TW201829766A - Filter-equipped bag for cells and production method therefor - Google Patents

Abstract

A container body 2 is formed by sandwiching, between flexible films 20 each of which includes at least a polyethylene layer 20a and which are disposed so as to have the respective polyethylene layers 20a facing each other, a filter member 4 comprising: a polyester or polyamide main mesh 4a having openings of 20-300 [mu]m; and polyethylene submeshes 4b which have openings of 100-4000 [mu]m and which are disposed on the front and back surfaces of the main mesh, and then by heat-sealing the flexible films at the periphery thereof. In a heat-sealed part 5 formed at the periphery of the container body 2, the submeshes 4b are melted so as to be fusion-bonded to each other while infiltrating voids in the main mesh 4a, and to be fusion-bonded with the polyethylene layers 20a of the flexible films 20. This configuration is able to enhance the sealing strength with the filter member when the interior of the container body is to be partitioned by the filter member so as to allow for a simple and efficient way of performing various operations necessary to carry out cell culturing, such as fractionation, filtration, and washing of cells.

Description

Cell bag with filter, and manufacturing method thereof

[0001] The present invention relates to a filter-attached cell bag for performing various operations such as division, filtration, and washing of cells or culture cells which are carried out in a simple and efficient manner, and a method for producing the same.

[0002] In recent years, in the pharmaceutical production, gene therapy, regenerative medicine, immunotherapy and other medical science. In the field of biochemistry, it is required to efficiently culture cells (including tissues, microorganisms, viruses, etc.) in an artificial environment. [0003] In order to cope with such requirements, the present applicant has repeatedly reviewed a cell culture system that can effectively perform cell culture while constructing a occlusion system environment and reducing the risk of contamination. [0004] For example, Patent Document 1 proposes a kit for cell culture, which is a culture container for culturing cells, a medium storage container for storing a medium, a cell for injecting cells, and a recovery culture. The cell recovery container of the post cell suspension is formed by a catheter connection to form a closed system environment. According to the cell culture kit, after the cells are injected, the cells can be added to the inside of the kit to be added to the medium, sampled, and recovered. [Prior Art Document] [Patent Document] [0005] [Patent Document 1] International Publication No. 2013/114845

[Problem to be Solved by the Invention] [0006] However, Patent Document 1 discloses an example in which, when the cultured cells are recovered, the culture container is allowed to stand, and the cells in the cell suspension are allowed to settle, and the cell suspension is discharged. After the liquid is reduced, the concentrated cell suspension is transferred from the culture container to the cell recovery container after the liquid amount is reduced. Further, in the cell culture, since it usually takes several days to several weeks, it is necessary to replace the culture medium as needed, and it is necessary to replace the medium as needed during the culture period without hindering the growth of the cell metabolite. When the medium is replaced, it is considered to remove the old medium by discharging the supernatant from the cell suspension, and a new medium is added instead. [0008] However, when performing cell recovery and medium exchange, the culture vessel must be allowed to stand before the cell suspension is discharged, and the cells in the cell suspension are allowed to settle until time before the cells settle. On the other hand, even if it takes a sufficient time to cause the cells in the cell suspension to settle, there is a case where the cells are mixed into the supernatant liquid by the discharge operation, and the cells are discharged together with the supernatant liquid. In particular, when the medium is replaced, it is desirable to remove the old medium as much as possible, and to add more new medium. However, the more the amount of the supernatant is discharged, the more the cells are prevented from entering. Further, when the medium is exchanged, in order to prevent the cells from being discharged together with the medium, it is also conceivable to install a filter in the flow path provided in the port of the culture container. However, since the flow path of the port is usually about 1 to 10 mm, and the cross-sectional area of the filter is relatively small, it is easy to cause the filter to be clogged by the captured cells, and there is a hindrance to the discharge of the old medium. Furthermore, there is also a cell that is captured by the filter does not return to the culture container, but is killed by the state captured by the filter. [0010] In view of these technical backgrounds, the Applicant considers that the interior of the container body is partitioned by the filter member, for example, when the medium is replaced, the cells in the culture are residing in one zone, and only the old zone is discharged from the other zone. The medium was subjected to various reviews in order to achieve this repetition. [0011] However, when used as a filter member, a sieve having a smaller mesh size to the extent that the cells are not allowed to pass is preferably a polyester or polyamide screen which can be easily obtained. When a container formed of a flexible material made of a polyethylene resin is used as the culture container of Patent Document 1, there is a problem that the sealing strength is weak. Therefore, in view of this problem, the applicant has repeated the results of active research and finally completed the present invention. [0012] In other words, the present invention provides various operations such as cell division, filtration, and washing performed in a cell culture in a simple and efficient manner, and aims to provide an improvement filter when the inside of the container body is partitioned by the filter member. A cell bag with a filter attached to the seal strength of the member, and a method of manufacturing the same. [Means for Solving the Problem] [0013] The cell bag with a filter according to the present invention includes a container body and a port for injection and discharge, and the inside of the container body is divided by a filter member, and is disposed in at least one division There is a port for injecting and discharging, and the filter member is made of a polyester mesh made of polyester of 20 to 300 μm or a polyamide mesh and a polyethylene of 100 to 4000 μm which is disposed on both sides of the front and back sides thereof. In the sub-screen, the container body is formed between the flexible films including at least the polyethylene layer, and the peripheral portion is thermally fused by sandwiching the filter member with the polyethylene layer, and the container body is In the heat fusion portion formed in the peripheral portion, the polyethylene sub-screen is melted, invades into the gap portion of the main screen, and is fused to each other, and is fused to the polyethylene layer of the flexible film. Further, the method for manufacturing a cell bag with a filter according to the present invention includes a container body and a port for injecting and discharging, wherein the inside of the container body is divided by a filter member, and the injection is provided in at least one division. The method for manufacturing a cell bag with a filter for the outflow port is configured to be a mesh of 100 to 4000 μm on the front and back sides of a mesh of 20 to 300 μm of a mesh or a polyamide mesh. a secondary screen made of ethylene, wherein the filter member is prepared, and between the flexible films including at least a polyethylene layer, the filter member is sandwiched by the polyethylene layer, and the peripheral portions are heated. Fusion. [Effect of the Invention] According to the present invention, it is possible to provide a cell bag with a filter which is formed by a flexible film forming a container body and a filter member which has improved sealing strength and improved bag breaking strength.

[0017] Hereinafter, preferred embodiments of the present invention will be described with reference to the drawings. 1 is an explanatory view showing an example of a cell bag with a filter according to an embodiment of the present invention, and FIG. 1(a) is a plan view of a cell bag 1 with a filter attached thereto, and FIG. 1(b) is attached. A side view of the cell bag 1 having a filter, and Fig. 1(c) is a cross-sectional view taken along line AA of Fig. 1(a). [0018] As a filter example, the cell bag 1 with a filter shown in FIG. 1 is provided with a container body 2 and injection/discharge ports 3a and 3b. Further, the inside of the container body 2 is divided into two chambers by the filter member 4, and injection/discharge ports 3a and 3b are provided in each of the divisions (the first division 1st and the second division 2nd). [0019] The injection/outlet ports 3a and 3b are provided at the entrance and exit of the medium or the cells during injection or discharge, and can be attached to the container body 2 by, for example, a tubular member through which a medium or a cell can flow. The tubular member forming the injection/outlet ports 3a and 3b is formed by, for example, injection molding, extrusion molding, or the like using a thermoplastic resin such as polyethylene, polypropylene, vinyl chloride, polystyrene elastomer, or FEP. Specific shape. [0020] The filter member 4 is composed of two screens having different mesh sizes, and is a sub-screen having a relatively small mesh size and a relatively large mesh screen disposed on both sides of the front and back sides. 4b made. The main screen 4a is a polyester mesh woven from a polyester resin fiber such as polyethylene terephthalate or a polyamide resin woven from nylon or the like. Screen mesh. The sub-screen 4b is a polyethylene screen woven from polyethylene resin fibers. Further, the polyethylene-based resin fiber forming the sub-screen 4b is preferably made of a polyethylene-based resin of the same type as the polyethylene layer 20a of the flexible film 20 to be described later. [0021] The mesh size of the screens 4a, 4b can be appropriately selected corresponding to the use of the cell bag 1 with the filter, but the main screen 4a is attached to the mesh of 20 to 300 μm. The range, regarding the sub-screen 4b, is in the range of 100 to 4000 μm of the mesh, and these mesh sizes are selected. In this case, it is preferable that the mesh of the sub-screen 4b is selected from the above range so as to be 2 to 10 times larger than the mesh of the main screen 4a. Further, an example of use of the cell bag 1 with a filter will be described later. [0022] The container body 2 is formed by bagging the flexible film 20 into a bag shape. As the flexible film 20 forming the container body 2, a flexible film 20 including at least a polyethylene layer 20a is used. The flexible film 20 may be made of, for example, polyethylene, ethylene-vinyl acetate elastomer, or tetrafluoroethylene-hexafluoropropylene copolymer, in addition to the polyethylene-based resin film containing the polyethylene layer 20a in a single layer. A laminated film in which one or two or more layers of a thermoplastic resin such as FEP) are laminated such that the polyethylene layer 20a is located on the surface layer. Further, the flexible film 20 forming the container body 2 preferably has transparency to confirm the progress of cell culture, the state of internal cells, or the color of the medium. [0023] The container body 2 can be overlapped by, for example, aligning two flexible films 20 of a desired size, and at this time, between the two flexible films 20, the polyethylene layer 20a is opposed to each other. The state of the filter member 4 is formed by thermally fusing the peripheral portions. In this case, the injection/outlet ports 3a and 3b are provided in the respective partitions 1st and 2nd divided by the filter member 4, and when the peripheral portion of the container body 2 is thermally fused, it is only required to be in the peripheral portion of the container body 2. The specific position between the flexible film 20 and the filter member 4 may be thermally fused by a tubular member forming the injection/outlet ports 3a and 3b. Moreover, in the case of heat fusion, the temperature is set to a temperature equal to or higher than the melting point of the polyethylene resin used for the sub-screen 4b and not to the melting point of the polyester resin or the polyamide resin used in the main screen 4a. In addition, FIG. 2 is an explanatory view showing a manufacturing example of the cell bag 1 with a filter according to the embodiment of the present invention, and shows the flexible film 20, the filter member 4, and the injection and discharge ports 3a and 3b. Configuration. [0024] In the container body 2 thus formed, in the heat fusing portion 5 formed at the peripheral portion thereof, the sub-screens 4b disposed on both sides of the front and back sides of the main screen 4a (specifically, the sub-screens 4b are formed) The polyethylene-based resin fiber is melted, invades into the void portion of the main screen 4a, and is fused to each other, and is fused to the polyethylene layer 20a of the flexible film 20. As a result, the sealing strength of the flexible film 20 forming the container body 2 and the filter member 4 is improved, and the breaking strength of the cell bag 1 with the filter is improved. [0025] When the peripheral portion is thermally fused to sandwich the container body 2 in a state in which the filter member 4 is interposed between the flexible films 20, the filter member 4 is also cut to a desired size, but is heated. The fusion portion 5 is preferably cut in such a manner that the end edge of the main screen 4a is located closer to the inside of the container than the end edge of the sub-screen 4b. Further, it is more preferable to make the edge of the sub-screen 4b more aligned than the edge of the flexible film 20 forming the container body 2 on the inside of the container. 3 corresponds to the BB cross section of FIG. 1(a), and shows the positional relationship between the respective edges of the flexible film 20, the main screen 4a, and the sub-screen 4b of the heat-fusible portion 5, and the flexible film 20 The end edge is indicated by a chain line α, the end edge of the main screen 4a is indicated by a chain line β, and the edge of the sub-screen 4b is indicated by a chain line γ. Thus, at the outermost edge portion of the heat-fusible portion 5, the polyethylene layers 20a of the flexible film 20 are directly or transmitted through the sub-mesh 4b, and the layers are melted and fused. As a result, the outermost edge portion of the container body 2 is more strongly thermally fused, and the bag breaking strength of the cell bag 1 with the filter is further improved. [0027] Next, an example of use of the cell bag 1 with a filter shown in FIG. 1 as an example of the present embodiment will be described. Fig. 4 shows an example in which the cell bag 1 with a filter shown in Fig. 1 is used as a culture bag which can be easily and efficiently exchanged with a medium. In the container body 2, which is filled with the medium, the injection/extraction port 3b is placed in the second partition 2nd, and the cultured cells C are injected and cultured. At this time, considering the size of the cell C to be cultured, the size of the sieve which does not allow the passage of the cell C is selected as the main screen 4a of the filter member 4. Thereby, the cell C in the culture can reside in the second division 2nd without moving to the first division 1st (refer to FIG. 4(a)). Further, the screen size of the sub-screen 4b of the filter member 4 may be larger than the size of the screen of the main screen 4a, and may be appropriately selected depending on the size of the cell C of the culture target. [0029] When the medium is exchanged during the culture period, the old medium is discharged from the injection/outlet port 3a provided in the first division 1st (refer to FIG. 4(b)). At this time, since the cells C in the culture can reside in the second division 2nd without being discharged together with the old medium, more old medium can be discharged. After the old medium is discharged, the same amount of the new medium (refer to FIG. 4(c)) is injected from the injection port 3a provided in the first division 1st (see FIG. 4(c)), and the culture is continued. As described above, in the present application example, the medium replacement during the culture can be carried out simply and efficiently, and in particular, the discharge operation of the old medium can be performed. [0031] Further, since the cell bag 1 with the filter attached to the present invention can be in a state in which the cells C reside in the second division 2nd, the injection and discharge port 3a provided in the first division 1st is discharged into the container. In the case of the culture container of the cell culture kit of Patent Document 1, the cell recovery operation exemplified in Patent Document 1 does not require a cell sedimentation time, and can be easily and efficiently performed. In addition, the cell suspension recovered from the culture container is transferred to the second partition 2nd using the culture container as the cell culture kit of Patent Document 1, and the injection and discharge port 3a provided in the first partition 1st alternates alternately. The cell washing operation is carried out by injecting and discharging the washing liquid, and the washing and collecting mechanism can be introduced into the cell culture kit of Patent Document 1. As described above, in the present example, as the main screen 4a of the filter member 4, the main screen which does not allow the cells C to be cultured to pass through is selected, and is injected in the division 2nd by one of the divisions of the filter member 4. When the cell C is cultured, the cell C can be placed in the state of the division 2nd, and the injection port 3a for the other portion 1st divided by the filter member 4 is injected, and the culture medium and the cleaning solution are injected and discharged. This use example is an example of use of the cell bag 1 with a filter according to the present embodiment, and the cell pack 1 with a filter according to the present embodiment can be easily used for cell culture. Various operations such as cell division, filtration, washing, or medium filtration performed at the time of cell culture are performed efficiently. The preferred embodiments of the present invention have been described above, but the present invention is not limited to the embodiments described above, and various modifications can be made without departing from the scope of the invention. For example, in the above-described embodiment, the case where the inside of the container main body 2 is divided into two chambers is exemplified, but a plurality of the same filter members 4 may be used to divide into three or more chambers. Further, although the examples of the injection/outlet ports 3a and 3b are provided one by one in each division, two or more injection/outlet ports may be provided as needed. Further, the injection outflow port may be provided in at least one division, or may be provided with a division in which the injection outflow port is not provided. The contents of the documents described in the present specification and the contents of the Japanese application filed on the basis of the priority of the present application are hereby incorporated by reference. [Industrial Applicability] [0037] The present invention is applicable to medicine. In the field of biochemistry, it is utilized as a technique for efficiently culturing cells.

[0038]

1‧‧‧cell bag with filter

2‧‧‧ container body

20‧‧‧Flexible film

20a‧‧‧polyethylene layer

3a, 3b‧‧‧injection port

4‧‧‧Filter components

4a‧‧‧Main screen

4b‧‧‧Sub Screen

5‧‧‧Hot fusion department

1st‧‧‧ first division

2nd‧‧‧ second division

[ Fig. 1] Fig. 1 is an explanatory view showing an example of a cell bag with a filter according to an embodiment of the present invention. Fig. 2 is an explanatory view showing a manufacturing example of a cell bag with a filter according to an embodiment of the present invention. Fig. 3 is an explanatory view showing the positional relationship of the respective edges of the flexible film, the main screen, and the sub-screen of the heat fusion portion. Fig. 4 is an explanatory view showing an example of use of a cell bag with a filter according to an embodiment of the present invention.

Claims (3)

A cell bag with a filter, comprising: a container body and a port for injecting and discharging, wherein the inside of the container body is divided by a filter member, and at least one partition is provided with a port for injection and discharge, and the filtering The device member is made of a polyester mesh made of 20~300μm mesh or a main mesh made of polyamide and a polyethylene mesh screen of 100~4000μm disposed on both sides of the front and back sides. Between the flexible films including at least the polyethylene layer, the peripheral portion is thermally fused with the filter member interposed therebetween, and the heat-fused portion formed at the peripheral portion of the container body is formed. The polyethylene sub-screen is melted, invaded into the void portion of the main screen, and fused to each other, and is fused to the polyethylene layer of the flexible film. The cell bag with a filter according to claim 1, wherein in the heat fusion portion, an end edge of the main screen is located inside the container than an edge of the sub-screen, at an outermost edge of the heat fusion portion. The polyethylene layer of the flexible film is melted and fused together directly or through the sub-screen. A method for producing a cell bag with a filter, comprising: a container body and a port for injecting and discharging, wherein the inside of the container body is divided by a filter member, and at least one partition is provided with a port for injection and discharge A method for producing a cell bag with a filter, which is characterized in that a polyethylene having a mesh of 100 to 4000 μm is disposed on the front and back sides of a mesh of 20 to 300 μm of a mesh or a polyamide mesh. In the sub-screen, the filter member is prepared, and between the flexible films including at least the polyethylene layer, the filter member is sandwiched by the polyethylene layer, and the peripheral portions are thermally fused.