JP2014113132A - Cell culture vessel seat and method for cell culture - Google Patents

Abstract

PROBLEM TO BE SOLVED: To suppress the temperature drop of a cell culture vessel when the culture vessel is taken out from an incubator.SOLUTION: A cell culture vessel 2 is placed on a cell culture vessel seat 1. The vessel seat 1 has a heat storage function at least at a part thereof contacting with the bottom of the culture vessel 2. The culture vessel seat 1, for example, is composed of a heat storage member 11 contacting with the bottom of the culture vessel 2, and a frame member 12 circumscribing around the heat storage member 11. The culture vessel 2 in a state being placed on the vessel seat 1 is accommodated in an incubator, then taken out therefrom, and an operation such as a microscopic observation is performed.

Description

  The present invention relates to an instrument used for cell culture.

  In recent years, regenerative medical technology using cultured cells has attracted attention. When culturing cells, a so-called “temperature-responsive cell culture vessel” in which the surface of the material is covered with a temperature-responsive polymer is sometimes used. This temperature-responsive polymer has, for example, N-isopropylacrylamide as a basic skeleton, and the temperature-responsive polymer surface is hydrophobic near a culture temperature (for example, 37 ° C.) and has a critical temperature (for example, Below 30 ° C., it is hydrophilic. Therefore, when the culture container is near the culture temperature, the cells adhere to the surface of the temperature-responsive polymer and can be made confluent. When this culture vessel is cooled to a temperature lower than the critical temperature, cells are peeled off from the surface of the temperature-responsive polymer, and a cultured tissue in which cells are connected in a sheet form, so-called “cell sheet”, can be collected. Thus, by using a temperature-responsive polymer as a cell culture substrate, it becomes possible to stably produce a cell sheet without impairing the function of the cell.

  Conventionally, cell culture has been performed manually. However, it is desired to develop an automatic culture apparatus for automatically producing cultured cells such as cell sheets. Automation of cell culture is expected to reduce costs related to strict hygiene management to prevent contamination and to supply cultured cells with stable quality. For example, Patent Literature 1 proposes a culture observation apparatus that automatically performs cell culture and observation. This culture observation apparatus is provided with a transport robot for carrying a culture container into an incubator section. The transfer robot includes a heater for heating the culture container in the arm portion, and suppresses the occurrence of condensation when the culture container is carried into the culture environment.

JP 2010-158185 A

  Periodic microscopic observation of cells is essential in the process of culturing cells. Microscopic observation of cells is performed by removing a culture container from an incubator for culturing cells. The inside of the incubator is managed so as to have an environment suitable for cell culture (for example, a temperature of 37 ° C., a humidity of 90% or more, and a carbon dioxide concentration of 5%). The temperature of the culture vessel taken out from the incubator decreases with the passage of time in the working chamber adjusted to about 20 ° C. Here, in the case where the culture vessel is a temperature-responsive cell culture vessel, if the temperature of the culture vessel falls below the critical temperature, the cells are peeled off from the surface of the temperature-responsive polymer, preventing good cell culture. .

  If the transfer robot described in Patent Document 1 is used to carry in and out the culture vessel from the incubator, the culture vessel during transfer can be maintained at the culture temperature by heating with the heater of the arm portion of the transfer robot. However, if the heater is provided in the transfer robot, there are disadvantages such as a complicated structure of the transfer robot and a running cost such as electricity bill. Furthermore, since the culture container can be released from the arm portion of the transport robot during the microscope observation, it is not possible to suppress the temperature drop of the culture container.

  When transporting a culture container using a transport robot, it was somewhat difficult to directly grip the culture container with a robot hand. This is because a culture container generally called a dish or petri dish is a round plate made of glass or plastic, and thus requires a slight adjustment of the gripping force and gripping position of the robot hand. Therefore, the inventors have devised to provide a container seat on which the culture container is placed in order to make it easier to hold the culture container by the robot hand. When the robot hand grips the container seat on which the culture container is placed, the allowable range of the gripping force and the gripping position of the robot hand is increased and control is facilitated. However, even if the container seat is used, there remains a problem of temperature drop of the culture container when the culture container is taken out from the incubator. Although it is conceivable to apply the transfer robot of Patent Document 1 to provide a heater on the container seat, there is a disadvantage that the structure becomes complicated and that running costs such as electricity costs are required.

  In the first place, in order to prevent cells from peeling from the surface of the temperature-responsive polymer in the culture vessel during operations such as microscopic observation, the culture vessel is critical over the work time without maintaining the culture vessel temperature at the culture temperature. What is necessary is just to be able to suppress the temperature drop of a culture container to such an extent that it does not become below temperature. Therefore, in the present invention, the container seat on which the culture container is placed is provided with a heat storage function, and the temperature drop of the culture container is suppressed so that the culture container does not become below the critical temperature during operations such as microscopic observation. Here, the critical temperature is a temperature at which the temperature-responsive polymer surface of the temperature-responsive cell culture vessel changes from hydrophobic to hydrophilic.

  The cell culture container seat according to the present invention is a cell culture container seat on which a cell culture container is placed, and at least a part of the cell culture container seat contacting the bottom of the culture container has a heat storage function.

  If the culture vessel placed on the cell culture vessel seat is accommodated in the incubator, the cell culture vessel seat is maintained at the same temperature as the culture vessel and stores heat. And if a culture container is taken out from an incubator with a cell culture container seat, since a culture container is heat-retained with the heat | fever stored in the cell culture container seat, the temperature fall of the said culture container can be suppressed. Thus, in order to suppress the temperature drop of the culture vessel, wiring, a power source, and the like are unnecessary, and handling is easier than in the case where a heater is provided. Then, since the rate of temperature decrease of the culture container when the culture container is removed from the incubator becomes slow, for example, when the culture container is a temperature-responsive cell culture container, the culture container decreases from the culture temperature to below the critical temperature. In the meantime, it becomes possible to finish the work such as the observation of a cell microscope.

  The cell culture container seat according to the present invention is a cell culture container seat on which a cell culture container is placed, and includes a heat storage member that contacts a bottom of the culture container, and a frame member that surrounds the periphery of the heat storage member. It is provided.

  If the culture vessel placed on the cell culture vessel seat is accommodated in the incubator, the heat storage member of the cell culture vessel seat is maintained at the same temperature as the culture vessel and stores heat. And if a culture container is taken out from an incubator with a cell culture container seat, since a culture container is heat-retained with the heat stored in the heat storage member of a cell culture container seat, the temperature fall of the said culture container can be suppressed. Thus, in order to suppress the temperature drop of the culture vessel, wiring, a power source, and the like are unnecessary, and handling is easier than in the case where a heater is provided. Then, since the rate of temperature decrease of the culture container when the culture container is removed from the incubator becomes slow, for example, when the culture container is a temperature-responsive cell culture container, the culture container decreases from the culture temperature to below the critical temperature. In the meantime, it becomes possible to finish the work such as the observation of a cell microscope. Further, if the culture vessel is handled in a state where the culture vessel is placed on the cell culture vessel seat, the culture vessel can be easily held by the hand or robot hand through the cell culture vessel seat.

  The heat storage member preferably has transparency. Thereby, in the state which mounted the culture container on the cell culture container seat, the cell in a culture container can be observed with a microscope.

  The culture vessel may be a circular flat plate, and the heat storage member may have a cylindrical shape having an outer diameter equal to or larger than the outer diameter of the bottom of the culture vessel. According to this configuration, the entire bottom surface of the culture vessel can be in contact with the heat storage member, which is suitable for effectively and evenly suppressing the temperature drop of the culture vessel.

  The frame member may protrude upward or downward from the heat storage member. If the frame member protrudes upward from the heat storage member, the movement of the culture vessel can be regulated by the frame member. In addition, if the frame member protrudes downward from the heat storage member, the cell culture container seat comes into contact with the work table or the like with the frame member, and contamination of the bottom surface of the heat storage member is prevented, or the work table is removed from the cell culture container seat. The heat transfer to can be reduced.

  The frame member preferably has a heat insulating property. Thereby, the heat transfer from the periphery of the cell culture container seat to the outside can be reduced, and the heat storage function of the cell culture container seat can be enhanced.

  The heat storage member can be made of glass, for example. The frame member can be made of, for example, polytetrafluoroethylene.

  In the cell culture method according to the present invention, the cell culture container is loaded into an incubator, stored in the incubator, and unloaded from the incubator while being placed on a container seat having a heat storage function. According to this method, the temperature drop when the culture container is taken out from the incubator can be reduced.

  Furthermore, it is preferable to perform microscopic observation of cells cultured in the culture container in a state where the culture container is placed on the container seat. According to this method, the temperature drop of the culture container during the microscopic observation of the cells can be reduced. In particular, when the culture vessel is a temperature-responsive cell culture vessel, it is possible to prevent the cells from being detached from the surface of the temperature-responsive polymer due to the temperature drop.

  The present invention has the following effects.

  If the culture vessel placed on the cell culture vessel seat described above is accommodated in the incubator, the cell culture vessel seat is maintained at the same temperature as the culture vessel and stores heat. And if a culture container is taken out from an incubator with a cell culture container seat, since a culture container is heat-retained with the heat | fever stored in the cell culture container seat, the temperature fall of the said culture container can be suppressed.

It is a perspective view of the container seat which concerns on embodiment of this invention. It is a perspective view of the container seat which mounted the culture container. It is a top view of a container seat. It is IV-IV sectional drawing of the container seat shown in FIG.

  Hereinafter, preferred embodiments of the present invention will be described with reference to the drawings. In the following description, the same or corresponding elements are denoted by the same reference symbols throughout the drawings, and redundant description thereof is omitted.

  FIG. 1 is a perspective view of a cell culture container seat (hereinafter simply referred to as a container seat 1) according to an embodiment of the present invention, and FIG. 2 is a perspective view of the container seat 1 on which the culture container 2 is placed. As shown in FIGS. 1 and 2, a container seat 1 according to the present invention is used with a culture container 2 mounted thereon. The culture vessel 2 is a round plate made of glass or plastic generally called a dish or petri dish. FIG. 2 shows the circular flat dish with a lid. The culture vessel 2 is a temperature-responsive cell culture vessel using a temperature-responsive polymer as a culture substrate. This temperature-responsive polymer has, for example, N-isopropylacrylamide as a basic skeleton, and the surface of the temperature-responsive polymer is hydrophobic near the culture temperature (37 ° C.) and has a critical temperature (for example, 30 ° C.). Below it is hydrophilic.

  3 is a plan view of the container seat 1, and FIG. 4 is a cross-sectional view taken along the line IV-IV of the container seat 1 shown in FIG. As shown in FIGS. 3 and 4, the container seat 1 is mainly formed of a heat storage member 11 and a frame member 12. The heat storage member 11 has a short cylindrical shape, and the outer diameter thereof is the same as or larger than the outer diameter of the bottom of the culture vessel 2. The height of the heat storage member 11 (that is, the volume of the heat storage member 11) is determined so as to have a predetermined heat storage amount. The predetermined heat storage amount is a heat storage amount sufficient to prevent the temperature of the cultured cells in the culture vessel 2 from dropping to a critical temperature or lower during work such as microscopic observation. The heat storage member 11 is made of a material having heat storage properties and transparency. The heat storage member 11 according to the present embodiment is made of glass, and the heat storage medium of the container seat 1 is glass.

  The frame member 12 has a short cylindrical shape that borders the periphery of the heat storage member 11. The inner diameter of the frame member 12 is substantially equal to the outer diameter of the heat storage member 11, and the outer diameter is also larger than the outer diameter of the culture vessel 2. The height of the frame member 12 is larger than the height of the heat storage member 11, and the frame member 12 protrudes upward and downward from the heat storage member 11. The frame member 12 is made of a material having chemical resistance and heat resistance. The heat resistance of the frame member 12 is sufficient if it can withstand the temperature of steam sterilization (for example, 120 ° C.). Furthermore, it is desirable that the constituent material of the frame member 12 has a heat insulating property. If the frame member 12 has heat insulation, heat transfer from the periphery of the container seat 1 to the outside can be reduced, and the heat storage function of the container seat 1 can be enhanced. As will be described later, since the container seat 1 substantially contacts the work table or the like with the frame member 12, heat exchange between the work table or the like and the container seat 1 is suppressed to reduce the temperature of the container seat 1 during operation. Can be prevented. The frame member 12 according to the present embodiment is made of polytetrafluoroethylene that is chemically stable, has excellent heat resistance and chemical resistance, and has heat insulation properties. Polytetrafluoroethylene is superior in impact resistance compared to glass.

  On the outer peripheral surface of the frame member 12, one or a plurality of grooves 21 for preventing slipping are formed. On the upper part of the frame member 12, an upper protrusion 22 that protrudes toward the inner periphery is formed continuously for one turn. The inner diameter of the upper protrusion 22 is smaller than the outer diameter of the heat storage member 11, and the lower surface of the upper protrusion 22 is in contact with the peripheral edge of the upper surface of the heat storage member 11. Further, the inner diameter of the upper protrusion 22 is substantially equal to the outer diameter of the bottom of the culture vessel 2. On the other hand, in the lower part of the frame member 12, a lower protrusion 23 that protrudes toward the inner periphery is formed continuously or intermittently. The inner diameter of the lower protrusion 23 is smaller than the outer diameter of the heat storage member 11, and the upper surface of the lower protrusion 23 is in contact with the peripheral edge of the lower surface of the heat storage member 11.

  The heat storage member 11 is press-fitted into the inner periphery of the frame member 12 from below. When the heat storage member 11 is pushed into the frame member 12, the lower protrusion 23 of the frame member 12 is pressed against the heat storage member 11 from the inner peripheral side, and the frame member 12 is elastically deformed so that the inner diameter increases. When the heat storage member 11 is further pushed into the frame member 12, the frame member 12 from which the lower end of the heat storage member 11 passes through the lower protrusion 23 and the external force is removed returns to the original shape, and the lower protrusion 23 is removed from the heat storage member 11. Acts as a stop.

  The container seat 1 having the above configuration has a short cylindrical shape as a whole, and has a diameter that is slightly larger than the diameter of the culture container 2. On the upper surface of the container seat 1, a circular recess is formed by the upper protrusion 22 of the frame member 12 and the upper surface of the heat storage member 11. By fitting the bottom of the culture vessel 2 into the recess on the upper surface of the vessel seat 1, the movement of the culture vessel 2 in a direction substantially parallel to the upper surface of the vessel seat 1 is restricted. Further, the outer diameter of the heat storage member 11 appearing in the recess on the upper surface of the container seat 1 is larger than the outer diameter of the bottom of the culture container 2. Thereby, the entire bottom surface of the culture vessel 2 can be brought into contact with the heat storage member 11, which is suitable for effectively and evenly suppressing the temperature drop of the culture vessel 2. On the other hand, on the bottom surface of the container seat 1, a circular recess is formed by the lower protrusion 23 of the frame member 12 and the upper surface of the heat storage member 11. Due to the recess on the bottom surface of the container seat 1, when the container seat 1 is placed on a work table or the like, the frame member 12 comes into contact with the work table or the like, and the bottom surface of the heat storage member 11 is separated from the work table or the like. Dirt such as dust adhering to the bottom surface of the heat storage member 11 can be prevented.

  Next, the usage example of the container seat 1 is shown. As shown in FIG. 2, the culture vessel 2 is placed on the vessel seat 1 such that the bottom portion of the culture vessel 2 is fitted into the recess on the upper surface of the vessel seat 1. At this time, the bottom of the culture vessel 2 is in contact with the heat storage member 11 of the vessel seat 1. In this way, the culture container 2 is handled in a state where the culture container 2 is placed on the container seat 1. When the culture container 2 is carried into the incubator from a work table or the like, the container seat 1 is grasped by hand or a robot hand, and the culture container 2 is moved together with the container seat 1 into the incubator. The container seat 1 is accommodated in the incubator together with the culture container 2, and is maintained at the culture temperature in the incubator. When the culture vessel 2 is taken out of the incubator, the vessel seat 1 is grasped with a hand or a robot hand, and the culture vessel 2 is moved from the inside of the incubator together with the vessel seat 1. By gripping the container seat 1 and moving the culture container 2 in this way, the gripping force and the allowable range of the gripping position of the robot hand are increased compared to when the culture container 2 is directly gripped. Control becomes easy. Also, when working with a human hand, gripping the container seat 1 instead of the culture container 2 makes it easier to work as compared to directly gripping the fragile and lightweight culture container 2.

  The container seat 1 and the culture container 2 are maintained at the culture temperature in the incubator, and the container seat 1 stores heat. The temperature of the culture vessel 2 taken out from the incubator controlled to the culture temperature of about 37 ° C. into the working chamber adjusted to about 20 ° C. gradually decreases with time. Here, since the culture container 2 is kept warm by the heat stored in the container seat 1 that is in contact with it, the temperature drop of the culture container 2 is suppressed. In other words, compared with the case where the container seat 1 does not exist, the temperature decrease rate of the culture container 2 becomes gentle. Therefore, it becomes possible to finish operations such as cell microscope observation until the temperature of the culture vessel 2 drops below the critical temperature. For example, observation with a cell microscope usually requires a work time of about 2 to 3 minutes and at most about 5 minutes. The temperature drop of the culture vessel 2 is suppressed by the vessel seat 1 so that the temperature of the culture vessel 2 does not fall below the critical temperature from the culture temperature during this working time. Thus, if the container seat 1 which has heat storage property is used in order to suppress the temperature drop of the culture container 2, wiring, a power supply, etc. are unnecessary and it is a simple structure compared with the case where heating means, such as a heater, are provided. And easy to handle.

  When observing cells, the culture vessel 2 is placed on the microscope stage with the culture vessel 2 placed on the vessel seat 1. In general, a phase contrast microscope is used for cell observation, but the heat storage member 11 of the container seat 1 transmits the irradiation light from below the microscope stage and does not block it. Therefore, with the culture vessel 2 placed on the vessel seat 1, the cells in the culture vessel 2 can be observed with a microscope in the same manner as when the vessel seat 1 does not exist. Also, when the culture container 2 is visually observed, the heat storage member 11 of the container seat 1 is transparent, so that the cells in the culture container 2 can be observed in the same manner as when the container seat 1 is not present.

  The preferred embodiment of the present invention has been described above. However, the present invention is not limited to the above-described embodiment, and various design changes can be made as long as they are described in the claims. is there.

  For example, the shape of the frame member 12 of the container seat 1 is not limited to the above embodiment. The frame member 12 of the container seat 1 protrudes above and below the heat storage member 11, but the frame member 12 of the container seat 1 has a shape protruding above or below the heat storage member 11. Also good. If the frame member 12 protrudes above the heat storage member 11, the movement of the culture vessel 2 can be regulated by the frame member 12. Further, if the frame member 12 protrudes downward from the heat storage member 11, the container seat 1 substantially contacts the work table or the like with the frame member 12, so that the bottom surface of the heat storage member 11 is prevented from being soiled, It is possible to reduce heat transfer from 1 to a work table or the like.

  The container seat of the cell culture container according to the present invention is not limited to the temperature reduction of the temperature-responsive cell culture container, and is generally widely available as a technique for suppressing the temperature reduction of the culture container.

DESCRIPTION OF SYMBOLS 1 Container seat 2 Culture container 11 Heat storage member 12 Frame member 21 Groove 22 Upper protrusion 23 Lower protrusion

Claims (10)

A cell culture container seat on which a cell culture container is placed,
A cell culture container seat, wherein at least a part of the culture container in contact with the bottom has a heat storage function. A cell culture container seat on which a cell culture container is placed,
A heat storage member in contact with the bottom of the culture vessel;
A cell culture container seat comprising a frame member that borders the periphery of the heat storage member.   The cell culture container seat according to claim 2, wherein the heat storage member has transparency. The culture vessel is a round flat plate,
The cell culture container seat according to claim 2 or 3, wherein the heat storage member has a cylindrical shape having an outer diameter equal to or larger than an outer diameter of a bottom of the culture container.   The cell culture container seat according to any one of claims 2 to 4, wherein the frame member protrudes to one or both of the upper side and the lower side of the heat storage member.   The cell culture container seat according to claim 5, wherein the frame member has a heat insulating property.   The cell culture container seat according to any one of claims 2 to 6, wherein the heat storage member is made of glass.   The cell culture container seat according to any one of claims 2 to 7, wherein the frame member is made of polytetrafluoroethylene.   A cell culture method in which a cell culture container is loaded into an incubator, stored in the incubator, and unloaded from the incubator in a state where the cell culture container is placed on a container seat having a heat storage function.   The cell culture method according to claim 9, wherein the cells cultured in the culture container are observed with a microscope in a state where the culture container is placed on the container seat.

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