Cell culturing method and kit
The present invention relates to a cell culturing method and kit. More specifically, it relates to a cell culturing method and kit using a support that is exposed to the air. It further relates to a method of culturing cells by allowing them to migrate onto a porous polyimide film.
1. A cell culturing method including:
(1) a step of supporting cells on one or more sheet-like porous supports,
(2) a step of applying a medium to the sheet-like porous support supporting the cells and wetting the sheet-like porous support with the medium, with the medium included in some or all of the pores of the sheet-like porous support,
(3) a step of disposing the sheet-like porous support in a culturing vessel that houses medium, in such a manner that all or a portion of the surface of the sheet-like porous support wetted with the medium is exposed to a gas phase, and
(4) a step of setting the culturing vessel in an incubator and culturing the cells,
wherein the wetted state of the sheet-like porous support surface and interior is maintained throughout the culturing,
wherein the sheet-like porous support is a porous polyimide film,
wherein the porous polyimide film has a multilayer structure having at least two surface layers (an A-surface and a B-surface), and a macro-void layer sandwiched between the two surface layers,
a mean pore size of the holes in the A-surface is smaller than a mean pore size of the holes in the B-surface, and
the macro-void layer has a partition bonded to the surface layers (the A-surface and the B-surface), and a plurality of macro-voids surrounded by the partition and the surface layers (the A-surface and the B-surface).
2. The method according to claim 1 , wherein in step (4), the medium is supplied into the culturing vessel in a continuous or intermittent manner, and all or a portion of the sheet-like porous support wetted with the medium is exposed to a gas phase throughout the culturing.
3. The method according to claim 1 , wherein in step (4), the step of culturing the cells further comprises supplying oxygen by an oxygen-supplying means.
4. The method according to claim 1 , wherein in step (3), the sheet-like porous support is mounted on a rigid body.
5. The method according to claim 4 , wherein the rigid body is a metal mesh.
6. The method according to claim 1 , wherein in step (3), a porous sheet with a larger mean pore size than the sheet-like porous supports is mounted in such a manner as to cover all or a portion of the top surface of the one or more sheet-like porous support.
7. The method according to claim 6 , wherein the porous sheet with a larger mean pore size than the sheet-like porous support is selected from the group consisting of nonwoven fabrics, gauzes and sponges.
8. The method according to claim 1 , wherein the culturing vessel is an open vessel.
9. The method according to claim 1 , wherein the culturing vessel is a closed vessel.
10. The method according to claim 1 , wherein in step (3), two or more sheet-like porous supports are disposed in a manner layered one over the other.
11. The method according to claim 1 , wherein in step (3), one or more sheet-like porous supports are disposed in a folded manner.
12. The method according to claim 1 , wherein the porous polyimide film includes a polyimide obtained from a tetracarboxylic dianhydride and a diamine.
13. The method according to claim 12 , wherein the porous polyimide film is a colored porous polyimide film obtained by forming a polyamic add solution composition from a tetracarboxylic dianhydride and a diamine, and a coloring precursor, and then heat treating the polyamic acid solution at 250° C. or higher.
14. The method according to claim 1 , wherein the cells are transformed by genetic engineering technology so as to express a substance.
15. The method according to claim 1 , wherein the cells are selected from the group consisting of animal cells, insect cells, plant cells, yeast cells and bacteria.
16. The method according to claim 15 , wherein the animal cells are cells derived from an animal belonging to the subphylum Vertebrata.
17. The method according to claim 16 , wherein the cells are selected from the group consisting of Chinese hamster ovary (CHO) cells, CHO-K1 cells, Vero cells and Madin-Darby Canine Kidney (MDCK) cells.
18. The method according to claim 1 , wherein a mean pore size of the A-surface is 0.01 μm to 15 μm, and a mean pore size of the B-surface is 20 μm to 100 μm.