JP6435201B2 - Well plate and object observation apparatus using the same - Google Patents

Description

  The present invention relates to a well plate for holding an object made of a solid or a liquid, and an object observation apparatus using the well plate.

  A well plate may be used for observing, selecting, inspecting, or culturing an object such as a cell or a cell aggregate. Generally, a well plate is composed of a flat plate main body and a plurality of wells provided independently on the plate main body (see, for example, Patent Document 1). Usually, a cell and its culture solution (medium) are put into each well by a chip capable of sucking and discharging cells. Furthermore, a reagent may be additionally supplied to the well containing cells by the chip for the purpose of reaction test or inspection.

  In order to control the temperature of the culture medium or the culture temperature of the cells, it may be necessary to adjust the temperature of the environment in which the well plate is placed. For example, when the basement membrane matrix Corning Matrigel (trade name of Corning) or BD Matrigel (trade name of Becton Dickinson) is used as the culture medium, the temperature of the culture medium may be 4 ° C. in certain tests. It is required to set the test environment. In addition, an observation image of the cells carried on the well plate is often captured by a camera. The imaging is often performed under transmitted illumination, with an illumination device disposed on the upper surface side of the well plate and a camera disposed on the lower surface side. For this reason, a plate made of a transparent resin member is generally used as the well plate.

JP 2006-109715 A

  In order to set the medium temperature, it is desirable to cool or heat the well plate directly. In this case, it is preferable to use a plate made of a material having excellent thermal conductivity from the viewpoint of accurately controlling the temperature. However, as a material having excellent heat conductivity, a material in which a base polymer is filled with a heat conductive filler is generally used, and the material is an opaque material. For this reason, if importance is attached to the thermal conductivity of the well plate, there arises a problem that it becomes difficult to capture an observation image by the camera.

  The present invention has been made in view of the above points, and a well plate capable of accurately adjusting a temperature while also clearly observing a supported object, and an object observation apparatus using the same. The purpose is to provide.

A well plate according to one aspect of the present invention is a well plate in which an object is inserted or removed by a chip that performs suction and discharge of the object, and is opposed to the first surface and the first surface. A plate body that is used with the first surface facing upward, an opening that is provided on the first surface and into which the object is placed, and a bottom that is located inside the plate body; A cylindrical wall surface extending downward from the opening toward the bottom, and one or a plurality of wells capable of holding the object, wherein the plate body is the first surface. An upper surface of the well, and an upper portion of the well is formed; a first member having good heat conductivity and non-transparency; a second upper surface; A second lower surface facing the second upper surface and serving as the second surface; A lower part including at least a part constituting the bottom of the well is formed, and a transparent second member, and the first lower surface of the first member and the second upper surface of the second member are: A well plate formed by joining the upper part and the lower part of the well in a state of being aligned , wherein the first member is a cylindrical part constituting a part excluding the vicinity of the lower end of the cylindrical wall surface And a first opening that is formed on the first upper surface and is an upper end edge of the cylindrical portion and serving as the opening, and a second opening that is formed on the first lower surface and serves as a lower end edge of the cylindrical portion, The second member includes a bottom portion and a lower end portion of the cylindrical wall surface, includes a recess having a third opening on the second upper surface, and the second opening and the third opening are aligned. In the state, the first member and the second member are joined .

  According to this well plate, one well plate is formed by joining the first member, which is the upper member, and the second member, which is the lower member, and the first member is a non-transparent member but has good heat conduction. The second member is a transparent member. Therefore, heat conduction to the object accommodated in the well is favorably performed by the first member. Further, since the transparency of the well is ensured by the second member, it is possible to improve the observability of the object accommodated in the well.

The first member includes a tubular portion which constitutes a portion excluding the vicinity of the lower end of the cylindrical wall, the first opening serving as the aperture a top edge of the formed on the first upper surface the cylindrical portion And a second opening formed on the first lower surface and serving as a lower end edge of the cylindrical portion, and the second member constitutes the lower portion and the vicinity of the lower end of the cylindrical wall surface, and is formed on the second upper surface. with a recess having a third opening, in a state in which said second opening and said third opening is aligned with said first member and said second member that are joined.

  According to this well plate, the transparent second member has the recess. Therefore, for example, when the object is a solid object, the solid object can be imaged with a camera or the like from the second surface side with the solid object in contact with the concave portion. In this case, the object can easily enter the range of the observation field of view as compared with a simple flat second member, so that the observability of the object can be improved.

  In this case, the second opening and the third opening may have the same horizontal cross-sectional size.

  According to this well plate, since the second opening and the third opening have the same size, a step does not occur on the inner surface of the well, and the object accommodation property can be improved.

  Alternatively, the horizontal opening size of the third opening may be larger than the horizontal cross-sectional size of the second opening.

  According to this well plate, it is possible to avoid the necessity of strictly performing the bonding accuracy between the first member and the second member, that is, the alignment accuracy between the second opening and the third opening. That is, even if the second opening is aligned with a slight misalignment with respect to the third opening, the entire area of the second opening can be opposed to the opening range of the third opening. Therefore, the joining work of the first member and the second member can be simplified.

  In the well plate, the first member includes a cylindrical portion constituting the cylindrical wall surface, a first opening formed on the first upper surface and serving as the opening at the upper edge of the cylindrical portion, A second opening formed on the first lower surface and serving as a lower end edge of the cylindrical portion, and the second upper surface of the second member may include a flat plate-like surface constituting the bottom portion. .

  According to this well plate, the second upper surface of the second member can be substantially flat. That is, a well plate that does not require any processing on the second member can be obtained, and the number of manufacturing steps can be reduced.

  In the well plate, the cylindrical wall surface is a predetermined region above the bottom, and has a first horizontal cross-sectional size capable of accommodating the object, and the upper part from the upper part of the accommodating part. It is desirable to include a receiving portion having a second horizontal cross-sectional size that is a region reaching the opening and is larger than the first horizontal cross-sectional size.

  According to this well plate, the volume of the well can be reduced without deteriorating the approachability of the chip to the well. Therefore, when an expensive liquid is accommodated in the well, the amount of use can be minimized.

  An observation apparatus for an object according to another aspect of the present invention is disposed on the second plate side of the well plate, a heat source that applies cold or heat to the first member of the plate body, and the plate body. An imaging device that captures an image of the object held in the well through the second member.

  According to this observation apparatus, it is possible to acquire an image of the object held in the well by the imaging apparatus in a state where the temperature of the well plate is adjusted by cold or warm heat input from a heat source.

  According to the present invention, a well plate capable of accurately adjusting the temperature of an object accommodated in a well while also clearly observing the accommodated object, and an object observation apparatus using the well plate are provided. Can be provided.

It is a top view of the well plate which concerns on 1st Embodiment of this invention. It is the II-II sectional view taken on the line of FIG. It is a figure which shows the structure of the observation apparatus of the target object which concerns on embodiment of this invention. It is a block diagram of the said observation apparatus. It is an expanded sectional view of the well plate. It is an exploded sectional view of the well plate. It is sectional drawing which shows the state in which the target object is thrown into the well with which the said well plate is provided, and the state where the target object is observed. It is sectional drawing which shows the well plate which concerns on 2nd Embodiment of this invention. It is sectional drawing which shows the well plate which concerns on 3rd Embodiment of this invention. (A)-(C) are sectional drawings which show the modification of a well.

  Hereinafter, embodiments of the present invention will be described in detail with reference to the drawings. FIG. 1 is a top view of a well plate 1 according to the first embodiment of the present invention, and FIG. 2 is a cross-sectional view taken along the line II-II in FIG. The well plate 1 is a plate capable of carrying an object made of a solid material, a liquid, or a mixture thereof. The operation of the object is provided with a tip opening, which is capable of sucking and holding the object through the tip opening, and discharging the held object from the tip opening. 6 (see FIG. 3) is used. With the chip 6 that performs such suction and discharge, the object is input to the well plate 1 or the input object is removed from the well plate 1.

  In the present invention, the object is not particularly limited, and various solids and liquids can be supported on the well plate 1. In this embodiment, a cell derived from a living body, particularly a cell aggregate is illustrated as a solid object. Further, as a liquid object, a cell aggregate medium (cell culture solution) such as Matrigel is exemplified. In this case, the well plate 1 is used for observing, selecting, inspecting, or culturing cells or cell aggregates. In addition, the object is a small electronic component or machine component, organic or inorganic crushed pieces or particles, solids such as pellets, liquids such as water, oil, biological fluids, solvents, reagents, or various solutions. Also good.

  The well plate 1 includes a plate body 2 and a plurality of wells 3 formed in the plate body 2. As illustrated in FIG. 1, here, a well plate 1 in which 4 × 6 = 24 wells 3 are arranged in a matrix is shown. It is sufficient that at least one well 3 is provided in the plate body 2. When the plate body 2 is provided with a plurality of wells 3, the arrangement mode may be a matrix arrangement of uniform arrangement or non-uniform arrangement, honeycomb form, or random arrangement.

  The plate body 2 is made of a flat plate member having a predetermined thickness, and includes an upper surface 21 (first surface) and a lower surface 22 (second surface facing the first surface). The well plate 1 is used with the upper surface 21 facing upward during normal use. The plate body 2 is composed of a first member 23 that is highly heat-conductive and non-transparent, and a second member 24 that is transparent, which are joined up and down. The first member 23 is a flat plate having a thickness that occupies most of the thickness of the plate body 2 in the vertical direction. The second member 24 is a thin flat plate having a thickness in the vertical direction of about ¼ to 1/15 compared to the first member, and is integrally joined to the lower surface of the first member 23.

  The well 3 is a bottomed hole provided in the plate body 2 and holds a solid object and / or a liquid object. The well 3 includes an opening 31, a bottom 32, and a cylindrical wall surface 33. The opening 31 is an opening into which an object is inserted. The opening 31 is a circular opening provided in the upper surface 21, and has an inner diameter that allows the chip to enter with a margin. The bottom 32 is located inside the plate body 2 and near the lower surface 22. The cylindrical wall surface 33 is a wall surface extending vertically downward from the opening 31 toward the bottom 32. The cylindrical wall surface 33 includes an accommodating portion 34 that occupies a region (predetermined region) that is about half of the upper and lower overall length of the well 3 upward from the bottom portion 32, and a receiving portion 35 that occupies a region that extends from the upper portion of the accommodating portion 34 to the opening 31. Including. The accommodating part 34 has the 1st horizontal cross-sectional size which can accommodate the solid object. The receiving part 35 has a second horizontal cross-sectional size that is larger than the first horizontal cross-sectional size. The shapes of the plate body 2 and the well 3 will be described in detail later with reference to FIGS.

  FIG. 3 is a diagram showing a configuration of an object observation apparatus S using the well plate 1 according to an embodiment of the present invention. Here, an observation apparatus S for observing the cell aggregate C held in the well plate 1 is illustrated. The object observation apparatus S includes a well plate 1 according to the present embodiment, a sorting container 4 for cell aggregates C, a camera unit 5 (imaging apparatus) including a camera lens 51, a head unit 61 on which a chip 6 is mounted, and A cooling unit 7 (heat source) for applying cold heat to the well plate 1 is provided.

  The sorting container 4 is a container for storing the cell culture solution 41 and holds a sorting plate 42 for cell sorting in a state of being immersed in the cell culture solution 41. The sorting plate 42 includes a plurality of holding recesses capable of individually supporting the cell aggregates C. The bottom surface of the holding recess is provided with an escape hole through which a cell aggregate C of a predetermined size cannot pass but a minute contaminant or the like can pass. A cell suspension containing a large number of cell aggregates C is introduced from above the sorting plate 42. The cell aggregate C having a predetermined size to be inspected is trapped in the holding recess, while impurities and the like fall from the escape hole to the bottom surface of the sorting container 4. Thereby, a state in which the cell aggregate C to be examined is carried on the upper surface of the sorting plate 42 is formed. FIG. 3 shows this state. If a small-diameter through hole reaching the lower surface 22 is provided on the bottom surface of the well 3, this can also be used as the sorting plate.

  The camera unit 5 captures an image of the cell aggregate C held in the sorting plate 42 in the sorting container 4 or the cell aggregate C held in the well 3 in the well plate 1. The camera unit 5 includes an image sensor such as a CCD image sensor. The camera lens 51 forms an optical image of the cell aggregate C on the light receiving surface of the image sensor. The camera unit 5 is disposed below the camera lens 51 so as to face the lower surfaces of the sorting container 4 and the well plate 1. That is, an image of the cell aggregate C carried on the sorting container 4 or the well plate 1 is taken from the lower surface side. The camera unit 5 can move in the horizontal direction between the lower side of the sorting container 4 and the lower side of the well plate 1 along the guide rail 52 as indicated by an arrow A2 in the drawing.

  The chip 6 is a tube-shaped member having a tip opening, and sucks and discharges the cell culture solution 41 including the cell aggregate C. Specifically, the chip 6 sucks the cell aggregate C together with the cell culture solution 41 from the sorting container 4 and discharges them to the well 3 of the well plate 1. Although not shown, the chip 6 aspirates a reagent solution or the like as needed and discharges it into the well 3 carrying the cell aggregate C. The chip 6 is attached to the lower end of the rod 6 </ b> R provided in the head unit 61. A piston mechanism is mounted on the rod 6R, and suction force and discharge force are applied to the tip opening of the chip 6 by the operation of the piston mechanism. Further, the rod 6R is movable in the vertical direction. The head unit 61 includes a power unit of the piston mechanism and the like, and is movable in the horizontal direction along the guide rail 62 as indicated by an arrow A1 in the drawing.

  The cooling unit 7 maintains the cell culture solution 41 held in the well 3 at a desired temperature by cooling the well plate 1 to a constant cooling temperature. The cooling unit 7 gives cold to the first member 23 having excellent thermal conductivity among the constituent members of the plate body 2. As the cooling unit 7, for example, a Peltier element can be preferably used. In this case, the heat absorption part of the Peltier element is brought into contact with a part of the first member 23. In addition, as the cooling unit 7, a metal block such as aluminum that has been cooled to a predetermined temperature in advance, a cold storage member such as a cooling agent, or an evaporator using a vaporization compression device and a vaporization absorption device can be applied.

  For example, Matrigel liquefies at 4 ° C. and is often used at a temperature of about 37 ° C. for cell culture. When the object observation device S is arranged in a room temperature environment and matrigel is accommodated in the well 3 as a culture medium, the cooling unit 7 applies cold heat to the first member 23 so that the well plate 1 can maintain 4 ° C. In addition, when the target object accommodated in the well 3 needs heating, it replaces with the cooling part 7 and a heating apparatus is used, and a 1st member 23 is given heat. In the case where a Peltier element is applied as the cooling unit 7, this heating can employ means that reverses the drive current applied to the Peltier element or uses the heat dissipation side of the Peltier element.

  FIG. 4 is a block diagram of the object observation apparatus S. The observation apparatus S includes a control unit 8 that controls operations of the camera unit 5, the head unit 61, the moving mechanism M, and the cooling unit 7. The moving mechanism M includes drive motors that move the camera unit 5 and the head unit 61 along the guide rails 52 and 62, respectively.

  The control unit 8 controls the camera unit 5 and the epi-illuminator 9 (see FIG. 7) when acquiring an image of the cell aggregate C carried on the sorting container 4 or the well plate 1, and under the epi-illumination. The imaging operation is executed. Further, the control unit 8 controls the power unit of the piston mechanism provided in the head unit 61, the vertical movement mechanism of the rod 6R, and the like to generate a suction force or a discharge force at the tip opening of the tip 6, and the rod The operation of moving the 6R up and down is controlled. Further, the control unit 8 operates the drive motor of the moving mechanism M to control the positions of the camera unit 5 and the head unit 61 on the guide rails 52 and 62. Further, the control unit 8 controls the cooling unit 7 to adjust the temperature of the well plate 1 (the temperature of the object in the well 3).

  FIG. 3 is also a diagram illustrating a process in which the cell aggregate C is transported from the sorting container 4 to the well plate 1. It is desirable to inject a predetermined amount of Matrigel into the well 3 of the well plate 1 before this transport. In this case, in order to prevent the matrigel from solidifying, the cooling of the well plate 1 by the cooling unit 7 is started in advance. With reference to FIGS. 3 and 4, first, the control unit 8 drives the moving mechanism M to move the head unit 61 over the sorting container 4 and the camera unit 5 below the sorting container 4. Next, the control unit 8 causes the camera unit 5 to perform an imaging operation, and acquires the position information of the cell aggregate C held in the holding recess of the sorting plate 42. Then, after positioning the chip 6 in one holding recess of the sorting plate 42, the control unit 8 operates the vertical movement mechanism of the head unit 61 to lower the rod 6R, so that the tip opening of the chip 6 becomes the holding recess. It is made to approach the cell aggregate C carried. Subsequently, the control unit 8 operates the piston mechanism to suck the cell aggregate C together with the surrounding cell culture solution 41 into the chip 6. Thereafter, the rod 6R is raised to the original height position. In FIG. 3, a head unit 61 indicated by a solid line shows a state in which the ascent is finished.

  Next, the cell aggregate C is moved to the well plate 1. The control unit 8 moves the head unit 61 along the guide rail 62 and stops it above the well plate 1 as indicated by a dotted line in FIG. At this time, the chip 6 holding the cell aggregate C and the one well 3 that is the input target of the cell aggregate C are aligned. The camera unit 5 is also moved below the well plate 1. Subsequently, the control unit 8 lowers the rod 6 </ b> R and causes the tip opening of the chip 6 to enter the well 3. And the control part 8 operates the said piston mechanism, and discharges the cell culture solution 41 containing the cell aggregate C from the front-end | tip opening of the chip | tip 6 in the well 3. FIG.

  The control unit 8 operates the cooling unit 7 as necessary to apply cold heat to the well plate 1 to adjust the temperature of the cell culture solution 41 and the cell aggregate C object in the well 3. In addition, the control unit 8 operates the camera unit 5 and the epi-illuminator 9 (see FIG. 7) to capture an observation image of the cell aggregate C accommodated in the well 3.

  Subsequently, the detailed structure of the well 3 provided in the well plate 1 of the first embodiment will be described in detail with reference to FIGS. 5 and 6. FIG. 5 is an enlarged sectional view of the well plate 1, and FIG. 6 is an exploded sectional view of the well plate 1. As described above, the plate body 2 includes a joined body of the non-transparent first member 23 and the transparent second member 24. The well 3 formed in the plate main body 2 has a circular shape in a horizontal cross section at any height position, and includes an opening 31, a bottom 32, and a cylindrical wall 33, and the cylindrical wall 33 includes an accommodating portion. 34 and the receiving part 35 are included. As shown in FIG. 1, the opening 31 and the bottom 32 have a circular shape in a top view, and the two are in a concentric positional relationship.

  The first member 23 has a first upper surface 23U that becomes the upper surface 21 of the plate body 2 and a first lower surface 23L that faces the first upper surface 23U, and an upper portion excluding the vicinity of the bottom 32 of the well 3 is formed. Yes. The first upper surface 23U and the first lower surface 23L are both flat. Specifically, the first member 23 includes a cylindrical portion 233 constituting a portion excluding the vicinity of the lower end of the cylindrical wall surface 33, a first opening 231 formed on the first upper surface 23U and serving as an upper end edge of the cylindrical portion 233, 1 includes a second opening 232 that is formed on the lower surface 23 </ b> L and serves as a lower edge of the cylindrical portion 233. The first opening 231 is an opening that becomes the opening 31 of the plate body 2. The cylindrical portion 233 is a portion that forms most of the cylindrical wall surface 33, and constitutes the majority of the receiving portion 35 except for the vicinity of the lower end of the accommodating portion 34.

  The second member 24 has a second upper surface 24U and a second lower surface 24L that is opposed to the second upper surface and serves as the lower surface 22 of the plate body 2, and a lower portion including a portion constituting the bottom 32 of the well 3 is formed. Has been. The second upper surface 24U and the second lower surface 24L are both flat. Specifically, the second member 24 includes a recess 241 that forms the bottom 32 and the vicinity of the lower end of the cylindrical wall surface 33. The recess 241 has a third opening 242 that opens to the second upper surface 24U, and includes a curved surface that is recessed downward. The vicinity of the bottom of the concave curved surface is the bottom 32 of the well 3, and the vicinity of the upper edge is the lower end of the cylindrical wall surface 33 (accommodating portion 34). You may make it provide the small hole penetrated from the deepest part of the recessed part 241 to the lower surface 22 as needed. The third opening 242 and the second opening 232 of the first member 23 are openings having the same diameter (the same horizontal cross-sectional size).

  The first member 23 and the second member 24 are stacked one above the other so that the first lower surface 23L and the second upper surface 24U are joined. At the time of this bonding, the upper portion and the lower portion of the well 3 are aligned, that is, the second opening 232 and the third opening 242 are aligned. By such joining, one plate body 2 composed of the first member 23 and the second member 24 is formed, and the well 3 communicated with the plate body 2 from the opening 31 to the bottom 32 in one space. It is formed. In the present embodiment, since the second opening 232 and the third opening 242 are circular openings having the same diameter, a step can be prevented from occurring on the inner surface of the well 3 if the two are accurately aligned. Thereby, the accommodating property of the cell aggregate C in the accommodating part 34 can be made favorable. Note that it is desirable to provide a member for positioning the first member 23 and the second member 24 so that the above alignment can be performed efficiently. For example, it is desirable to project a positioning rib that engages with a part of the first lower surface 23 </ b> L of the first member 23 on the second upper surface 24 </ b> U of the second member 24.

  The material of the plate body 2 is not particularly limited, and is a material capable of forming the well 3 by drilling or molding, and is chemically applied to the object to be carried (cell aggregate C, cell culture solution 41, etc.). Any material that does not affect the environment is acceptable. For example, the plate body 2 made of a resin molded product is one of the preferred examples. However, the first member 23 is required to have good thermal conductivity, and the second member 24 is required to be transparent.

  As a material for forming the first member 23, for example, a resin composition formed by filling a base resin with an appropriate amount of a heat conductive filler can be used. The base resin is not particularly limited. For example, epoxy resin, polypropylene resin, polycarbonate resin, acrylic resin, polyester resin, nylon, PPS (polyphenylene sulfide), PBT (polybutylene terephthalate resin), polyamide resin, PPS (polyphenylene) Examples thereof include sulfide), PET (polyethylene terephthalate), POM (polyoxymethylene), CA (cellulose acetate), CAB (cellulose acetate butyrate), phenol resin, phenol resin, polyurethane resin and the like. Moreover, an inorganic filler can be illustrated as a heat conductive filler. Inorganic fillers include aluminum oxide, aluminum nitride, magnesium oxide, aluminum hydroxide, magnesium hydroxide, boron nitride, copper, gold, silver, titanium, SUS, iron, graphite, silicon, silicon carbide, boron nitride, silicon nitride, etc. Can be illustrated.

  The material forming the second member 24 is not particularly limited as long as it is a resin having transparency, and examples thereof include methacrylic resin, polycarbonate resin, polystyrene resin, acrylic resin, and polyurethane resin.

  The inorganic filler-containing resin composition as described above is generally non-transparent. For this reason, the 1st member 23 is inferior in transparency, and it is difficult to observe the cell aggregate C hold | maintained at the well 3 from the side surface direction (horizontal direction). However, since the first member 23 is a member having excellent thermal conductivity, for example, when cold heat is applied from the side periphery of the first member 23 by the cooling unit 7 (see FIG. 3), the cold heat is transferred to each well 3. Good heat transfer. Therefore, the well plate 1 of the present embodiment is useful for maintaining the low temperature of the cell culture solution 41 held in the well 3 (for example, 4 ° C.) and for the purpose of floating or embedding the cell aggregate C in Matrigel.

  The first member 23 is a non-transparent member, but the second member 24 is a transparent member. Therefore, it is possible to observe the cell aggregate C accommodated in the accommodating portion 34 of the well 3 through the second member 24 from the lower surface 22 side of the well plate 1. When viewed from the lower surface 22 side of the well plate 1, the first lower surface 23 </ b> L of the first member 23 is visually recognized through the transparent second member 24. In this case, when the illumination light is applied to the upper surface 21 side of the well plate 1, the cylindrical portion 233 is a hole penetrating the first member 23 up and down, so the second opening 232 becomes a bright region through which the illumination light passes. The part becomes a dark region. When the cell aggregate C is accommodated in the accommodating part 34 of the well 3, the image can be visually recognized. Therefore, by arranging the camera lens 51 immediately below the bottom 32 of the well 3, an image of the cell aggregate C can be acquired.

  That is, according to the well plate 1 of the present embodiment, the heat conduction to the well 3 is favorably performed by the first member 23, and the transparency of the well 3 is ensured by the second member 24. Both the heat controllability and the observability of the cell aggregate C thus obtained can be improved. Further, the transparent second member 24 has a recess 241. Accordingly, the cell aggregate C can be imaged from the lower surface 22 side in a state where the cell aggregate C is brought into contact with the recess 241. Since the concave portion 241 has a concave curved surface shape, when the cell aggregate C is in contact, the cell aggregate C is stabilized and the cell aggregate C enters the range of the observation field. There is an advantage that the range in which the image can be directly captured becomes wider.

  There is no restriction | limiting in particular about the method of joining the 1st member 23 and the 2nd member 24. FIG. It is one of preferable methods to apply an adhesive to the first lower surface 23L of the first member 23 or the second upper surface 24U of the second member 24 and to bond them together. In this case, a jig that can separate and approach the first member 23 and the second member 24 in the vertical direction in a state where the second opening 232 and the third opening 242 are aligned is used. In a state where the first member 23 and the second member 24 are separated from each other by the jig, the adhesive is applied. After that, the two are brought close to each other so that the first lower surface 23L and the second upper surface 24U By making the contact surface, both can be joined. In addition, the first lower surface 23L and the second upper surface 24U are each formed with a fitting portion that engages with each other, and a method of joining the two by fitting them together, or a method of mechanically coupling the two by screwing or the like. Can be illustrated.

  Next, the shape of the well 3 will be described in detail. The opening 31 of the well 3 is connected to the upper end of the cylindrical wall surface 33 (receiving portion 35), and the bottom 32 is connected to the lower end of the cylindrical wall surface 33 (accommodating portion 34). The inner diameter of the opening 31 is set to a diameter sufficient to receive the tip of the chip 6. The cylindrical wall surface 33 has a circular cross section at any height. The accommodating portion 34 of the cylindrical wall surface 33 is a wall surface having a predetermined first inner diameter over the entire height, and defines a cylindrical space. That is, the horizontal cross-sectional size of the accommodating portion 34 is not substantially changed in the vertical direction. The first inner diameter is selected to a size that does not cause the cell agglomerates C that have entered the accommodating portion 34 to swing in the horizontal direction.

  The receiving part 35 is generally a wall surface having a second inner diameter that is larger than the first inner diameter. In this embodiment, the taper-shaped wall surface in which the horizontal cross-sectional size of the receiving part 35 changes continuously in the up-down direction is illustrated. The inner diameter (horizontal cross-sectional size) of the receiving portion 35 has the largest inner diameter at the upper end portion connected to the opening portion 31 and the smallest inner diameter at the lower end portion serving as a boundary portion with the accommodating portion 34, and the inner diameter between them. Is changing continuously. A frustoconical space is defined by such a receiving portion 35.

  According to the well 3 having such a shape feature, it is possible to reduce the internal volume of the well 3 while maintaining good approachability when the chip 6 (see FIG. 3) enters the well 3. In FIG. 5, a rectangular shape R indicated by a dotted line is a vertical sectional shape of the well when the well extends to the position of the bottom 32 with the same diameter as the inner diameter of the opening 31. Compared with the rectangular shape R, the horizontal cross-sectional size of the well 3 of this embodiment is suppressed. In general, a shape such as the rectangular shape R can be said to be a shape that can simplify the shape of a mold used when producing the well plate 1 (first member 23). However, the well 3 according to the present embodiment includes the accommodating portion 34 whose horizontal cross-sectional size is intentionally suppressed according to the size of the cell aggregate C, and the truncated cone-shaped receiving portion 35 whose maximum size portion is the opening 31. A cylindrical wall surface 33 is formed. As a result, the internal volume of the well 3 is suppressed. For example, when it is necessary to put an expensive reagent solution into the well 3, it is possible to save the amount of the reagent solution used, thereby reducing the cost of testing and inspection. To contribute.

  Even if the internal volume of the well 3 is suppressed, the approach of the chip 6 to the well 3 is not impaired because the vicinity of the opening 31 has the maximum size. That is, the opening 31 has the same size as that of the rectangular shape R, and even if the chip 6 is slightly misaligned and enters the well 3, it can be received. In addition, since the receiving part 35 has a tapered shape that decreases in diameter as it goes downward, when a target object such as a cell aggregate C, a cell culture solution, or a reagent solution is put into any part of the receiving part 35 However, it is possible to smoothly guide the object to the lower accommodating portion 34 using gravity. And the cell clump C which entered the accommodating part 34 as above-mentioned is hold | maintained favorably. If the above cost requirement is not particularly considered, the well 3 having a right cylindrical hole shape such as the rectangular shape R may be used.

  FIG. 7 is a vertical cross-sectional view showing a state where the cell aggregate C is put into the well 3 from the chip 6. As described above with reference to FIG. 3, when the cell aggregate C is put into the well 3, the chip 6 holding the cell aggregate C and the cell culture solution 41 is lowered from above toward the opening 31 of the well 3. The Then, the lowering of the chip 6 is stopped at a position where the tip opening 6T of the chip 6 enters the receiving portion 35 by a predetermined depth. Thereafter, the cell aggregate C and the cell culture solution 41 are discharged from the tip opening 6T. The left well 3 in FIG. 7 shows a state immediately before the ejection is performed. Note that a predetermined amount of cell culture solution may be poured into the well 3 in advance before the discharge. On the other hand, the center and right wells 3 in FIG. 7 show a state in which the ejection is completed. In these wells 3, the cell culture solution 41 fills the entire storage part 34 of the well 3 and about the lower half of the receiving part 35, and the cell aggregate C is confined on the side by the storage part 34. Therefore, it is in contact with or floats near the bottom 32. The amount of the cell culture solution 41 is appropriately adjusted. In the embodiment in which the cell culture solution is poured into the well 3 in advance, the liquid of the cell culture solution to be held in the well 3 in advance in consideration of the amount of the cell culture solution held in the chip when the cell aggregate C is aspirated. The amount is determined.

  Note that the lowering of the tip 6 may be stopped at a position where the tip opening 6T does not enter the receiving portion 35. That is, the opening 31 only needs to have an opening of a size that allows the tip opening 6T of the chip 6 to enter, and when the cell aggregate C is introduced, the tip opening 6T actually exceeds the opening 31 and the well 3 It is not necessary to enter inside. For example, the tip opening 6T may be stopped about several mm above the position of the opening 31, and the cell aggregate C and the cell culture solution 41 may be discharged from the tip opening 6T at that position. In this case, if the tip opening 6T is not directly above the bottom portion 32 but is positioned above the opening portion 31, the discharged cell aggregate C and the cell culture solution 41 can be transferred to the tapered wall surface of the receiving portion 35. Can be guided to the housing part 34.

  FIG. 7 schematically illustrates a state in which the camera unit 5 and the epi-illuminator 9 are positioned with respect to the right well 3. The epi-illuminator 9 is arranged above the well plate 1 and irradiates the illumination light L from directly above the well 3 carrying the cell aggregate C to be imaged. The camera unit 5 is disposed below the well plate 1 and the optical axis and focus of the camera lens 51 are aligned with the cell aggregate C to be imaged. In this state, the imaging operation of the cell aggregate C is executed. The imaging operation of such an embodiment cannot be performed when the entire well plate 1 is formed of a highly heat conductive material (inorganic filler-containing resin composition) and is an opaque member. However, in the well plate 1 of the present embodiment, the visibility of the well 3 from below is ensured by the second member 24, so that the image capturing operation of the cell aggregate C held in the well 3 through the second member 24 is performed. Can be executed without problems.

  FIG. 8 is a sectional view showing a well plate 1A according to the second embodiment of the present invention. The second embodiment exemplifies a well plate 1A having a second member 24A made of a flat plate that does not have the recess 241 as in the first embodiment.

  The well plate 1A includes a plate body 2A and a plurality of wells 3A formed in the plate body 2A. The plate body 2 </ b> A includes a joined body of a first member 23 that is exactly the same as that of the first embodiment and a second member 24 </ b> A that does not include the recess 241. The well 3 </ b> A includes an opening 31, a bottom 32 </ b> A, and a cylindrical wall surface 33, and the cylindrical wall surface 33 includes a storage portion 34 and a receiving portion 35. The difference from the first embodiment is that the bottom 32A is a flat horizontal surface. In the second embodiment, the cylindrical portion 233 of the first member 23 constitutes the entire length of the receiving portion 35 as well as the entire length of the receiving portion 35.

  The second upper surface 24AU of the second member 24A is composed of a flat surface having no irregularities. Therefore, in a state where the second member 24A is joined to the first member 23, a part of the second upper surface 24AU facing the second opening 232 of the cylindrical portion 233 becomes the bottom portion 32A. That is, an arbitrary portion of the second upper surface 24AU can be the bottom 32A of each well 3A.

  According to the well plate 1A of the second embodiment, the second upper surface 24AU of the second member 24A can be substantially flat. For this reason, a simple flat plate material that is not subjected to any processing can be used as the second member 24A. In addition, as in the first embodiment, the alignment between the concave portion 241 and the second opening 232 is not necessary, so that the joining operation of the first member 23 and the second member 24A can be simplified. As a result, the number of manufacturing steps for the well plate 1A can be reduced. Furthermore, since the well 3A is entirely constituted by the first member 23 having substantially good thermal conductivity, it becomes possible to more accurately perform the heat control on the object accommodated in the well 3A.

  FIG. 9 is a sectional view showing a well plate 1B according to the third embodiment of the present invention. In the third embodiment, a well plate 1B including a second member 24B having a recess 241B different from the recess 241 of the first embodiment is illustrated.

  The well plate 1B includes a plate body 2B and a plurality of wells 3B formed in the plate body 2B. The plate body 2B is composed of a joined body of a first member 23 that is exactly the same as that of the first embodiment and a second member 24B having a recess 241B having a large opening diameter. The well 3 </ b> B includes an opening 31, a bottom 32 </ b> B, and a cylindrical wall surface 33, and the cylindrical wall surface 33 includes a storage portion 34 and a receiving portion 35. The concave portion 241B constitutes the bottom portion 32B and the vicinity of the lower end of the cylindrical wall surface 33. The recess 241B has a third opening 242B that opens to the second upper surface 24BU, and has a curved surface that is recessed downward.

  The third opening 242B and the second opening 232 of the first member 23 are circular openings. The inner diameter (horizontal cross-sectional size) of the third opening 242B is set to be about 5% to 50% larger than the inner diameter of the second opening 232. In the first embodiment, the third opening 242 </ b> B and the second opening 232 have the same diameter, whereas in the present embodiment, the third opening 242 </ b> B has a larger diameter than the second opening 232.

  According to the well plate 1B of the third embodiment, it is possible to avoid the necessity of strictly performing the bonding accuracy between the first member 23 and the second member 24B, that is, the alignment accuracy between the second opening 232 and the third opening 242B. be able to. That is, even if the second opening 232 is aligned with a slight misalignment with respect to the third opening 242B, the entire region of the second opening 232 can be made to face the opening range of the third opening 242B. Therefore, the joining work of the first member 23 and the second member 24B can be simplified. Moreover, since the 3rd opening 242B has a concave-curved shape, when the cell aggregate C contact | connects similarly to 1st Embodiment, while the said cell aggregate C is stabilized, while the cell aggregate C is stabilized, it is a cell aggregate C with a perspective line. There is an advantage that the range in which the outer surface of the image can be directly imaged is widened.

  In addition, various shapes can be adopted as the shape of the well. In the above-described embodiment, the wells 3, 3A, and 3B including the receiving portion 35 having a tapered wall surface are illustrated. The well may not include a portion such as a tapered portion. FIG. 10 shows another example of the well. In the well 3C shown in FIG. 10A, the cylindrical wall surface portion formed in the first member 23 defines a cylindrical space having the same diameter, and the bottom formed in the second member 24 defines a hemispherical space. doing. In the well 3D shown in FIG. 10B, a cylindrical wall surface portion formed in the first member 23 defines a cylindrical space having the same diameter, and a bottom portion formed in the second member 24 defines a conical space. doing. The well 3E shown in FIG. 10C includes a cylindrical space having the same diameter from the cylindrical wall surface portion formed in the first member 23 to the bottom portion formed in the second member 24. Such wells 3C, 3D, and 3E can also be employed in the present invention.

  As described above, according to the present embodiment, the temperature of the cell aggregate C (object) accommodated in the well 3 can be accurately adjusted, while the cell aggregate C can be clearly observed. Can be provided.

1 Well plate 2 Plate body 21 Upper surface (first surface)
22 Lower surface (second surface)
23 first member 231 first opening 232 second opening 233 tube portion 23U first upper surface 23L first lower surface 24 second member 241 recess 242 third opening 24U second upper surface 24L second lower surface 3 well 31 opening portion 32 bottom portion 33 tube Shaped wall 34 Storage part 35 Receiving part 4 Sorting container 5 Camera unit (imaging device)
6 Chip 7 Cooling section (heat source)
8 Control unit

Claims (5)

A well plate in which the object is inserted or removed by a chip that sucks and discharges the object,
A plate body including a first surface and a second surface facing the first surface, the plate body being used with the first surface facing upward;
An opening provided on the first surface and into which the object is inserted; a bottom located inside the plate body; and a cylindrical wall surface extending downward from the opening toward the bottom, One or a plurality of wells capable of holding an object,
The plate body is
A first upper surface serving as the first surface; and a first lower surface facing the first upper surface, wherein an upper portion of the well is formed, and has a good thermal conductivity and a non-transparent first member; ,
A transparent second member having a second upper surface and a second lower surface opposite to the second upper surface and serving as the second surface, wherein a lower portion including at least a portion constituting the bottom of the well is formed. And comprising
Wherein the first lower surface of the first member and the second upper surface of said second member, said upper portion and said lower portion of said well a well plate ing are joined while being aligned ,
The first member includes a cylindrical portion constituting a portion excluding the vicinity of the lower end of the cylindrical wall surface, a first opening formed on the first upper surface and serving as the opening at the upper edge of the cylindrical portion, A second opening formed on the first lower surface and serving as a lower edge of the cylindrical portion,
The second member comprises the bottom and the vicinity of the lower end of the cylindrical wall surface, and includes a recess having a third opening on the second upper surface,
A well plate in which the first member and the second member are joined in a state where the second opening and the third opening are aligned . The well plate according to claim 1 ,
The well plate, wherein the second opening and the third opening have the same horizontal cross-sectional size. The well plate according to claim 1 ,
The well plate, wherein a horizontal sectional size of the third opening is larger than a horizontal sectional size of the second opening. The well plate according to any one of claims 1 to 3 ,
The cylindrical wall surface is
An accommodating portion having a first horizontal cross-sectional size that is a predetermined region above the bottom and capable of accommodating the object;
A well plate including a receiving portion having a second horizontal cross-sectional size larger than the first horizontal cross-sectional size, which is an area extending from above the housing portion to the opening. The well plate according to any one of claims 1 to 4 ,
A heat source for applying cold or hot to the first member of the plate body;
An imaging device that is disposed on the second surface side of the plate body and that captures an image of the object held in the well through the second member;
A device for observing an object.

Images