Patent Application
20160264918
The present invention relates to a cell culture apparatus to culture a cell using a culture container.
Conventionally, work of culturing a cell has been performed by manual labor of a skilled worker under a strict manufacturing process in a clean room which is unlimitedly sterilized. Thus, enormous labor cost and facility cost are required in the case of culturing the cell in large amount, which causes a great barrier to industrialization.
It is possible to automate a series of culture operations, which has been performed by the manual labor, using a robot, and to reduce the labor cost. However, it is not necessarily avoidable to a risk of contamination from outside since there is an operation to open an inside of a culture container, for example, in order to replace a culture medium by opening a lid of the culture container. Thus, there is a need of installing the entire system including the robot in a large-scale clean room, and it is difficult to significantly reduce the facility cost.
Accordingly, a system has been proposed which cultures a cell by forming a single sealed system by connecting a liquid bag with a culture medium therein and a culture container or the like (hereinafter, referred to as a closed culture system), and performing replacement of the culture medium or the like inside the closed culture system. An example thereof is described in JP2011-142837 A (PTL 1), for example. Accordingly, the cleanliness of a location in which the system has been installed may be kept while enabling the exclusion of the risk of contamination from the outside, and the significant reduction of the facility cost becomes possible.
In addition, a method of controlling liquid supply in a closed culture system which has a plurality of culture containers is described in JP 2009-125027 A (PTL 2), for example.
PTL 1: JP 2011-142837 A
PTL 2: JP 2009-125027 A
In general, it is considered to increase a culture area in order to increase a yield of cell culture. At the time, dozens or hundreds of types of culture containers having different sizes may be prepared to selectively use the culture container according to the yield, which is diseconomy. Therefore, it is economical to prepare culture containers having a limited number of types of sizes, and to change the yield by changing the number of the containers. Such a technique of simultaneously culturing the same cells in a plurality of the culture containers is often performed in order to increase the yield. The important point at this time is that there is no variation in quality of results of the culture. In order for this, it is important to uniformly perform a culture operation such as culture medium replacement among the plurality of culture containers. This is also applied to a closed culture system which has a plurality of culture containers.
A method of connecting culture containers in series and a method of connecting culture containers in parallel are considered in order to form a single closed culture system with the plurality of culture containers.
An example of advantages of the method of connecting the culture containers in series is that it is easy to control the liquid supply such as the culture medium replacement. Since there is no branch, the liquid reliably and uniformly spreads to each culture container although being on a rotating basis. However, there is a problem that the quality of liquid is easily degraded as going down from the upstream to the downstream in the case of the series connection. It is also necessary to obtain uniformity in terms of quality as well as of amount of the supplied liquid in order to minimize a variation in the culture quality. In addition, there is another problem that, in a case in which contamination occurs at the upstream, the downstream is also contaminated.
It is possible to solve the above-described problem when the culture containers are connected in parallel to separately supply the liquid. A method of supplying a liquid by switching culture containers (set), which are connected in parallel, using a valve is described in PTL 2, described above. Accordingly, a homogeneous liquid supply with respect to the plurality of culture containers becomes possible, but there is a problem that the number of the valves increases in the case of increasing the number of branches when the valve is provided for each branch, which causes an increase of cost.
Therefore, the present invention has been made in view of the above-described problems, and a representative object thereof is to provide a cell culture apparatus which enables a uniform and homogeneous liquid supply among a plurality of culture containers while keeping cost low.
Other objects and novel characteristics in addition to the above-described ones of the present invention will be apparent from description of the present specification and the attached drawings.
An overview of representatives of the invention to be disclosed in the present application will be simply described as follows.
That is, a representative cell culture apparatus is a cell culture apparatus in which a plurality of sealed culture containers having fluid introduction ports and discharge ports are provided, and the plurality of culture containers are connected in parallel, forming a single closed culture system. The cell culture apparatus has a single flow channel switching mechanism for switching a plurality of flow channels connected to the plurality of culture containers.
More preferably, the flow channel switching mechanism in the cell culture apparatus makes a flow channel resistance of one individual flow channel from among the individual flow channels branching and being connected to the culture containers less than the flow channel resistance of the remainder of the individual flow channels.
More preferably, the flow channel switching mechanism in the cell culture apparatus forms an individual flow channel having a small flow channel resistance by placing a single flow channel switching member in the closed culture system and changing at least one of an orientation and a position of the flow channel switching member.
An effect that can be obtained by the representatives of the invention to be disclosed in the present application will be simply described as follows.
That is, the representative effect is that it is possible to provide a cell culture apparatus which enables a uniform and homogeneous liquid supply among a plurality of culture containers while suppressing cost.
In the embodiments below, although a description will be given by dividing the embodiment into a plurality of sections or embodiments if necessary for convenience, these are not irrelevant to each other excepting the case that is particularly demonstrated, but are in a relationship in which one is a modified example of part or all of the other, a detailed description, a supplementary description, or the like. In addition, in the embodiments below, when the number of elements and the like (including the number, a numeric value, a quantity, a range, and the like) are stated, the embodiment is not limited to a particular number excepting the case that is particularly demonstrated or a case in which the embodiment is clearly limited, in principle, to the particular number, and the number may be more than or less than the particular number.
Further, in the embodiments below, it is obvious that the constituent components (including component steps and the like) are not necessarily required, excepting the case that is particularly demonstrated or a case in which the components are clearly required in principle. Similarly, in the embodiments below, when shapes, positional relationships and the like of the constituent components are stated, it is assumed that those substantially approximate to or analogous to the shapes and the like are included excepting the case that is particularly demonstrated or a case in which the components are obviously inappropriate in principle. This also applies to the numeric value and the range described above.
First, an overview of an embodiment will be described. The overview of the embodiment will be described by providing reference numeral or the like of constituent components corresponding to the embodiment in parentheses, for example.
A representative cell culture apparatus of the present embodiment is a cell culture apparatus in which a plurality of sealed culture containers (culture containers 1, 35, 41 and 44) having fluid introduction ports and discharge ports are provided, and the plurality of culture containers are connected in parallel, forming a single closed culture system. The cell culture apparatus has a single flow channel switching mechanism (flow channel switching mechanism 8 or 19) for switching a plurality of flow channels connected to the plurality of culture containers.
More preferably, the flow channel switching mechanism in the cell culture apparatus makes a flow channel resistance of one individual flow channel from among the individual flow channels branching and being connected to the culture containers less than the flow channel resistance of the remainder of the individual flow channels.
More preferably, the flow channel switching mechanism in the cell culture apparatus forms an individual flow channel having a small flow channel resistance by placing a single flow channel switching member (flow channel switching member 9, 12, 13, 29, 37 or 39) in the closed culture system and changing at least one of an orientation and a position of the flow channel switching member.
Hereinafter, a detailed description will be given regarding the respective embodiments based on the overview of the embodiment described above with reference to the drawings. Incidentally, the same reference numerals will be attached to members having the same function, in principle, in the entire drawing for describing the respective embodiments, and the repetitive description thereof will be omitted. In addition, the description of the same or similar portions will not be repeated in principle unless particularly required in the respective embodiments.
In addition, there is a case in which hatching is not illustrated even in a cross-sectional view in order for the viewability of the drawing in some drawings to be used in the respective embodiments. In addition, there is a case in which hatching is illustrated even in a plan view in order for the viewability of the drawing.
A cell culture apparatus according to the present embodiment will be described with reference to
The cell culture apparatus according to the present embodiment includes a culture container 1, a supply bag 2 in which a culture medium or the like is housed, a collection bag 3 which collects the used culture medium or the like, and a flow channel switching mechanism 8. The culture container 1, the supply bag 2, and the collection bag 3 are connected via flow channels in this cell culture apparatus. A plurality of (four in the example of
Since the cell culture apparatus according to the present embodiment is the closed culture system, it is necessary to apply a driving force of a liquid from the outside of system. An example of the means for applying the driving force is a peristaltic pump that supplies the liquid by squeezing an elastic tube from an outer side. A least a part of the common flow channel may have the elasticity so as to enable the liquid supply using the peristaltic pump. The peristaltic pump may be installed in the common flow channel 4 on the upstream side or in the common flow channel 7 on the downstream side. In addition, the peristaltic pump may be installed in both the common flow channel 4 on the upstream side and the common flow channel 7 on the downstream side.
When the peristaltic pump (not illustrated) is driven, the liquid inside the supply bag 2 is transferred to the culture container 1. At the time, the liquid is transferred while changing the culture containers 1 by a switching operation of the flow channel switching mechanism 8. The liquid which has been originally contained in the culture container 1 is sent to be pushed out to the collection bag 3.
The flow channel switching mechanism 8 includes a flow channel switching member 9 and a storage compartment 10 in which low channel switching member 9 is stored. The common flow channel 4 on the upstream side and the plurality of (four in the examples of
The flow channel switching member 9 has a cylindrical shape in
Incidentally, the up and down may be reversed in
Even if a part of the liquid flows out to an undesired branch in the closed culture system that switchably supplies the liquid to the plurality of culture containers 1 being connected in parallel, such a liquid is originally planned to flow, and thus, there is no influence on the culture quality. Thus, the branch for which the liquid supply is not desired is not necessarily closed completely, but may be provided with a difference in flow channel resistance against a branch for which the liquid supply is desired to make the liquid hardly flow thereto in terms of an object of homogeneously supplying the liquid to the respective culture containers 1.
Therefore, a size of the flow channel switching member 9 may be a size which is slightly smaller than the storage compartment 10 so as to enter the storage compartment 10 with a gap. The supplied liquid can flow out to the undesired branch from the slight gap between the flow channel switching member 9 and the storage compartment 10, but there is no problem on the quality as described above even if the liquid has flown out. In addition, it is not preferable to raise liquid supply speed from a point of view of a fixing property of a cell in fact, and the above-described flowing out rarely occurs when the flow channel resistance is large to some extent since the liquid supply pressure is not high.
The flow channel switching mechanism 8 of the present embodiment is schematically drawn like
(r1+r)<<(r2+R) Formula 1
When the relation of Formula 1 is established, the liquid rarely flows to the R side. The above-described Formula 1 is satisfied when r1 and r2 are set to be sufficiently smaller than R, and r<<R in the flow channel switching mechanism 8.
The following effect is obtained using a flow channel switching method according to the flow channel switching mechanism 8 of the present embodiment.
First, it is possible to lower a manufacturing cost of the flow channel switching mechanism 8. A high dimensional accuracy is required for the fitting of the flow channel switching member 9 and the storage compartment 10 in order to eliminate or infinitely suppress the flowing out of the supplied liquid to the branch other than the desired one. It is necessary to perform machining in some cases, thereby causing extremely high cost. The closed culture system including the flow channel switching mechanism 8 according to the present embodiment may be reused after performing sterilizing treatment for each culture, but may be preferably discarded after each culture in order to lower a risk of contamination. Accordingly, the manufacturing cost is extremely important. It is possible to loosen the fitting dimension in a case in which the slight flowing out of the supplied liquid is allowable as in the present embodiment. When a level of molding dimensional accuracy of a general molded article may be enough, the significant cost reduction is possible.
Another effect is that a force for switching the orientation of the flow channel switching member 9 is not required much. The flow channel switching member 9 and the storage compartment 10 are fitted with each other with the gap, and do not strongly contact each other in the flow channel switching method of the present embodiment. Thus, it is not necessary to exert a large force in order to change the orientation of the flow channel switching member 9. Accordingly, it is also possible to provide simple mechanism and structure for switching the flow channel switching member 9.
Hereinafter, a description will be given regarding modified examples of the structure of the flow channel switching mechanism in order with reference to
The flowing out of the supplied liquid other than the desired one in the flow channel switching mechanism is wasteful although is not problematic in terms of the culture quality, it is desirable to tighten the fitting dimension between the flow channel switching member 9 and the storage compartment 10 as much as possible. Alternatively, a size of the flow channel switching member 9 may be set to be larger than the storage compartment 10 using a material with a low Young's modulus. An example thereof is illustrated in
When the flow channel switching member 12 is in strong contacts the storage compartment 10, it is necessary to exert a large force to change an orientation of the flow channel switching member 12, and thus, it is preferable if there is a scheme to reduce the force. For example, it is preferable to reduce the contact surface as much as possible in order to decrease friction on the sliding surface. A material with high lubricity like Teflon may be used.
The material having the low Young's modulus may be applied to either side or both sides of the flow channel switching member and the storage compartment. Alternatively, the material may be applied partially not to the entire member. An example thereof is illustrated in
The connecting flow channel to be formed in the flow channel switching member 9 is not limited to a tube structure, but may be a structure like a groove. Examples thereof are illustrated in
In addition, the branching flow channel 5 to be connected together can be provided on a lower surface as illustrated in
It may be enough when the connecting flow channel of the flow channel switching member forms the single path having the low the flow channel resistance at the time of supplying the liquid, and the number thereof is not limited to one but may be plural. Examples thereof are illustrated in
It is advantageous when the flow channel switching member, which is switched by the rotation, has the connecting flow channel such that an end thereof is positioned at a central axis of rotation. When the common flow channel is arranged on the central axis of rotation, the common flow channel and the connecting flow channel necessarily face each other even when the rotation of the flow channel switching member is arbitrary.
Meanwhile, the flow channel switching member may have the connecting flow channel of which one end is not intentionally positioned at the central axis of rotation. Examples thereof are illustrated in
It is possible to obtain the following effect with the above-described structure.
In a case in which one end of the connecting flow channel is positioned at the central axis of rotation, the number of the connecting flow channels is two at most on the upper surface and the lower surface of the cylinder. However, it is possible to provide an innumerable connecting flow channels in principle if one end of the connecting flow channel is not necessarily provided at the central axis of rotation.
When a plurality of connecting flow channels are provided, it is possible to perform the switching of the flow channels of the plurality of common flow channel using the single flow channel switching member 9. Examples thereof are illustrated in
When the above-described flow channel switching member is provided, two flow channel switching mechanisms 8a and 8b, illustrated in
<Method of Switching Flow Channel Switching Member>
Next, a description will be given regarding a method of switching the orientation of the flow channel switching member. Hereinafter, a description will be given regarding examples and modified examples of the method of switching the flow channel switching member in order with reference to
Since the flow channel switching member is inside the closed system, it is difficult to directly contact the member for switching, and another scheme is required. One of the switching method uses a remotely applied force. Examples of the remotely applied force that can be used includes a magnetic force and gravity.
An electromagnet may be used as the magnetic field generation means.
The orientation of the force is a single direction and is not changed in the case of using the gravity, and thus, the switching is difficult as compared to the case of using the magnetic force, but it is possible to switch the flow channel by using a switching member of which the center of gravity is made eccentric and rotating the storage compartment side.
Next, a description will be given regarding a method of indirectly grabbing and switching the flow channel switching member. Examples thereof are illustrated in
It is preferable to provide a scheme that facilitates the grabbing in the flow channel switching member 9. For example, a concave structure 24 is provided in a part of the flow channel switching member 9, and the flow channel switching member 9 is grabbed by an external force transmission mechanism 25 having a counterpart convex structure via the film 23 as illustrated in
A part of the flow channel switching member 9 may be formed to protrude from the storage compartment 10 as illustrated in
Although the film 23 has elasticity or flexibility, there is a limit in variable amount, and thus, a scheme is required to prevent the film 23 from being rotated more than necessary in the state of being grabbed. For example, there is a method of dividing the amount of rotation and changing the grabbing instead of rotating at a time. It is possible to repeat the rotation operation as deformation of the film is recovered by the elasticity or the deflection is recovered at the time of releasing the film for changing the grabbing. It is also effective to combine the above-described scheme with the scheme for reducing the amount of rotation at the time of the switching by providing the plurality of connecting flow channel s in the flow channel switching member as illustrated in
As illustrated in
It may be enough when the flow channel switching member 9 has the connecting flow channel being arranged inside the closed system, and thus, it is not necessarily to accommodate the flow channel switching member 9 in the storage compartment 10. It may be configured such that the structure which is called the storage compartment is not provided as illustrated in
As illustrated in
The flow channel switching mechanism 8 may be arranged on the downstream in the cell culture apparatus according to the present embodiment as illustrated in
In this manner, the flow channel switching mechanism 8 is arranged at the merging portion between the branching flow channels 6 on the downstream side and the common flow channel 7 on the downstream side.
In this flow channel switching mechanism 8, the liquid may enter the gap between the flow channel switching member and the storage compartment upon the structure, which becomes the dead volume. Such a liquid becomes wasteful if the flow channel switching mechanism 8 is at the upstream, but is not wasteful if the flow channel switching mechanism 8 is at the downstream which is a drainage side. In addition, when the flow channel switching mechanism 8 is arranged at the downstream, there is also an advantageous that the restriction with respect to the material of the member forming the flow channel switching mechanism 8 is mitigated. Since the liquid flows to the culture container 1 passing through the flow channel switching mechanism 8 in a case in which the flow channel switching mechanism 8 is arranged at the upstream, it is essential that the material of the component does not affect the culture, but the material is not limited as long as the flow channel switching mechanism 8 is at the downstream on the drainage side.
As described above, it is possible to uniformly and homogeneously supply the liquid among the plurality of culture containers 1 while making the cost low, as the representative effects, by including the single flow channel switching mechanism 8 to switch the plurality of flow channels connected with the plurality of culture containers 1 according to the cell culture apparatus according to the present embodiment. As a result, it is possible to achieve homogenization in culture quality among the culture containers 1. In addition, it is possible to perform the cell culture simultaneously in the plurality of culture containers 1, and thus, it is possible to increase a yield by increasing the number of the culture containers 1. In addition, it is possible to change the number of the culture containers 1 or the like in accordance with a target yield, and thus, a flexible response with respect to the cell culture becomes possible. Other effects are as described as in the present embodiment.
Further, since it is possible to obtain a plurality of cultures, it is possible to provide one of the obtained cultures for inspection in addition to one for transplantation, for example, which is an additional effect. In the case of an invasive inspection such as a staining test, the inspection becomes a so-called destructive inspection, and thus, it is difficult to provide the inspected cell for transplantation. When a plurality of cells are cultured simultaneously as in the present embodiment, it is possible to allow a usage in which one of the cultures is taken out for inspection prior to transplantation, and the other cultures are used for transplantation after confirming that there is no problem in terms of quality.
A cell culture apparatus according to the present embodiment will be described with reference to
A flow channel switching member may perform reciprocating motion such as linear motion or screwing motion without being limited to the rotational motion as in Embodiment 1 described above. A description will be given regarding an example of the flow channel switching member which performs the reciprocating motion in the present embodiment. A point that is different from Embodiment 1 described above will be mainly described in the present embodiment.
In
It may be configured such that the connecting flow channel is formed in a shape of the through-hole 31 and a groove 32 are combined, and the common flow channel 4 is directly connected together with the storage compartment 30 as illustrated in
As illustrated in
The flow channel switching member performing the rotational motion can be switched only by changing the orientation, and a range of motion of the flow channel switching member is not changed, and thus, it is possible to downsize the storage compartment. However, it is necessary to three-dimensionally arrange the common flow channel, the branching flow channel, and the connecting flow channel. On the contrary, it is possible to two-dimensionally arrange the above-described channels in the reciprocating motion equation as in the present embodiment, and thus, the latter configuration is suitable in a case in which it is desired to provide a thin flow channel switching mechanism.
Incidentally, a shape or the number of the connecting flow channels, a method of switching the flow channel switching member, and the like also can be considered similarly as those of the rotational motion equation even in the reciprocating motion equation.
As described above, it is possible to obtain a different effect as follows in addition to the same effects of Embodiment 1 described above according to the cell culture apparatus according to the present embodiment. For example, a representative effect is that it is possible to provide the thin flow channel switching mechanism since it is possible to two-dimensionally arrange the common flow channel, the branching flow channel, and the connecting flow channel by providing the flow channel switching member 29 which performs the reciprocating motion such as the linear motion and the screwing motion. Other effects are as described as in the present embodiment.
A description will be given regarding a cell culture apparatus according to the present embodiment with reference to
Further, each of
A description will be given regarding the example of the configuration of the integrated flow channel which includes the flow channel switching mechanism in the present embodiment. A point that is different from Embodiments 1 and 2 described above will be mainly described also in the present embodiment.
In a case in which the plurality of culture containers are connected in parallel, the corresponding number of branches are required, and the number of flow channels increases, which is complicated. When the individual flow channels are connected one by one, it takes a lot of man-hours, and further, there is a risk of erroneous piping. Therefore, it is preferable to collect these flow channels to form an integrated member (integrated flow channel member). Examples thereof are illustrated in
A configuration which includes the culture container and the integrated flow channel member enclosing the flow channel switching member as illustrated in
A culture container 35 has a culture surface 35a, an inflow channel 35b, and a discharge flow channel 35c. An integrated flow channel member 36 has an inflow port 36a, an upstream common flow channel 36b, an upstream branching flow channel 36c, a downstream branching flow channel 36d, a storage compartment 36e serving as the flow channel switching member, a downstream common flow channel 36f, and a discharge port 36g. In addition, the culture container 35 and the integrated flow channel member 36 have a connection port 35d and ports 36h, respectively, to enable connection with each other. The culture container 35 has the single connection port 35d, and the integrated flow channel member 36 has a plurality of the ports 36h to enable connection with a plurality of containers. In addition, a flow channel switching member 37 is provided inside the storage compartment 36e.
The connection port 35d of the culture container 35 is linked with the inflow channel and the discharge flow channel, and the ports 36h of the integrated flow channel member 36 are linked with upper and lower branching flow channels. When the integrated flow channel member 36 and the culture container 35 are connected, the upstream branching flow channel 36c and the inflow channel 35b of the culture container 35, and the downstream branching flow channel 36d and the discharge flow channel 35c of the culture container 35 are caused to face each other. Desirably, each of the connection port 35d and the port 36h has one surface, and the above-flow channels are linked with the respective surfaces, and accordingly, the flow channels face each other by causing the surfaces match each other. In this manner, it is possible to attach and detach the culture container 35 from a single direction with respect to the integrated flow channel member 36, thereby facilitating the attachment and detachment. Each of the connection port 35d and the port 36h may have a plurality of surfaces in order for positioning or sealing, but an angle formed with respect to each normal line of surface need not exceed 180°. In this manner, the attachment and detachment from the single direction is possible even if the plurality of surfaces are provided although depending on the arrangement of surfaces.
An inlet 35e and an outlet 35f of the liquid in the culture surface 35a of the culture container 35 may be arranged to oppose each other with the culture surface 35a interposed therebetween such that the liquid supply spreads over the entire surface. Meanwhile, it is preferable to allow the inflow channel 35b and the discharge flow channel 35c to be linked with one surface of the connection port 35d as described above in order for easiness of connection with the integrated flow channel member 36, and thus, a scheme is required for the arrangement of flow channels inside the culture container 35. For example, the two inflow channel 35b and discharge flow channel 35c, which are parallel in a tangential direction of the circular culture surface 35a, are drawn out to be linked with the connection port 35d as illustrated in
When being installed, the culture container 35 may be pushed to be fit and fixed in the single direction using a snap-fit structure so as to enable the simple installation. Examples of the structure thereof are illustrated in
A scheme for preventing a leakage is provided between the ports 36h of the integrated flow channel member 36 and the connection port 35d of the culture container 35 using a seal such as a gasket 38p illustrated in
It is also preferable to form the ports 36h of the integrated flow channel member 36 and the connection port 35d of the culture container 35 in asymmetric shapes so as not to be attached upside down.
When such shapes are formed, the closed culture system is formed only by connecting the culture container 35 to each of the ports 36h of the integrated flow channel member 36, and connecting the supply bag and the collection bag to the inflow port 36a and the discharge port 36g of the integrated flow channel member 36, respectively. As a result, it does not take the man-hours, and the erroneous piping with the culture container 35 hardly occurs. In addition, it is possible to lower the cost as the upper and lower common flow channels and branching flow channels are integrated.
Incidentally, there is a method of dividing the integrated flow channel member into two divided members 36s and 36t, inserting the flow channel switching member 37 therebetween, and then joining the two divided members 36s and 36t using, for example, ultrasonic welding, as illustrated in
Incidentally, it may be enough that the switching of the flow channels allows the flow channel switching member 39 and a desired branching flow channel inside the integrated flow channel member 38 to be aligned to face each other, and the drive target is not necessarily limited to the flow channel switching member. It may be configured such that the flow channel switching member is fixed, and the integrated flow channel member is driven.
As illustrated in
In addition, a stopper 42, formed in accordance with the shape of the connection port may be provided. It may be enough if the stopper 42 has a function of sealing a liquid, and thus, it is possible to manufacture the stopper 42 with low cost. It may be configured such that a required number of the culture containers are connected to the integrated flow channel member, and the stopper 42 is used for the remaining port.
The culture container can be detached. An application of detaching the culture container for inspection is considered. The stopper may be used after detaching the culture container, and it is preferable when there is a device which is capable of blocking each of the integrated flow channel member side and the culture container side to be aseptically separated at the time of detaching the culture container.
It is preferable that the culture container, the integrated flow channel member, and the flow channel switching member be resin-molded products in terms of cost. Polystyrene, polypropylene, polycarbonate or the like, which is a material to be used for a general culture container, may be used as the material thereof.
As illustrated in
The structure as illustrated in
As described above, it is possible to obtain a different effect as follows in addition to the same effects of Embodiment 1 described above according to the cell culture apparatus according to the present embodiment. For example, a representative effect is that it is possible to prevent the occurrence of erroneous piping with respect to the culture container without taking the man-hours in a case in which the plurality of culture containers are connected in parallel using the culture container set which includes the culture container and the integrated flow channel member enclosing the flow channel switching member. Other effects are as described as in the present embodiment.
A description will be given regarding a cell culture apparatus according to the present embodiment with reference to
A description will be given regarding an example of the cell culture which uses the cell culture apparatus with the closed culture system as described above in the present embodiment. A point that is different from Embodiments 1 to 3 described above will be mainly described also in the present embodiment.
The plurality of culture containers 35-1 to 35-4 are connected to the integrated flow channel member 36 including the flow channel switching member 37 and are installed inside an incubator 47 as an integrated body in
In addition, common flow channels 4 and 7 are connected to the upstream side and the downstream side of the integrated flow channel member 36, and further, are connected with supply bags 2-1 and 2-2, and the collection bag 3, respectively, thereby forming the closed culture system.
As illustrated in
These supply bags 2-1 and 2-2 are formed such that the selection of liquid supply can be made by a switching valve 48. This switching means needs to be a switching means different from the means that has been described hereinbefore in order to completely block a flow channel at a location where the flow of liquid is not desired. For example, there is a method of providing a pinch valve for each flow channel, but a description will not be particularly given regarding the method in the present invention. The supply bags 2-1 and 2-2 may be kept in a cool box 49 in order to keep the quality of content.
The collection bag 3 is not particularly limited in terms of the installation location as long as being the drainage, and may be installed also in the cool box in order to delay degradation during a culture period. In addition, a plurality of the collection bags may be connected so as to separate a recovered material using the switching valve similarly as the switching of the liquid supply of the plurality of supply bags although not illustrated.
A peristaltic pump 50, which is a pump mechanism, is installed in the common flow channel 4 as a driving source of liquid supply. The common flow channel 4 is made of silicone rubber, for example, and is capable of supplying the liquid by squeezing the common flow channel 4 using the elasticity thereof.
A flow channel switching member driving mechanism 51, which is a control mechanism, is provided for switching of the flow channel switching member 37, as a mechanism using, for example, a stepping motor or a servomotor which is capable of changing an orientation and a position of the flow channel switching member 37.
A desired liquid supply is realized by controlling the switching valve 48, the peristaltic pump 50, and the flow channel switching member driving mechanism 51 using a control unit 52 in the above-described configuration of the cell culture apparatus.
Next, a description will be given regarding control at the time of seeding a cell, that is, supplying a cell suspension to the culture container. An example of this control time chart is illustrated in
The flow channel switching member 37 is oriented toward the desired culture container (for example, 35-1) using the flow channel switching member driving mechanism 51. The switching valve 48 selects (turns ON) the supply bag (for example, 2-1) containing the cell suspension, and the liquid supply to the desired culture container 35-1 is started when the peristaltic pump 50 is driven (turned ON) in this state. When the liquid is supplied for a predetermined time in this state, and then the culture container 35-1 is filled with the cell suspension, the flow channel switching member 37 is driven and the liquid supply to the different culture container (for example, 35-2) is started. It is possible to perform the seeding to all the plurality of installed culture containers 35-1 to 35-4 by repeating the above-described control.
Although the content contained in advance is extruded by a liquid to be input from the behind thereof in the present closed culture system, the extruding material is not necessarily the liquid. It may be configured such that a supply bag which contains a sterilized gas is connected instead of the supply bag containing the cell suspension or the culture medium, and the cell suspension or the culture medium contained in advance is pushed out by the gas.
The liquid on the upstream side of the culture container can be used at the subsequent liquid supply, but it is desirable that the amount of such a liquid be minimized as much as possible since the degradation of liquid is accelerated in the state of being placed outside the cool box. When a required amount of liquid is supplied, and the gas is supplied from the behind thereof to extrude the liquid while allowing the liquid to remain inside the cool box, that is, the supply bag as much as possible, the degradation of liquid is little, and it is possible to supply the liquid without waste.
Outside air may be taken in via a HEPA filter instead of the supply bag containing the gas. When a diameter of the HEPA filter is set to be small, it can be regarded as a substantially closed system.
Since the liquid supplied from the cool box is cooled, the liquid is preferably supplied to the culture container after being heated to the temperature that does not affect the culture prior to entering the culture container. A heat source dedicated for heating may be provided, but it is more economic to use the heat of environment inside the incubator. It is preferable to form a common flow channel from the entrance of the incubator to the culture container to be long such that the liquid is heated in a stationary state. Volume tanks may be installed instead of the long common flow channel. An arbitrary stirring means may be provided prior to the supply of liquid to the culture container after performing the heating in the stationary state.
Since the culture surface of the culture container is large as compared to the inflow port and the discharge port, air is likely to remain at the time of supplying the liquid from an empty culture container. The liquid may be supplied while tilting the culture container so as to allow the air to pass without remaining in the culture container.
Since the culture containers 35-1 to 35-4 described in the present embodiment are integrated with the integrated flow channel member 36, it may be enough when the integrated flow channel members 36 is tilted. It is preferable that a tilting mechanism 53 configured for this purpose be simultaneously controlled by the control unit 52 (illustrated in
In addition, a camera 54 is provided in the incubator 47 as illustrated in
Although the plurality of culture containers are provided in the present embodiment, it is desirable to enable the observation of the entire culture container. To do so, the camera 54 needs to be set so as to enable not only a Z-direction control for focusing but also two-dimensional movement in XY directions as illustrated in
As described above, it is possible to obtain a different effect as follows in addition to the same effects of Embodiment 1 described above according to the cell culture apparatus according to the present embodiment. For example, a representative effect is that it is possible to select the liquid to be supplied, to maintain the quality of content, and further, to realize a desired liquid supply in the cell culture using the cell culture apparatus. Other effects are as described as in the present embodiment.
Although the description has been given in detail regarding the invention made by the present inventor based on the embodiments as above, the present invention is not limited to the embodiments, and, of course, can be modified in various ways within a scope not departing from a gist thereof. For example, the above-described embodiments have been described in detail in order to describe the present invention in an easily understandable manner, and are not necessarily limited to one including the entire configuration that has been described above. In addition, some configurations of a certain embodiment can be substituted by configurations of another embodiment, and further, a configuration of another embodiment can be added to a configuration of a certain embodiment. In addition, addition, deletion or substitution of other configurations can be made with respect to some configurations of each embodiment.
The present invention has the following characteristics in relation to the culture container set or the like, for example, in addition to the cell culture apparatus described in the claims.
(1) A culture container set to be used in the cell culture apparatus described in the claim (for example, any one of claims 4 to 12), the culture container set including:
a plurality of culture containers having connection ports; and
an integrated flow channel member having a plurality of ports capable of attaching and detaching the plurality of culture containers,
in which the integrated flow channel member includes common flow channels on an upstream side and a downstream side, a plurality of branching flow channels branching from the respective common flow channels and connected to the plurality of ports, and a flow channel switching member, which has a connecting flow channel therein, and is moved to enable communication between a desired flow channel among the plurality of branching flow channels and the connecting flow channel, being enclosed in a branching portion or a merging portion between the common flow channel and the branching flow channel.
(2) A culture container set to be used in the cell culture apparatus described in the claim (for example, claim 13 or 14), the culture container set including:
a plurality of culture containers having connection ports; and
an integrated flow channel member having a plurality of ports capable of attaching and detaching the plurality of culture containers,
in which the integrated flow channel member includes a common flow channel on an upstream side or a downstream side, a plurality of branching flow channels branching from the common flow channel and connected to the plurality of ports, branching flow channels on the downstream side or the upstream side extending from the plurality of ports, and an enclosed flow channel switching member which has a part exposed to the outside of a closed culture system, a sealing portion provided at a boundary thereof, and a connecting flow channel, which serves a common flow channel function, therein, and is moved to enable communication between a desired flow channel among the plurality of branching flow channels the downstream side or the upstream side and the connecting flow channel.
(3) The culture container set described in the above-described (1) or (2), in which each of the culture containers has the single connection port, and is capable of being attached and detached from a single direction.
(4) The culture container set described in any one of the above-described (1) to (3), in which the culture containers have different sizes of culture surfaces while having the same size of the connection ports.
(5) The culture container set described in any one of the above-described (1) to (3), further including a stopper having the same size as the connection port.
(6) The culture container set described in any one of the above-described (1) to (3), in which the culture container, the integrated flow channel member, and the flow channel switching member are resin-molded articles.
(7) A cell culture apparatus described in the claim (for example, any one of claims 4 to 15), the cell culture apparatus including a pump mechanism which controls a supply amount of a liquid depending on a type of a connected culture container.
(8) The cell culture apparatus described in the claim (for example, any one of claims 4 to 15), the cell culture apparatus including a tilting mechanism to tilt a culture container set including the culture container.
(9) The cell culture apparatus described in the claim (for example, any one of claims 4 to 15), the cell culture apparatus including a driving mechanism to fix the flow channel switching member and to drive the culture container side.
(10) The cell culture apparatus described in the claim (for example, any one of claims 4 to 15), the cell culture apparatus including a two-axis control camera driving mechanism which is controlled along an axis in a horizontal direction and an axis in a height direction.
(11) A culture container set including: a plurality of culture containers having connection ports; and
an integrated flow channel member which includes a plurality of ports capable of attaching and detaching the plurality of culture containers, common flow channels on an upstream side and a downstream side, and a plurality of branching flow channels branching from the respective common flow channels and connected to the plurality of ports,
in which each of the culture containers has the single connection port, and is capable of being attached and detached from a single direction.
(12) The culture container set described in the above-described (11), in which the culture containers have different sizes of culture surfaces while having the same size of the connection ports.
(13) The culture container set described in the above-described (11), further including a stopper having the same size as the connection port.
(14) The culture container set described in the above-described (11), in which the culture container, the integrated flow channel member, and the flow channel switching member are resin-molded articles.
| Number | Date | Country | Kind |
|---|---|---|---|
| 2013-231257 | Nov 2013 | JP | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/JP2014/073073 | 9/2/2014 | WO | 00 |
