Cell recovery method and cell recovery jig

Abstract

This invention provides a cell recovery method and a cell recovery jig that allow for easy cell recovery from cell culture bags. [Solution] A cell recovery method and cell recovery jig for recovering cultured cells from a cell culture bag having a flexible container body and at least one port for injecting and discharging contents, wherein when discharging contents from the container body with the port positioned vertically downward, the container body containing the contents is partitioned into a first chamber having a port for discharging contents and a second chamber not having a port for discharging contents, with the contents distributed into each chamber. After discharging the contents distributed into the first chamber through the port for discharging contents, the partition between the first and second chambers is released, and the contents distributed into the second chamber are transferred to the first chamber and discharged through the port for discharging contents.

Description

Technical Field

[0001] The present invention relates to a cell recovery method and a cell recovery jig.

Background Art

[0002] In recent years, in the fields of pharmaceutical production, gene therapy, regenerative medicine, immunotherapy, etc., it has been required to efficiently culture and induce differentiation of cells (including tissues, microorganisms, viruses, etc.) in a large amount in an artificial environment. In such cell culture, a cell culture bag may be used to avoid the risk of contamination. The cell culture bag is a sterile closed system, and culture can be performed in a state where the contents are not exposed to contaminated outside air.

[0003] For example, Patent Document 1 discloses a bag-shaped culture container in which two multilayer sheets are overlapped and the periphery is sealed.

Prior Art Documents

Patent Documents

[0004]

Patent Document 1

Summary of the Invention

Problems to be Solved by the Invention

[0005] To recover cells from a cell culture bag, one might consider tilting the bag to allow gravity to drain the contents through a port provided on the bag. However, simply tilting the bag so that the port is positioned vertically downwards may leave cells behind. In particular, many cell culture bags block the inflow of air into the bag during draining, making draining difficult. To ensure that no cells remain in the cell culture bag, it is possible to inject air into the bag before or during draining to facilitate the draining of contents. Alternatively, washing with culture medium or similar substances may be performed to recover any remaining cells. However, performing such operations during the cell culture process is time-consuming and therefore undesirable. Cells must be handled aseptically to avoid contamination, requiring that the instruments used for processing, the culture medium and air injected into the cell culture bag be sterile, and that the processing be carried out under sterile conditions such as a clean bench.

[0006] The present invention aims to provide a cell recovery method and a cell recovery jig that allow for easy recovery of cells from a cell culture bag when performing cell culture using a cell culture bag. [Means for solving the problem]

[0007] One embodiment of the present invention is a cell recovery method for recovering cultured cells from a cell culture bag having a flexible container body and at least one port for injecting and discharging contents, wherein when discharging contents from the container body with the port positioned vertically downward, the container body containing the contents is partitioned into a first chamber having a port for discharging contents and a second chamber not having a port for discharging contents, with the contents distributed into each chamber; after discharging the contents distributed into the first chamber through the port for discharging contents, the partition between the first chamber and the second chamber is released, and the contents distributed into the second chamber are transferred to the first chamber and discharged through the port for discharging contents.

[0008] Furthermore, one embodiment of the present invention is a cell recovery jig for recovering cultured cells from a cell culture bag having a flexible container body and at least one port for injecting and discharging contents, the cell recovery jig comprising a partition member that abuts the container body from the outside and brings the inner surfaces of the container body into close contact, thereby dividing the container body into a first chamber having a port for discharging contents and a second chamber not having a port for discharging contents. [Effects of the Invention]

[0009] According to the present invention, a cell recovery method and a cell recovery jig can be provided that allow for easy recovery of cells from a cell culture bag when performing cell culture using a cell culture bag. [Brief explanation of the drawing]

[0010] [Figure 1] This is an explanatory diagram illustrating an example of a cell culture bag used in embodiments of the present invention, where (a) is a plan view, (b) is an enlarged view of (a) along line AA', and (c) is an enlarged cross-sectional view of (b) along line BB'. [Figure 2] This is an explanatory diagram illustrating a schematic of a cell recovery method according to one embodiment of the present invention. [Figure 3]This is an explanatory diagram illustrating a schematic cell recovery method according to another embodiment of the present invention. [Figure 4] This is an explanatory diagram illustrating a schematic of a cell retrieval jig according to one embodiment of the present invention. [Figure 5] This is an explanatory diagram illustrating a schematic method for using a cell retrieval device according to one embodiment of the present invention. [Figure 6] This is an explanatory diagram illustrating a schematic of a cell retrieval jig according to another embodiment of the present invention. [Figure 7] This is an explanatory diagram illustrating a schematic method for using a cell retrieval device according to another embodiment of the present invention. [Figure 8] This is a schematic diagram illustrating another example of a cell culture bag used in embodiments of the present invention. [Modes for carrying out the invention]

[0011] Preferred embodiments of the present invention will be described below with reference to the drawings.

[0012] [Cell culture bag] First, before describing the cell recovery method and cell recovery jig according to this embodiment, an example of the cell culture bag 10 used in this embodiment will be briefly described with reference to Figure 1. Figure 1(a) is a plan view of the cell culture bag 10, (b) is an enlarged view of (a) along line AA', and (c) is an enlarged cross-sectional view of (b) along line BB', which is a schematic explanatory diagram showing the cross-section. Note that the wall thickness of the container body 11 shown in the cross-section of Figure 1(c) is exaggerated.

[0013] The cell culture bag 10 used in this embodiment comprises a flexible container body 11 and at least one port 12 attached to the container body 11 for injecting and discharging the contents. In the illustrated example, the planar shape of the container body 11 of the cell culture bag 10 is substantially rectangular. The size of the container body 11 is not particularly limited, but for example, it can be 50 to 500 mm in length and 50 to 500 mm in width. In the illustrated example, the port 12 is attached to one of the corners of the planar shape of the container body 11.

[0014] The container body 11 can be formed by overlapping plastic films and heat-sealing the peripheral portions, and includes an upper surface 11a and a lower surface 11b.

[0015] On the lower surface 11b, a plurality of recesses 11c each serving as a cell culture part are formed. By providing the plurality of recesses 11c on the bottom surface of the container body, the movement of cells in the cell culture bag 10 can be suppressed, and cell aggregates (also referred to as cell aggregates or spheroids) can be formed. In the cell culture bag 10, since cells tend to gather at the bottom of the recess 11c, as shown in Fig. 1(c), the recess 11c has a spherical crown shape. The recess 11c preferably has an opening diameter (diameter) D of 0.3 to 10 mm and a depth d of 0.1 mm or more in order to suppress the movement of cells in the container body 11 and make the cells in culture stay in one recess 11c.

[0016] As shown in Fig. 1(c), the upper surface 11a has a bulging shape in a stepped form composed of a substantially flat top surface portion 11d covering the entire plurality of recesses 11c and a side wall portion 11e formed around the top surface portion 11d. Thereby, compared with a flat pouch-shaped container formed by simply overlapping two plastic films and sealing the peripheral portions, for example, when the container body 11 is filled with a culture medium in a state where the lower surface 11b is placed in contact with the ground surface, the deformation of the peripheral portion of the lower surface 11b being lifted is suppressed.

[0017] The container body 11 is preferably formed of a plastic film having gas permeability. The gas permeability of the plastic film is such that the oxygen permeability measured at a test temperature of 37°C in accordance with the gas permeability test method of JIS K 7126 is 5000 mL / (m ,

[0017] , , , 2 ,

[0016] , , , ·day·atm) or more. Examples of materials used for the plastic film forming the container body 11 include thermoplastic resins such as polyethylene, polypropylene, ethylene-vinyl acetate copolymer, polyester, silicone-based elastomer, polystyrene-based elastomer, and tetrafluoroethylene-hexafluoropropylene copolymer (FEP). These may be used alone or by laminating the same or different materials. However, considering the heat fusion property when heat-sealing the peripheral portion, it is preferable to have a layer that functions as a sealant layer.

[0018] The port 12 is composed of a tubular member through which a culture medium, cells, etc. can flow, and a tube 13 can be connected thereto. The port 12 can be formed of a thermoplastic resin such as polyethylene, polypropylene, vinyl chloride, polystyrene-based elastomer, FEP, etc.

[0019] [Cell recovery method] Regarding the cell recovery method of the present invention, a preferred embodiment will be described. The cell recovery method of the embodiment described below is a method for recovering the cultured cells C by discharging the contents contained in the container body 11 of the cell culture bag 10 from the port 借 12 by gravity. In the present invention, the term "cell" includes not only single cells but also cell aggregates.

[0020] If the contents can be discharged from the cell culture bag 10 by gravity, it is preferable because there is no need to install and operate a large-scale device equipped with a pump or the like. However, if the contents of the container body 11 are to be discharged by gravity, for example by suspending the cell culture bag 10 so that the port 12 is located vertically below the container body 11, and air inflow into the container body 11 is blocked, there is a risk that the inner surfaces of the film forming the container body 11 will come into contact with each other before all the contents are discharged, causing a part of the container body 11 to become blocked and some of the contents to remain inside the container body 11. Furthermore, when attempting to discharge the contents of the container body 11 by gravity in this way, the settling cells may accumulate on the inner wall of the film, and a concentration distribution may occur within the contents, causing liquids such as the culture medium to be discharged from the cell culture bag 10 before the cells. In this case, the concentration of cells in the culture medium will increase towards the end of the discharge process, and even if the volume of contents remaining in the container body 11 is small, there is a risk that a large number of cells will not be discharged from the container body 11. Furthermore, because the culture medium and other liquids are discharged from the cell culture bag 10 first, there is a risk that cells may get caught on the uneven surface of the inner surface of the container body 11 or around the port 12, resulting in cells not being properly discharged and remaining inside the cell culture bag 10. Similarly, there is a risk that cells may remain inside the tube 13 connected to the port 12 for discharging the contents.

[0021] To solve these problems, it is conceivable to inject air into the container body 11 before discharging the contents from the port 12 to prevent the inner surfaces of the container body 11 from coming into contact with each other as the contents are discharged, or to inject culture medium back into the container body 11 to recover any remaining contents by washing them in. However, the cells need to be handled aseptically. Performing such work would require placing the cell culture bag 10 on an incubator or the like for culturing, then moving it to a sterile space such as a clean bench when discharging the contents, and ensuring that the instruments, culture medium, and air used in the work are sterile, which is time-consuming and therefore undesirable.

[0022] Therefore, in the cell recovery method of this embodiment, the port 12 for discharging contents is positioned vertically downward relative to the container body 11, and in order to discharge the contents from the container body 11 by gravity, the flexible container body 11 containing contents such as culture medium and cells is partitioned into a first chamber 15 where the port 12 for discharging contents is located and a second chamber 16 where the port 12 for discharging contents is not located, with the contents distributed between the two chambers. The contents distributed in the first chamber 15 are discharged through the port 12 for discharging contents, and after the contents distributed in the first chamber 15 have been mostly discharged, the partition between the first chamber 15 and the second chamber 16 is released, and the contents distributed in the second chamber 16 are transferred to the first chamber 15 and discharged through the port 12 for discharging contents. This allows the contents contained inside the cell culture bag 10 to be discharged in two stages, and any cells remaining after the discharge of the contents of the first chamber 15 can be washed away along with the discharge of the contents of the second chamber 16. Therefore, according to this embodiment, it is possible to recover cells with a simple operation, reducing the risk of contamination and decreasing the amount of cells remaining in the cell culture bag, without having to go through steps such as injecting air or culture medium into the container body 11 when discharging the contents.

[0023] 1. First embodiment of a cell harvesting method A preferred cell recovery method according to the first embodiment of the present invention will be described in detail below with reference to Figure 2. Figure 2 is an explanatory diagram illustrating a schematic of a preferred cell recovery method according to this embodiment. Figure 2 shows the cell culture bag 10 suspended so as to hold one diagonal end of the port 12, with the port 12 positioned vertically at the lowest point of the cell culture bag 10. In the following explanation, we will assume the use of the cell culture bag 10 shown in Figure 1. In the following explanation, port 12 is a port for injecting and discharging the contents.

[0024] In the cell recovery method of this embodiment, when discharging the contents from the container body 11, it is preferable to first prevent the contents from being discharged from the container body 11 by, for example, clamping the port 12 or the tube 13 connected to the port 12 with a clip 14, and then tilting the cell culture bag 10 so that the port 12 provided on the cell culture bag 10 is positioned vertically downward relative to the container body 11, and holding it there for a predetermined time (Figure 2(a)). This allows most of the cells C contained within the container body 11 to settle vertically downwards.

[0025] Positioning the port 12 for discharging the contents vertically downward relative to the container body 11 is not limited to the configuration in which the cell culture bag 10 is suspended as shown in the figure. For example, the container body 11 may be tilted at an angle of 30° to 80° with respect to the ground by lifting one end of the cell culture bag 10 on the diagonal side of the port 12, starting from a state where the container body 10 of the cell culture bag 10 is placed on a horizontal surface with the upper surface 11a or lower surface 11b facing downward, so that the port 12 is positioned vertically downward relative to the container body 11.

[0026] Before partitioning the container body 11 into the first chamber 15 and the second chamber 16, the time for holding the container body 11 with the port 12 for discharging contents positioned vertically downward relative to the container body 11 can be appropriately set according to the type and size of the cells, as long as the cells contained within the container body 11 can be allowed to settle vertically downward. For example, cell aggregates tend to settle faster than cells (single cells), and by holding them for about 5 minutes, for example, the cell aggregates on the vertically upward side of the container body 11 will generally settle vertically downward within the container body 11.

[0027] Next, the flexible container body 11 containing the contents is partitioned into a first chamber 15 where the port 12 is located and a second chamber 16 where the port is not located (Figure 2(b)). In this embodiment, when partitioning the container body 11 into the first chamber 15 and the second chamber 16, the contents are distributed into the first chamber 15 and the second chamber 16, respectively. As a result, in the second chamber 16 located vertically above the container body 11, the contents distributed inside can be made to contain almost no cells C.

[0028] To partition the container body 11, the partition member 4 is brought into contact with the container body 11 from the outside, bringing the inner surfaces of the films forming the container body 11 into close contact, thereby preventing fluid connection between the first chamber 15 and the second chamber 16. Furthermore, partitioning the container body 11 while distributing contents between the first chamber 15 and the second chamber 16 can be easily achieved by partitioning the container body 11 while ensuring that the contents are not excessively concentrated in either the area corresponding to the first chamber 15 or the area corresponding to the second chamber 16, preferably while maintaining a generally constant thickness of the container body 11. In the illustrated example, the container body is partitioned into the first chamber 15 and the second chamber 16 with the port 12 for discharging contents suspended vertically downward from the container body, which can be achieved, for example, by using the cell retrieval jig 100 described later. However, the method is not limited as long as the flexible container body 11 containing the contents can be partitioned into a first chamber 15 and a second chamber 16, with the contents distributed between them. For example, a clip that can clamp the container body 11 can be used as the partitioning member 4, and the container body 11 can be partitioned into a first chamber 15 and a second chamber 16 by clamping it from the outside with the clip. In this case, by placing at least a part of the container body 11 on a flat surface or by reducing the angle of inclination of the container body 11 with respect to the horizontal plane, so that the thickness of the container body 11 is not extremely biased towards the first chamber 15 or the second chamber 16, the container body 11 can be easily partitioned by clamping it with the clip, with the contents distributed between the first chamber 15 and the second chamber 16.

[0029] Next, with the container body 11 still partitioned into the first chamber 15 and the second chamber 16, the clips 14 attached to the port 12 and tube 13 are removed, and the contents distributed in the first chamber 15 are naturally discharged by gravity through the port 12 located on the vertically downward side (Figure 2(c)). At this time, it is anticipated that cells may remain on the inner surface of the first chamber 15, near the port 12, and inside the tube 13 as the contents of the first chamber 15 are discharged, causing the inner surfaces of the films forming the container body 11 to come into contact and the container body 11 to become blocked, or cells to accumulate on the inner wall of the films, or the culture medium to be discharged out of the cell culture bag 10 first.

[0030] Once the contents distributed in the first chamber 15 have been largely discharged from the container body 11 of the cell culture bag 10, the partition member 4 is removed to release the partition between the first chamber 15 and the second chamber 16 in the container body 11. This connects the first chamber 15 and the second chamber 16, allowing the contents that were distributed in the second chamber 16 to be transferred to the first chamber 15 and discharged from the port 12 located on the first chamber 15 side (Figures 2(d), (e)).

[0031] 2. Second Embodiment of Cell Recovery Method A preferred cell recovery method according to the second embodiment of the present invention will be described in detail below with reference to Figure 3. Figure 3 is an explanatory diagram illustrating a schematic of a preferred cell recovery method according to this embodiment.

[0032] In the first embodiment, a method of partitioning the cell culture bag 10 into a first chamber 15 and a second chamber 16 was described, such as by suspending the port 12 of the cell culture bag 10 so that it is held at one diagonal end, so that the port 12 is positioned vertically downward relative to the container body 11. In this embodiment, the method of partitioning the first chamber 15 and the second chamber 16 differs from that of the first embodiment.

[0033] In the cell recovery method of this embodiment, when discharging the contents from the container body 11, it is preferable to first prevent the contents from being discharged from the container body 11 by, for example, clamping the port 12 or the tube 13 connected to the port 12 with a clip 14, and then tilting the cell culture bag 10 so that the port 12 provided on the cell culture bag 10 is located vertically downward relative to at least a part of the container body 11, and holding it there for a predetermined time. In the illustrated example, the container body 11 is placed on a nearly flat surface (flat) (Figure 3(a)), and then, while keeping a portion of the container body 11 on the side to which the port 12 for discharging the contents is connected resting on the surface, a portion of the container body 11 of the cell culture bag 10 is lifted by holding one end of the cell culture bag 10 diagonally opposite to the port 12, so that the port 12 on the cell culture bag 10 is positioned vertically below at least a portion of the container body 11, and this position is held for a predetermined time (Figure 3(b)). In this case, when partitioning the container body 11 into a first chamber 15 where the port 12 exists and a second chamber 16 where the port 12 does not exist in a subsequent step, it is preferable to lift a portion of the container body 11 so that at least a portion of the area corresponding to the second chamber 16 is in contact with the inner surfaces of the films forming the container body 11. This allows most of the cells C contained within the container body 11 to be moved to the side to which the port 12 for draining the contents is connected, which in the illustrated example is the side to which the container body 11 is placed on its mounting surface. Here, not only the cells C, but also most of the liquid such as culture medium contained as contents in the container body 11 will be moved to the side to which the port 12 for draining the contents is connected.

[0034] Positioning the port 12 for discharging contents vertically downward relative to at least a portion of the container body 11 is not limited to the method of lifting a portion of the container body 11 of the cell culture bag 10 as shown in the figure. For example, as described in the first embodiment of the cell recovery method, the cell culture bag 10 may be suspended, or the container body 11 may be tilted at an angle of 30° to 80° with respect to the ground surface by lifting one end on the diagonal side of the port 12 for discharging contents of the cell culture bag 10 from a state where the upper surface 11a or lower surface 11b of the container body 11 of the cell culture bag 10 is facing downward, so that the port 12 for discharging contents is positioned vertically downward relative to the container body 11.

[0035] Before partitioning the container body 11 into the first chamber 15 and the second chamber 16, the time for holding the container body 11 with the port 12 for discharging contents positioned vertically downward relative to at least a portion of the container body 11 can be appropriately set according to the type and size of the cells, provided that the cells that have moved to the port-connected side of the container body 11 are allowed to settle onto the inner surface of the film on the ground surface on which the container body 11 is placed (for example, the film forming the bottom surface 11b of the container body 11). For example, as shown in Figure 3, if the container body 11 is placed flat and a portion of the side with the port 12 connected is placed on the mounting surface, and one end of the cell culture bag 10 diagonally opposite the port 12 is held, holding it for a few minutes will cause the cell aggregates in the container body 11 to settle onto the inner surface of the film on the ground surface on the side with the port connected (the first chamber 15 side).

[0036] Next, a rod-shaped partition member 4, whose length spans the container body 11, is placed on a horizontal surface. Then, the container body 11 is slowly lowered onto the partition member 4, ensuring that cells that have settled on the inner surface of the film on the side where the port is connected (the first chamber 15 side) do not float up. The entire container body 11 is then placed on the mounting surface again so that it straddles the partition member 4 (Figure 3(c)). The partition member 4 is positioned so as to contact from below the boundary portion that will divide the container body 11 into the first chamber 15 where the port 12 exists and the second chamber 16 where the port 12 does not exist, preferably the region where cells exist and the region where cells do not exist. In the illustrated example, the partition member 4 is placed so as to contact from below the boundary between the raised portion and the unraised portion of the container body 11, and then the raised portion of the container body 11 is slowly lowered to place the entire container body 11 on the mounting surface again. The partition member 4, which is preferably used in this embodiment, is a rod-shaped member that protrudes less than the thickness of the first chamber 15 containing the contents. For example, if the thickness of the contents of the container body 11 when moved to the side where the port inside the container body 11 is connected (the first chamber 15 side) is 8 to 10 mm, the partition member 4 can be designed to protrude 1 to 8 mm from the mounting surface. By placing such a partition member 4 between the container body 11 and the mounting surface, the partition member 4 pushes up the container body 11 from below, causing the lower surface of the film forming the container body 11 to flex and bulge, thereby dividing the container body 11 into a first chamber 15 where the port 12 exists and a second chamber 16 where the port 12 does not exist. In this embodiment, the partition member 4 only pushes up the container body 11 from below, and since the container body 11 contains contents, the inner surfaces of the films forming the container body 11 are not in complete contact with each other, and the first chamber 15 and the second chamber 16 are at least slightly fluidly connected.

[0037] Here, when the container body 11 is placed back on the mounting surface, the first chamber 15, which had been held for a predetermined time in a position vertically below the second chamber 16, contains not only the cells contained in the container body 11 but also a large amount of culture medium. On the other hand, when the container body 11 is placed back on the mounting surface, the second chamber 16, which had been held for a predetermined time in a position vertically above the first chamber 15, is expected to contain almost no cells and a smaller amount of culture medium compared to the first chamber 15. As mentioned above, simply placing the container body 11 on the mounting surface on which the partition member 4 is installed results in at least a slight fluid connection between the first chamber 15 and the second chamber 16. The liquid culture medium contained in the first chamber 15 moves into the second chamber 16, following the movement of the film as the lifted portion of the container body 11 is slowly lowered, until the container body 11 as a whole is leveled to approximately the same thickness or until the thickness of the first chamber 15 side falls below the protrusion height of the partition member 4 from the mounting surface. The cells, which are part of the contents of the first chamber 15, settle on the bottom side of the container body 11 and will not float up or move from the inner surface of the settled film unless a large liquid flow occurs. Therefore, by slowly and gently lowering the lifted container body 11 over a period of 5 to 10 seconds, for example, without causing any liquid flow to the contents, only the liquid culture medium that was in the first chamber 15 moves to the second chamber 16, leaving the cells on the inner surface of the settled film. Furthermore, the partition member 4, which protrudes from the mounting surface, contacts the boundary between the first chamber 15 and the second chamber 16 from below the container body 11. As a result, the cells, which are part of the contents of the first chamber 15, cannot overcome the portion that is being pushed up by the partition member 4, thus more effectively keeping the cells in the first chamber 15. In this embodiment, the contents distributed inside the second chamber 16 include the culture medium and are in a state where there are almost no cells C present.

[0038] Next, the partition member 4 is lifted upward while maintaining its position relative to the container body 11 (Figure 3(d)). As a result, the container body 11 is lifted up by bending at the partition member 4, and is pushed up from below by the partition member 4. At the same time, the weight of the contents contained inside causes the inner surfaces of the film forming the container body 11 to come into close contact with each other at the point where the partition member 4 contacts. In this embodiment, the first chamber 15 and the second chamber 16 are partitioned in this way so that they are not fluidly connected. The contents are distributed into the first chamber 15 and the second chamber 16 of the container body 11, respectively. The port 12 attached to the end of the container body 11 is generally assumed to be located vertically below the first chamber 15 of the container body 11. However, depending on the arrangement of the partition member 4, the arrangement may be changed so that the port 12 is located vertically below the first chamber 15 of the container body 11, by sliding the container body 11 on the partition member 4, without fluid connection between the first chamber 15 and the second chamber 16.

[0039] Next, with the container body 11 still partitioned so that the first chamber 15 and the second chamber 16 are not fluidly connected, the clips 14 attached to the port 12 and tube 13 are removed, and the contents of the first chamber 15 are naturally discharged from the port 12 by gravity (Figure 3(e)).

[0040] Once the contents distributed in the first chamber 15 have been largely discharged from the container body 11 of the cell culture bag 10, the end of the container body 11 on the second chamber 16 side is lifted so that the second chamber 16 of the container body 11 is positioned vertically above the first chamber 15. This releases the partition between the first chamber 15 and the second chamber 16 in the container body 11, which is made possible by the partition member 4, connecting the first chamber 15 and the second chamber 16, and the contents that were distributed in the second chamber 16 are transferred to the first chamber 15 and discharged from the port 12 located on the first chamber 15 side (Figures 3(f), (g)).

[0041] This embodiment differs from the first embodiment in the points described above, but other configurations are the same as those of the first embodiment, so redundant explanations will be omitted.

[0042] As described above, according to the cell recovery method of the present invention, when recovering cultured cells from a flexible container body 11, even if cells remain on the inner surface of the container body 11, near the port 12, and inside the tube 13 after the contents distributed in the first chamber 15, where the port 12 for discharging contents is located, these cells can be washed away by the contents distributed in the second chamber 16. This reduces the number of remaining cells in the cell culture bag, making it possible to recover cells. In particular, if, in the preceding step, the container body 11 is held for a predetermined time with the port 12 for discharging contents located vertically below at least a part of the container body 11, and then the container body 11 is partitioned into the first chamber 15 and the second chamber 16, the contents distributed in the second chamber 16 can be a culture medium containing almost no cells, thereby further enhancing the effect of washing away cells remaining on the inner surface of the container body 11 and near the port 12 when the contents distributed in the first chamber 15 are discharged. Furthermore, since there is no need to move the cell culture bag 10 to a sterile space for processing, and no need to reinject air or culture medium into the container body 11, it is less time-consuming. Because air or culture medium is not reinjected into the container body 11, the aseptically closed system state is maintained, reducing the risk of contamination.

[0043] [Cell retrieval device] Preferred embodiments of the cell recovery jig of the present invention will now be described. The cell recovery jigs 100 and 200 of the embodiments described below are suitable for use in the cell recovery method described above and are jigs for recovering contents containing cultured cells from a cell culture bag 10 having a flexible container body 11 and at least one port 12 for injecting and discharging the contents. The cell recovery jigs 100 and 200 of the present invention are equipped with a partition member 4 that contacts the container body 11 of the cell culture bag 10 from the outside, bringing the inner surfaces of the container body 11 into close contact, and dividing the container body 11 into a first chamber 15 in which a port 12 for discharging the contents is located and a second chamber 16 in which a port 12 for discharging the contents is not located. As a result, when the contents are discharged from the container body 11 by gravity through the port 12 for discharging the contents, the flexible container body 11 containing the contents such as culture medium and cells is partitioned into a first chamber 15 where the port 12 for discharging the contents is located and a second chamber 16 where the port 12 for discharging the contents is not located, with the contents distributed between the two chambers. The contents distributed in the first chamber 15 are discharged through the port 12 for discharging the contents, and after the contents of the first chamber 15 have been mostly discharged, the partition between the first chamber 15 and the second chamber 16 is released, and the contents distributed in the second chamber 16 are transferred to the first chamber 15 and discharged through the port 12 for discharging the contents.

[0044] 1. First Embodiment of Cell Recovery Fixture The following describes in detail, with reference to Figure 4, a cell recovery jig 100 suitable for use in the cell recovery method described above, particularly in the embodiment described as a preferred example of the first embodiment. Figure 4 is an explanatory diagram illustrating the general structure of the cell retrieval jig 100 according to this embodiment, and is shown as an exploded perspective view showing the configuration of the cell retrieval jig 100 and the cell culture bag 10.

[0045] The cell retrieval jig 100 is a jig for holding a cell culture bag 10 containing cells and culture medium, and comprises a first member 2 and a second member 3 that clamp the container body 11 of the cell culture bag 10, and a partition member 4 that, while clamping the cell culture bag 10, abuts against the container body 11 from the outside, bringing the inner surfaces of the container body 11 into close contact, and divides the container body 11 into a first chamber 15 having a port 12 for discharging the contents and a second chamber 16 not having a port 12 for discharging the contents. In Figure 4, other components are omitted as appropriate in order to show the relationship between the cell culture bag 10, the first component 2, the second component 3, and the partition component 4, which will be explained below.

[0046] The first member 2 and the second member 3 are arranged facing each other at a predetermined distance, and are configured to sandwich the container body 11 of the cell culture bag 10 between them.

[0047] In the illustrated example, the first member 2 and the second member 3 are plate-like members having a substantially rectangular planar shape larger than the area of ​​the upper surface 11a or lower surface 11b of the container body 11 of the cell culture bag 10. Each of them has a contact surface 3a or contact surface 2a that contacts the upper surface 11a or lower surface 11b of the container body 11 of the cell culture bag 10. The first member 2 and / or the second member 3 may be partially or entirely transparent so that the internal conditions of the cell culture bag 10 can be observed. In that case, such parts can be formed from, for example, highly transparent synthetic resin or glass.

[0048] The first member 2 and the second member 3 are designed to hold the cell culture bag 10 containing its contents at a distance such that the cell culture bag 10 has a predetermined thickness. The distance between the first member 2 and the second member 3 is not limited and can be appropriately designed according to the thickness of the cell culture bag 10, etc.

[0049] Specifically, the first member 2 and the second member 3 can be opened to approximately 90° relative to each other using, for example, hinges or latches (not shown), and can be closed and fixed so that their contact surfaces 2a and 3a are substantially parallel to each other with a predetermined distance between them. With this structure, the first member 2 and the second member 3 are opened relative to each other, and for example, the first member 2 is placed on the ground surface with the first member 2 facing downwards. The cell culture bag 10 is then placed so that the lower surface 11b of the container body 11 of the cell culture bag 10 is in contact with the contact surface 2a of the first member 2. After that, the first member 2 and the second member 3 are closed relative to each other, thereby making it possible to sandwich the cell culture bag 10 between the first member 2 and the second member 3.

[0050] In this embodiment, the partition member 4 consists of an opening 41 that opens to a portion of the first member 2 or the second member 3, and a lid portion 42 that is detachably formed on the opening 41. In the illustrated example, the partition member 4 is provided on the second member 3.

[0051] The opening 41 is formed in a range corresponding to the second chamber 16 of the first member 2 or the second member 3. In the illustrated example, the container body 11 of the cell culture bag 10 has a roughly rectangular shape in plan view, and the port 12 is located at one of the four corners of the container body 11's plan view. For a cell retrieval jig 100 that holds such a cell culture bag 10, the opening 41 can be designed so that a portion of the area including the corner of the container body 11 located diagonally opposite the port 12 of the cell culture bag 10 held inside the cell retrieval jig 100 is partitioned off as a second chamber 16.

[0052] When the lid portion 42 is attached to the first member 2 or the second member 3, it closes the opening 41 and forms a contact surface 2a or contact surface 3a as part of the first member 2 or the second member 3. When the lid portion 42 is removed from the first member 2 or the second member 3, an opening 41 is formed in the first member 2 or the second member 3.

[0053] Furthermore, it is preferable that the cell retrieval jig 100 includes a holding means 5 that maintains the port 12 of the held cell culture bag 10 so that it is positioned vertically downward relative to the container body 11. In the illustrated example, the cell retrieval jig 100 has a suspension hole 51 on the edge diagonally opposite to the area where the port 12 of the cell culture bag 10 is located, for suspending the cell retrieval jig 100 together with a hook or the like, and this suspension hole 51 is used as a holding means 5.

[0054] When using a cell retrieval jig 100 equipped with such suspension holes 51, the cell culture bag 10 may also be provided with holes drilled at positions corresponding to the suspension holes 51 of the cell retrieval jig 100. With a cell culture bag 10 equipped with such holes, the cell retrieval jig 100 and the cell culture bag 10 held inside it can be suspended together using hooks or the like, thereby ensuring stable holding of the cell culture bag 10 inside the cell retrieval jig 100 without the risk of it sliding down when the cell culture bag 10 is tilted. However, the cell culture bag 10 does not need to have such holes, as long as the container body 11 can be held in place without slipping down between the first member 2 and the second member 3 of the cell retrieval jig 100. For example, the container body 11 may be fixed in place so that its edge is sandwiched between the periphery of the first member 2 and the periphery of the second member 3, preventing it from slipping down due to the inclination or suspension of the cell retrieval jig 100. In this case, the periphery of the first member 2 and the periphery of the second member 3 may protrude from each other, and the periphery of each member may be in contact with each other when the contact surfaces 2a and 3a are closed and fixed so that they are substantially parallel and facing each other with a predetermined distance between them.

[0055] A partitioning method for dividing the container body 11 of the cell culture bag 10 into a first chamber 15 and a second chamber 16 using the partition member 4 of the cell retrieval jig 100 according to this embodiment will be explained with reference to Figure 5. In the illustrated example, the partition member 4 is provided on the second member 3, and here, the explanation will be given using the configuration in which the partition member 4 is provided on the second member 3 as an example. It goes without saying that the container body 11 can be partitioned in the same way even if the partition member 4 is provided on the first member 2.

[0056] First, the container body 11 of the cell culture bag 10, which contains the contents, is sandwiched between the first member 2 and the second member 3 so that the port 12 of the cell culture bag 10 is located diagonally opposite to the suspension hole 51 of the cell retrieval jig 100. The cell retrieval jig 100 is then suspended using the suspension hole 51 while the cell culture bag 10 is held between the first member 2 and the second member 3 (Figure 5(a)).

[0057] Next, the lid portion 42 of the second member 3 is removed to form an opening 41 in the second member 3 (Figure 5(b)).

[0058] Next, a portion of the container body 11 of the cell culture bag 10, held between the first member 2 and the second member 3 of the cell retrieval jig 100 (the area that will become the second chamber 16), is pulled out from the opening 41 to the outside of the cell retrieval jig 100, and the container body 11 is folded outwards from the second member 3 along the lower side 41a that defines the opening 41 (Figure 5(c)).

[0059] Next, the lid portion 42 is reattached to the second member 3 by sandwiching the two films that form the container body 11 (Figure 5(d)). As a result, one side 41a of the opening 41 and the corresponding side 42a of the lid portion 42 abut against the container body 11 from the outside, bringing the inner surfaces of the container body 11 into close contact. The portion that extends outside the cell retrieval jig 100 from the opening 41 becomes the second chamber 16, thus dividing the container body 11 into the first chamber 15 and the second chamber 16.

[0060] To release the partition by the partition member 4 and connect the first chamber 15 and the second chamber 16, the lid 42 that was holding the container body 11 should be removed, and a portion of the container body 11 that had been pulled out to the outside of the cell recovery jig 100 should be pushed into the space between the first member 2 and the second member 3 of the cell recovery jig 100. If necessary, the lid 42 may be reattached to the second member 3 without holding the container body 11 to close the opening 41.

[0061] According to the cell retrieval jig 100 of this embodiment, a cell culture bag 10 having a flexible container body 11 and a port 12 for discharging contents is held, and when discharging contents from the container body 11 with the port 12 positioned vertically downward, the container body 11 containing the contents is partitioned into a first chamber 15 where the port 12 for discharging contents is located and a second chamber 16 where the port 12 is not located, with the contents distributed between the two chambers. After discharging the contents distributed in the first chamber 15 through the port 12, the partition between the first chamber 15 and the second chamber 16 is released, and the contents distributed in the second chamber 16 are transferred to the first chamber 15 and discharged through the port 12. Furthermore, by clamping the cell culture bag 10 having a flexible container body 11 between the first member 2 and the second member 3, the thickness of the container body 11 can be kept approximately constant even when the cell culture bag 10 is suspended or tilted. This helps to prevent the contents from shifting to the lower vertical side when the cell culture bag 10 is tilted, which could cause some of the inner surfaces of the film forming the container body 11 to come into contact and become blocked, or cause the film to bend and wrinkle. Furthermore, by keeping the thickness of the container body 11 of the cell culture bag 10 constant, the process of dividing the container body 11 into a first chamber 15 and a second chamber 16 with the contents distributed into each can be made easier. In addition, by keeping the thickness of the container body 11 of the cell culture bag 10 constant, when dividing the container body 11 into a first chamber 15 and a second chamber 16 with the contents distributed into each, the volume of each can be kept relatively constant to avoid variations from one operation to the next, or the approximate volumes of the first chamber 15 and the second chamber 16 can be easily determined.

[0062] 2. Second embodiment of the cell retrieval device The following describes in detail, with reference to Figure 6, a cell recovery jig 200 suitable for use in the cell recovery method described above, particularly the embodiment described as a preferred example of the second embodiment. Figure 6 is an explanatory diagram illustrating the general structure of the cell retrieval jig 200 according to this embodiment. Figure 6(a) shows one usage state of the cell retrieval jig 200, and Figure 6(b) shows another usage state of the cell retrieval jig 200.

[0063] The cell retrieval jig 200 is a jig for placing a cell culture bag 10 containing cells and culture medium, and comprises a mounting plate 6 for placing the container body 11 of the cell culture bag 10, and a partition member 4 installed across the mounting plate 6, which pushes the container body 11 up from below to divide it into a first chamber 15 having a port 12 for discharging the contents and a second chamber 16 not having a port 12 for discharging the contents.

[0064] In the illustrated example, the mounting plate 6 is a plate-like member having a substantially rectangular planar shape larger than the area of ​​the upper surface 11a or lower surface 11b of the container body 11 of the cell culture bag 10. The mounting plate 6 has a contact surface 6a that is in contact with the upper surface 11a or lower surface 11b of the container body 11 of the cell culture bag 10.

[0065] The partition member 4 is preferably a rod-shaped member that protrudes 1 to 8 mm from the mounting plate 6.

[0066] In the illustrated example, the mounting plate 6 is composed of a first mounting portion 61 and a second mounting portion 62, with a partition member 4 in between (Figure 6(a)). The first mounting portion 61 and the second mounting portion 62 each have contact surfaces 61a and 62a that contact the upper surface 11a or lower surface 11b of the container body 11. The second mounting portion 62 is rotatable around the partition member 4 so that its inclination angle with respect to the first mounting portion 61 can be changed. It is preferable that the second mounting portion 62 be structured to be rotatable until the opposite surface of its contact surface 62a contacts the opposite surface of the contact surface 61a of the first mounting portion 61, as shown in Figure 6(b). Furthermore, it is even more preferable that the second mounting portion 62 is structured to be able to rotate around the partition member 4 such that the angle between the first mounting portion 61 and the second mounting portion 62, with the partition member 4 as the apex, is such that the region sandwiched between the contact surfaces 61a and 62a has a less favorable angle (i.e., on the opposite side from the state shown in Figure 6(b)). Such a second mounting portion 62 can be fixed to the partition member 4 by, for example, a hinge (not shown).

[0067] After placing the container body 11 on the mounting plate 6, the second mounting portion 62 is rotated around the partition member 4 so that the angle between the first mounting portion 61 and the second mounting portion 62, with the partition member 4 as the apex, is such that the area sandwiched between the contact surfaces 61a and 62a has a dominant angle, causing the second mounting portion 62 to be inclined relative to the first mounting portion 61. As a result, the flexible container body 11 placed on the mounting plate 6 bends at the partition member 4, is pushed up from below by the partition member 4, and both ends of the container body 11 are pulled by the weight of the contents contained inside. Consequently, at the portion where the partition member 4 abuts from below, the inner surfaces of the film forming the container body 11 come into close contact, making it possible to partition the first mounting portion 61 side and the second mounting portion 62 side in a state where fluid connection is generally not established.

[0068] The cell retrieval jig 200 preferably includes a holding means 5 for supporting and lifting the partition member 4. In the illustrated example, the cell retrieval jig 200 includes a holding means 5 extending from the edge of the mounting plate 6 near the partition member 4. More specifically, the holding means 5 shown in the illustration is connected to the end of the mounting plate 6 in the region from a part of the first mounting portion 61 to the partition member 4 on one side of the mounting plate 6 that includes the first mounting portion 61 and the second mounting portion 62, and in the region of the second mounting portion 62, it extends parallel to the end of the second mounting portion 62 without being connected. With this holding means 5, the user can support and lift the partition member 4 by pinching the holding means 5 of the cell retrieval jig 200 placed on the mounting surface with their fingers or the like. More specifically, when the cell retrieval jig 200 is lifted by the holding means 5 shown in the figure, the first mounting part 61 connected to the holding means 5 tilts upward from the horizontal surface on which it was placed, as the free end side of the holding means 5, which is on the opposite side of the first mounting part 61, is lifted vertically upward. At this time, the partition member 4, which is located on the free end side of the holding means 5 relative to the first mounting part 61, is lifted by the holding means 5 so that it is positioned vertically upward relative to the first mounting part 61. On the other hand, as the partition member 4 is lifted vertically upward, the second mounting part 62 rotates around the partition member 4 and gradually tilts so that the partition member 4 side is vertically upward. In other words, the second mounting portion 62 is gradually tilted relative to the first mounting portion 61 so that the angle between the first mounting portion 61 and the second mounting portion 62, with the partition member 4 as the apex, is such that the region sandwiched between the contact surfaces 61a and 62a has a dominant angle. In the illustrated example, when the cell retrieval jig 200 is completely separated from the mounting surface, the second mounting portion 62 can rotate around the partition member 4 until it is approximately parallel to the vertical direction. The first mounting portion 61 is approximately parallel to the vertical direction when the holding means 5 is held so that it is approximately parallel to the vertical direction.

[0069] Furthermore, the holding means 5 shown in the figure is equipped with a suspension hole 51 at the free end on the tip side extending from the end of the mounting plate 6. The cell retrieval jig 200 can be suspended by suspending the suspension hole 51 of such a holding means 5 using a hook or the like.

[0070] However, the holding means 5 is not limited to the illustrated example, as long as it is capable of supporting and lifting the partition member 4. For example, suspension holes may be provided at both ends of the partition member 4, and each suspension hole may be simultaneously suspended by a string. Alternatively, the holding means 5 may be structured to support the partition member 4 from below so as to lift it to a predetermined height.

[0071] Furthermore, it is preferable that the cell retrieval jig 200 is equipped with fixing means 7 such that the contact surface 61a of the first mounting portion 61 and the contact surface 62a of the second mounting portion 62 are fixed in a state where they are substantially flush. In the illustrated example, the cell retrieval jig 200 is equipped with a fixing means 7 on the free end side of the holding means 5 for fixing the holding means 5 and the second mounting portion 62. By fixing the holding means 5 and the second mounting portion 62 with the fixing means 7, the first mounting portion 61 and the second mounting portion 62 connected to the holding means 5 can be fixed in a state where they are almost flush with each other.

[0072] A partitioning method for dividing the container body 11 of the cell culture bag 10 into a first chamber 15 and a second chamber 16 using the cell retrieval jig 200 according to this embodiment will be explained with reference to Figure 7.

[0073] First, the cell retrieval jig 200 is placed on a horizontal surface, and the cell culture bag 10 is placed on the mounting plate 6 with the first mounting section 61 and the second mounting section 62 almost flush (Figure 7(a)). Here, the part of the cell culture bag 10 with a port 12 for discharging the contents is placed on the side of the first mounting section 61.

[0074] Next, with the cell retrieval jig 200 in place, and the side of the cell culture bag 10 with the port 12 still on the mounting plate 6 (first mounting section 61) of the cell retrieval jig 200, lift one end of the cell culture bag 10 on the diagonal side of the port 12 ((Figure 7(b)). This allows the port 12 to be positioned vertically downward relative to at least a portion of the lifted container body 11. Alternatively, the angle between the first mounting section 61 and the second mounting section 62, with the partition member 4 as the apex, can be adjusted so that the area sandwiched between the contact surfaces 61a and 62a has a less favorable angle. If the mounting portion 62 is rotatable around the partition member 4, the second mounting portion 62 may be lifted and tilted together with the container body 11. That is, the angle between the first mounting portion 61 and the second mounting portion 62, with the partition member 4 as the apex, is adjusted so that the angle of the region sandwiched between the contact surfaces 61a and 62a is 90° to 120° by rotating the second mounting portion 62 around the partition member 4 and tilting it relative to the first mounting portion 61. This allows the port 12 placed on the first mounting portion 61 to be positioned vertically downward relative to the container body 11 placed on the second mounting portion 62.

[0075] Next, after a predetermined time has elapsed, the diagonal end of the port 12 of the cell culture bag 10 that had been lifted is slowly lowered, and the entire cell culture bag 10 is placed back on the mounting plate 6 (Figure 7(c)). Alternatively, the second mounting section 62 that had been lifted is slowly lowered, and the cell retrieval jig 200 is returned to a state where the first mounting section 61 and the second mounting section 62 are almost flush.

[0076] Next, at least a portion of the container body 11 diagonally opposite to the port 12 is fixed to the second mounting portion 62, and then the cell retrieval jig 200 is lifted using the holding means 5. The means for fixing the container body 11 to the second mounting portion 62 is not particularly limited, as long as it is fixed in a way that prevents the container body 11 from sliding down when the second mounting portion 62 is tilted vertically upward relative to the first mounting portion 61 in a subsequent step. For example, the container body 11 and the second mounting portion 62 may be fixed using clips. If the cell culture bag 10 has a hole drilled near the peripheral portion diagonally opposite to the port 12 of the container body 11, a fixing device (not shown) for locking the hole at the corresponding position on the second mounting portion 62 may be provided. By locking the hole of the cell culture bag 10 with the fixing device, the cell culture bag 10 can be fixed to the second mounting portion 62 in a way that prevents it from sliding down inside the cell retrieval jig 100 when the cell culture bag 10 is tilted. By lifting the cell retrieval jig 200 using the holding means 5, the partition member 4 is separated from the horizontal surface on which the cell retrieval jig 200 was placed by the holding means 5, and the mounting plate 6 gradually tilts so that the first mounting portion 61 and the second mounting portion 62 are on the vertically upward side toward the partition member 4. (Figure 7(d)) When the cell retrieval jig 200 is lifted up by suspending the suspension hole 51 of the holding means 5 with a hook or the like, until it is completely separated from the horizontal surface on which it was placed, the first mounting part 61 and the second mounting part 62 become approximately parallel to the vertical direction ((Figure 7(e)). As a result, the partition member 4 presses against the container body 11 that was placed on the mounting plate 6 from below and pushes it up, and the weight of the contents inside the container body 11 brings the inner surfaces of the container body 11 into close contact, thereby partitioning the container body 11 into a first chamber 15 and a second chamber 16.

[0077] To release the partition by the partition member 4 and connect the first chamber 15 and the second chamber 16, the first mounting portion 61 is positioned vertically below the second mounting portion 62, and the second mounting portion 62 on which the second chamber 16 is mounted is rotated around the partition member 4 and lifted up so that the first mounting portion 61 and the second mounting portion 62 are flush, and then fixed in that state using the fixing means 7 ((Figure 7(f)). According to the cell retrieval jig 200 shown, by fixing the second mounting portion 62 to the fixing means 7, the first mounting portion 61 is positioned vertically below the second mounting portion 62, and the first mounting portion 61 and the second mounting portion 62 are fixed in a state where they are almost flush.

[0078] According to the cell recovery jig 200 of this embodiment, a cell culture bag 10 having a flexible container body 11 and a port 12 for discharging contents is placed on the jig, and when discharging contents from the container body 11 with the port 12 positioned vertically downward, the container body 11 containing the contents is partitioned into a first chamber 15 where the port 12 for discharging contents is located and a second chamber 16 where the port 12 for discharging contents is not located, with the contents distributed between the two chambers. After discharging the contents distributed in the first chamber 15 through the port 12 for discharging contents, the partition between the first chamber 15 and the second chamber 16 is released, and the contents distributed in the second chamber 16 are transferred to the first chamber 15 and discharged through the port 12 for discharging contents. Furthermore, when the container body 11 is held for a predetermined time with the port 12 for discharging the contents positioned vertically downward relative to at least a part of the container body 11, and then the container body 11 is partitioned into a first chamber 15 and a second chamber 16 with the contents distributed to each, it is possible to suitably avoid the culture medium being unevenly distributed to the first chamber 15.

[0079] Although the present invention has been described above with reference to preferred embodiments, it goes without saying that the present invention is not limited to the embodiments described above, and various modifications can be made within the scope of the present invention.

[0080] For example, in the embodiments described above, a cell culture bag 10 was given in which a plurality of recesses are formed on the lower surface 11b of the container body 11 of the cell culture bag 10. However, the cell culture bag to which the cell culture method and cell recovery jig of the present invention are applied is not limited to this. For example, the shape of the lower surface 11b of the cell culture bag 10 may be planar, similar to the upper surface 11a.

[0081] Furthermore, in the embodiments described above, an example of a cell culture bag 10 was given in which the planar shape of the container body 11 is substantially rectangular, but the planar shape of the container body 11 of the cell culture bag 10 is not limited to this. For example, the planar shape of the container body 11 can be various shapes such as a square, a polygon such as a hexagon, an ellipse, or a circle. The port 12 for discharging the contents is preferably provided on the container body 11 so that it can be easily positioned vertically downward relative to the container body 11 when the cell culture bag 10 is tilted, but it goes without saying that the position of the port 12 can be appropriately designed depending on the planar shape of the container body 11.

[0082] Furthermore, in the above-described embodiment, an example of the cell culture bag 10 was given in which the planar shape of the container body 11 is substantially rectangular and a port 12 is attached to one of the corners of the planar shape. However, the attachment position and number of ports 12 are not limited to this. For example, as shown in Figure 8, the planar shape of the container body 11 may be substantially rectangular, and ports 12 may be attached to each corner on the same long side of the planar shape. If there are multiple ports, the port for discharging the contents should be treated in the same way as the port 12 described in the above-described embodiment when retrieving cells.

[0083] Furthermore, while the first embodiment of the cell retrieval jig described above included a configuration in which the partition member 4 has an opening 41 and a lid 42, the specific configuration of the partition member 4 is not limited as long as it has a structure that can divide the container body 11 into a first chamber 15 and a second chamber 16. For example, the partition member 4 may be configured such that one of the first member 2 or the second member 3 has a slit opening in a region corresponding to the area where the partition is provided, and a partition piece that can protrude from the slit toward the cell culture bag 10 held by the cell retrieval jig 100. With such a configuration, by making the partition piece protrude from the slit toward the cell culture bag 10, it can contact the container body 11 from the outside, pressing and sandwiching the container body 11 against the second member 3 or the first member 2 facing the slit, bringing the inner surfaces of the container body 11 into close contact and partitioning the container body 11 into a first chamber 15 and a second chamber 16.

[0084] Furthermore, in the first embodiment of the cell retrieval jig described above, a suspension hole 51 was given as the holding means 5. However, the specific configuration of the holding means 5 is not limited to a suspension hole 51, as long as it can hold the cell culture bag 10 so that the port 12 for discharging the contents is located vertically downward relative to the container body 11. For example, in order to tilt the container body 11 at an angle of inclination of 30° to 80° with respect to the ground surface so that the port 12 for discharging the contents is located vertically downward relative to the container body 11, a support structure such as an inclined plate that supports the container body 11 in an inclined state at a predetermined angle may be provided as the holding means 5. When such a holding means is provided, the cell retrieval jig 100 may be equipped with fixing devices on the first member 2 and / or the second member 3 for engaging the hole drilled near the periphery of the cell culture bag 10. By securing the holes of the cell culture bag 10 with a fixing device, it is preferable that the cell culture bag 10 can be held stably within the cell retrieval jig 100 without the risk of it sliding down when the cell culture bag 10 is tilted. [Explanation of Symbols]

[0085] 100,200 Cell Retrieval Fixture 2. First component 3. Second component 4. Partition members 5 Holding means 6 Mounting plate 10 cell culture bags 11 Container body 12 ports

Claims

1. A cell recovery method for recovering cultured cells from a cell culture bag having a flexible container body and at least one port for injecting and discharging contents, When discharging the contents from the container body by positioning the port vertically downward, the container body containing the contents is partitioned into a first chamber where a port for discharging the contents exists and a second chamber where no port for discharging the contents exists, with the contents distributed between the two chambers. After the contents distributed in the first chamber are discharged through the port for discharging the contents, the partition between the first chamber and the second chamber is released. The contents distributed in the second chamber are transferred to the first chamber and discharged through the port for discharging the contents. A method for recovering cells characterized by the following features.

2. The cell recovery method according to claim 1, wherein the container body is held for a predetermined time with the port positioned vertically downward with respect to at least a part of the container body, and then the container body is partitioned into the first chamber and the second chamber.

3. A cell retrieval jig for retrieving cultured cells from a cell culture bag having a flexible container body and at least one port for injecting and discharging contents, A partition member that contacts the container body of the cell culture bag from the outside, bringing the inner surfaces of the container body into close contact, thereby dividing the container body into a first chamber having a port for discharging the contents and a second chamber not having a port for discharging the contents. A cell retrieval jig characterized by having the following features.

4. The cell retrieval jig according to claim 3, comprising a first member and a second member arranged opposite to each other at a predetermined interval, for clamping the container body of the cell culture bag.

5. The cell recovery jig according to claim 4, further comprising a holding means for holding the cell culture bag such that the port for discharging the contents is located vertically downward relative to the container body.

6. The cell recovery jig according to claim 4 or 5, wherein the partition member includes an opening in the first member or the second member that opens to a range corresponding to the second chamber, and a lid portion that is detachably formed on the opening.

7. It is equipped with a mounting plate on which to place cell culture bags, The cell recovery jig according to claim 3, wherein the partition member is installed so as to span the aforementioned mounting plate and is configured to push up the container body from below, thereby dividing it into the first chamber and the second chamber.

8. The cell retrieval jig according to claim 7, wherein the mounting plate is composed of a first mounting portion and a second mounting portion sandwiching the partition member, and the second mounting portion is rotatably mounted around the partition member.

9. The cell retrieval jig according to claim 7 or 8, further comprising a holding means for supporting and lifting the partition member.

Images