# US 11,078,455 B2 · Closed culture vessel for adherent cells > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 11,078,455 B2 - Document ID: US11078455B2 - Status: Granted - Kind: B2 - Application number: 15/311,880 - Filed: 2015-06-09 - Published: 2017-04-27 - Granted: 2021-08-03 - Earliest claimed: 2014-06-09 - Applicants: Seiichi Yokoo - Inventors: Seiichi Yokoo - Web page: https://patentgrep.com/patent/US11078455B2 ## CPC - C12M 37/02: {Filters} - C12M 23/14: {Bags} - C12M 23/24: {Gas permeable parts} - C12M 23/48: {Holding appliances; Racks; Supports (holding devices for laboratory apparatus B01L9/00)} - C12M 29/20: {Degassing; Venting; Bubble traps (means for collection or storage of gas C12M23/36; gas collection apparatus for laboratory use B01L5/02)} - C12M 33/00: {Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus (chemical or physical laboratory apparatus in general B01L, devices for taking cell samples A61B10/0045, withdrawing or distributing predetermined quantities of fluid B01L99/00)} - C12M 37/04: {Seals} - C12M: APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; {APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS} ## Abstract The present invention provides a culture vessel with which cells can be aseptically cultured and aseptically transferred to a place where the cells are to be used (such as an operating room) and from which the cells can be simply taken out. The present invention provides a closed culture vessel including an openable airtight vessel, and a cell culture member disposed in the airtight vessel and including a cell culture surface, in which the cell culture member is provided detachably from the airtight vessel. ## Claims ### Claim 1 A closed cell culture system, comprising: (i) a closed culture vessel for adherent cells comprising: a gas permeable cell culture bag; and an amnion membrane; (ii) medium supply means for supplying a culture medium to the gas permeable cell culture bag; and (iii) medium discharge means for discharging the culture medium from the gas permeable cell culture bag, wherein the amnion membrane is provided inside the gas permeable cell culture bag; wherein the amnion membrane is provided detachably from the gas permeable cell culture bag; wherein the closed culture vessel comprises no culture medium; wherein the supply means and the discharge means are aseptically closed; and wherein the gas permeable cell culture bag has been aseptically sealed. ### Claim 2 (depends on 1) The closed cell culture system according to claim 1, wherein an inner surface of the gas permeable cell culture bag is non-cell-adhesive. ### Claim 3 (depends on 1) The closed cell culture system according to claim 1, wherein the gas permeable cell culture bag includes a cell inlet that is closed with a rubber plug through which adherent cells can be aseptically introduced onto a cell adhesion surface of the amnion membrane by using a syringe equipped with a needle that can penetrate through the rubber plug. ### Claim 4 (depends on 1) A method, comprising: providing a closed culture vessel for adherent cells according to claim 1; unsealing the gas permeable cell culture bag; introducing an adhesive cell in a culture medium onto the amnion membrane inside the vessel to allow the adhesive cell to attach to the amnion membrane; sealing the gas permeable cell culture bag to include a space therein formed by filling of the gas permeable cell culture bag with a culture medium and optionally a gas, thereby protecting a cell culture surface of the amnion membrane and cells adhered to the amnion membrane from physical damage; and then, culturing the cell on the amnion membrane to form a cell sheet. ### Claim 5 (depends on 4) The method according to claim 4, further comprising aseptically transferring the closed culture vessel for adherent cells comprising the cell sheet to an operating room. ### Claim 6 A method, comprising: providing a closed culture vessel for adherent cells, comprising: a gas permeable cell culture bag having a cell inlet; and an amnion membrane, wherein the amnion membrane is provided inside the gas permeable cell culture bag, wherein the amnion membrane is provided detachably from the gas permeable cell culture bag; wherein the closed culture vessel comprises no culture medium; and wherein the cell inlet is aseptically closed with a rubber plug, introducing an adhesive cell in a culture medium at least onto the amnion membrane inside the bag through the rubber plug by using a syringe equipped with a needle that can penetrate through the rubber plug to allow the adhesive cell to attach to the amnion membrane, sealing the cell inlet of the gas permeable cell culture bag to include a space therein formed by filling of the gas permeable cell culture bag with a culture medium and optionally a gas, thereby protecting a cell culture surface of the amnion membrane and cells adhered to the amnion membrane from physical damage, and then, culturing the cell on the amnion membrane to form a cell sheet. ### Claim 7 (depends on 6) The method according to claim 6, further comprising aseptically transferring the closed culture vessel for adherent cells comprising the cell sheet to an operating room.