ml / min or more, further preferably 70 ml / min or more, and preferably 700 ml / min or less, preferably 600 ml / min or less. More preferably, it is ml / min or less. The contact time of the liquid flow is preferably 1 second or longer, more preferably 10 seconds or longer, preferably 60 seconds or shorter, and more preferably 30 seconds or shorter. When the flow velocity and contact time are equal to or higher than the lower limit, the foreign matter existing at the bottom of the container and, in some cases, at the bottom of the storage compartment can be within a predetermined range. Further, when the flow velocity and the contact time are equal to or less than the upper limit value, it is possible to substantially prevent the cell handling container from moving or being damaged by the liquid flow.

The foreign matter removing means using the transfer member can be carried out by bringing the transfer member into contact with the inner surface of the bottom of the container of the cell handling container obtained in the container forming step and, in some cases, the inner surface of the bottom of the storage compartment.

In the present invention, "transfer" means peeling an object from the surface of one member and attaching (that is, transferring) it to the surface of another member. In the foreign matter removing means using the transfer member, various known members having a transfer layer capable of transferring dust or the like can be used as the transfer member. Since it is used in contact with the inner surface of the bottom of the cell handling container and, in some cases, the inner surface of the bottom of the storage compartment, it is preferable to use a stick-shaped or sheet-shaped transfer member having an adhesive transfer layer.

In the foreign matter removing means using the transfer member, the contact pressure of the transfer member is determined based on the type of the transfer member used, the shape and size of the cell handling container, the shape and size of the existing foreign substance, and the like. It can be set as appropriate. For example, the transfer member used has a stick shape with an adhesive transfer layer, and the cell handling container has an opening width of 30 to 60 mm at the side wall of the container in a top view in the arrangement at the time of use. In the case of a circular petri dish that is usually used for culturing fertilized eggs in the cell culture field, in which the height of the side wall of the container is in the range of 5 to 20 mm and the bottom of the container is circular, translocation is performed. The contact pressure of the member ^{is preferably 1 kgf / cm 2} or more, more preferably 2 kgf / cm ² or more, and preferably 6 kgf / cm ² or less. When the contact pressure is equal to or higher than the lower limit value, foreign matter existing at the bottom of the container and, in some cases, at the bottom of the storage compartment can be within a predetermined range. Further, when the contact pressure is equal to or less than the upper limit value, it is possible to substantially avoid damage to the inner surface of the bottom of the container and, in some cases, the inner surface of the bottom of the storage compartment due to the contact of the transfer member.

When this step is carried out by various foreign matter removing means described above, it is preferable to fix the cell handling container by using a container fixing member having a recess into which the cell handling container can be fitted. By using the container fixing member, it is possible to substantially prevent the cell handling container from moving or being damaged by the treatment of the foreign matter removing means.

Hereinafter, the present invention will be described in more detail with reference to Examples. However, the technical scope of the present invention is not limited to these examples.

<I: Foreign matter removal effect by foreign matter removing means>
[Example I-1: Foreign matter removing means using gas flow]
A polystyrene circular petri dish usually used for culturing fertilized eggs in the cell culture field (opening width of the side wall of the container: 35 mm, height of the side wall of the container: 10 mm, opening width of the side wall of the storage compartment: 8 mm, height of the side wall of the containment compartment: 2 mm, opening width of the recess: 280 μm, depth of the recess: 160 μm, number of recesses: 25) were manufactured and prepared. This petri dish was allowed to stand with the lid open until foreign matter adhered to the inner surface of the bottom of the containment compartment. Fig. 9 shows a photograph showing the relationship between the recess and the foreign matter. FIG. 9 (a) shows the foreign matter existing on the inner surface of the bottom of the storage compartment and the opening of the recess, and FIG. 9 (b) shows the foreign matter having entered the inside of the recess. A petri dish with four foreign substances attached to the inner surface of the bottom of the containment compartment was selected. By observation under a microscope, it was confirmed that the maximum length of the adhered foreign matter was in the range of 30 to 120 μm.

The petri dish was allowed to stand on a laboratory table. From above the petri dish, use an air gun (TD-60-1R manufactured by Trusco Nakayama; nozzle length: 100 mm; caliber: 2 mm) to apply a gas stream of nitrogen (flow rate: 500 cm ³ / s) for 1 second. Sprayed on the inner surface of the bottom of the container and the bottom of the containment compartment. The same treatment was performed by changing the distance between the petri dish and the air gun. In addition, the wind speed near the inner surface of the bottom of the petri dish and the bottom of the storage compartment was measured in the treatment of each distance. After the treatment was completed, the number of foreign substances present on the inner surface of the bottom of each petri dish was counted. The relationship between the distance between the petri dish and the air gun and the wind speed of the gas flow is shown in Fig. 10, and the foreign matter existing on the inner surface of the bottom of the storage compartment of the petri dish before and after the treatment is shown in Fig. 11. The relationships are shown in Table 1, respectively.

Fig. 11 (a) shows the inner surface of the bottom of the petri dish before treatment and the foreign matter existing there, and (b) shows the inner surface of the bottom of the petri dish after treatment with a gas flow of 6.2 m / s. And the foreign matter present there, (c) is the inner surface of the bottom of the petri dish's storage compartment after treatment with a gas flow of 9.0 m / s and the foreign matter present there, (d) is 12.4 m / s. The inner surface of the bottom of the petri dish after being treated with a gas stream is shown, and the circles in the figure indicate foreign substances. As shown in Table 1, 3 foreign substances were removed when the wind speed of the nitrogen gas flow was 9.0 m / s or more, and all (that is, 4) foreign substances were removed when the wind speed was 12.4 m / s or more. In addition, the maximum length of foreign matter existing on the inner surface of the bottom of the petri dish before the start of treatment was in the range of 30 to 120 μm (measured maximum length: 30, 40, 50, 120 μm). , The maximum length of the foreign matter present after the treatment with a gas flow of 9.0 m / s was 40 μm (Figs. 11 (a), (b) and (c)).

[Example I-2: Foreign matter removing means using a liquid flow]
The same polystyrene circular petri dish used in Example I-1 was prepared. This petri dish was allowed to stand with the lid open until foreign matter adhered to the inner surface of the bottom of the containment compartment. A petri dish with 5 foreign substances attached to the inner surface of the bottom of the containment compartment was selected. The maximum length of the attached foreign matter was confirmed by observation under a microscope.

From above the petri dish, a liquid stream of tap water was circulated on the inner surface of the bottom of the petri dish container and the bottom of the storage compartment for 30 seconds. The same treatment was performed by changing the flow velocity of tap water. After the treatment was completed, the number of foreign substances present on the inner surface of the bottom of each petri dish was counted. Figure 12 shows the foreign matter present on the inner surface of the bottom of the petri dish's storage compartment before and after the treatment.

FIG. 12 (a) shows the inner surface of the bottom of the petri dish before treatment and foreign substances existing there, and (b) shows the inner surface of the bottom of the petri dish after treatment with a liquid flow of 70 ml / min. , (C) show the inner surface of the bottom of the petri dish's containment compartment after treatment with a liquid flow of 600 ml / min. As a result, all (that is, 5) foreign substances were removed regardless of whether the flow rate of tap water was 70 ml / min or 600 ml / min (FIGS. 12 (b) and (c)).

[Example I-3: Foreign matter removing means using a transfer member]
The same polystyrene circular petri dish used in Example I-1 was prepared. This petri dish was allowed to stand with the lid open until foreign matter adhered to the inner surface of the bottom of the containment compartment. A petri dish with three foreign substances attached to the inner surface of the bottom of the containment compartment was selected. By observation under a microscope, it was confirmed that the maximum length of the adhered foreign matter was in the range of 30 to 140 μm.

From above the petri dish, a cleaning stick (surface diameter of transfer layer: 5 mm; manufactured by Isshin Sangyo Co., Ltd.) was brought into contact with the inner surface of the bottom of the storage compartment of the petri dish. The contact pressure of the cleaning stick was measured using a commercially available pressure measuring film (prescale 4LW (for micro pressure) and LLLW (for ultra-low pressure); manufactured by FUJIFILM Corporation). After the treatment was completed, the number of foreign substances present on the inner surface of the bottom of each petri dish was counted. Figure 13 shows the foreign matter present on the inner surface of the bottom of the petri dish's storage compartment before and after the treatment.

Fig. 13 (a) shows the inner surface of the bottom of the petri dish's storage compartment before treatment and foreign matter existing there, and (b) shows the petri dish after contacting the cleaning stick with a contact pressure of ^{0.51 to 2.04 kgf / cm 2.} (C) shows the inner surface of the bottom of the petri dish and the foreign matter present therein, and (c) shows the inner surface of the bottom of the petri dish after contacting the cleaning stick with a contact pressure of ^{2.04 to 6.12 kgf / cm 2.} As a result, the treatment with a contact pressure of 0.51 to 2.04 kgf / cm ² did not remove foreign substances, but the ^{treatment with a contact pressure of 2.04 to 6.12 kgf / cm 2} removed all (that is, three) foreign substances. In addition, the maximum length of foreign matter present on the inner surface of the bottom of the petri dish's storage compartment before the start of treatment was in the range of 30 to 140 μm, but it was present after the treatment was completed with a contact pressure of ^{2.04 to 6.12 kgf / cm 2.} The foreign matter disappeared (Figs. 13 (a), (b) and (c)).

1… Cell handling container
10… Internal space of the container
11… Bottom of the container
12… Container side wall
20 ... dent
21 ... The bottom of the depression
22 ... Side wall of the recess
30 ... Containment compartment
31… Bottom of containment compartment
32 ... Side wall of containment compartment
100 ... Foreign matter
r… Opening width of the opening of the recess
d ... Depth of the depression
p… Maximum length of foreign matter
A ... cells
M ... Drop of culture solution
O ... Mineral oil

Claims (2)

A foreign matter removal method of the cells handling container,
The cell handling container has a container bottom and a container side wall, and n recesses having an opening width r are arranged at the container bottom, and a foreign substance which is a dust, a fiber piece, or a plastic piece is arranged at the container bottom. Have,
The method is a method of reducing the number m of foreign matter having a maximum length p at the bottom of the container to n-1 or less by a foreign matter removing means using a gas flow, a liquid flow or a transfer member.
However,
The number n of the recesses is an integer of 3 or more,
The number m of foreign matter is an integer of 1 or more,
The opening width r of the recess and the maximum length p of the foreign matter satisfy r ≧ p.
The above method, wherein the number n of the recesses and the number m of the foreign matter satisfy n-1 ≧ m. The bottom of the container of the cell handling containers, accommodating compartment bottom and the housing compartment is accommodated partition configuration having a side wall portion, the recessed portion in the housing compartment bottom that are located The method of claim 1.

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