# US 11,306,286 B2 · Cell culture substrate > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 11,306,286 B2 - Document ID: US11306286B2 - Status: Granted - Kind: B2 - Application number: 16/467,520 - Filed: 2017-12-12 - Published: 2019-11-07 - Granted: 2022-04-19 - Earliest claimed: 2016-12-22 - Assignees: DIC Corporation - Applicants: DIC Corporation - Inventors: Hirohide Nakaguma; Tetsuo Takada; Ayako Isshiki - Web page: https://patentgrep.com/patent/US11306286B2 ## CPC - C12N 5/0068: {General culture methods using substrates (for specific animal cell type C12N5/06)} - C08F 220/12: of monohydric alcohols or phenols - C08F 220/16: of phenols or of alcohols containing two or more carbon atoms - C08F 220/387: {and containing nitrogen and oxygen} - C08F 299/024: {the unsaturation being in acrylic or methacrylic groups} - C08L 33/14: of esters containing halogen, nitrogen, sulfur, or oxygen atoms in addition to the carboxy oxygen - C08L 87/00: Compositions of unspecified macromolecular compounds, obtained otherwise than by polymerisation reactions only involving unsaturated carbon-to-carbon bonds - C12N 5/0607: {Non-embryonic pluripotent stem cells, e.g. MASC (induced pluripotent stem cells C12N5/0696)} - C12N 5/0696: {Artificially induced pluripotent stem cells, e.g. iPS} - C08G 81/00: Macromolecular compounds obtained by interreacting polymers in the absence of monomers, e.g. block polymers (involving only carbon-to-carbon unsaturated bond reactions C08F299/00 {; polyester-amides C08G69/44; polyester-imides C08G73/16; polyamides-imides C08G73/14; block- or graft polymers containing polysiloxane sequences C08G77/42}) - C08G 81/02: at least one of the polymers being obtained by reactions involving only carbon-to-carbon unsaturated bonds - C08G 81/024: {Block or graft polymers containing sequences of polymers of C08C or C08F and of polymers of C08G} - C12N 2533/30: Synthetic polymers - C12N 2533/52: Fibronectin; Laminin - C12N 2533/54: Collagen; Gelatin - C12N 2533/90: Substrates of biological origin, e.g. extracellular matrix, decellularised tissue - C12N 2539/00: Supports and/or coatings for cell culture characterised by properties - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) - C08F: MACROMOLECULAR COMPOUNDS OBTAINED BY REACTIONS ONLY INVOLVING CARBON-TO-CARBON UNSATURATED BONDS - C08L: COMPOSITIONS OF MACROMOLECULAR COMPOUNDS (compositions based on polymerisable monomers C08F, C08G; artificial filaments or fibres D01F; textile treating compositions D06) - C08G: MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS ## Abstract The present invention is to provide a cell culture substrate including a block polymer including a segment having a lower critical solution temperature and a hydrophobic segment, the cell culture substrate further including an adhesive matrix, in which the adhesive matrix is an extracellular matrix and/or an adhesive synthetic matrix. Furthermore, the invention is to provide a cell culture substrate in which the extracellular matrix is at least one selected from laminin, fibronectin, vitronectin, cadherin, and fragments thereof, and/or the adhesive synthetic matrix is poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl) ammonium hydroxide] or an oligopeptide-carrying polymer. ## Claims ### Claim 1 A cell culture substrate comprising a block polymer comprising a segment having a lower critical solution temperature and a hydrophobic segment, the cell culture substrate further including an adhesive matrix, wherein a degree of polymerization of the segment having a lower critical solution temperature is 400 to 10,000, and wherein the adhesive matrix is an extracellular matrix and/or an adhesive synthetic matrix. ### Claim 2 (depends on 1) The cell culture substrate according to claim 1, wherein the extracellular matrix is at least one selected from laminin, fibronectin, vitronectin, cadherin, and fragments thereof. ### Claim 3 (depends on 1) The cell culture substrate according to claim 1, wherein the adhesive synthetic matrix is poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl) ammonium hydroxide] or an oligopeptide-carrying polymer. ### Claim 4 (depends on 1) The cell culture substrate according to claim 1, wherein the hydrophobic segment is obtainable by polymerizing a monomer represented by the following Formula (1): wherein in Formula (1), R1 represents a hydrogen atom or a methyl group; and R2 represents any one of a phenyl group, a carboxyalkyl group having an alkyl with 1 to 8 carbon atoms, a carboxyaralkyl group having an aralkyl with 7 or 8 carbon atoms, a group represented by the following Formula (2), or a group represented by the following Formula (3): wherein in Formula (2), n represents 2 or 3; and R3 represents an alkyl group having 1 to 3 carbon atoms, wherein in Formula (3), R4 and R5 each independently represent a hydrogen atom or an alkyl group having 1 to 6 carbon atoms; and the total number of carbon atoms of R4 and R5 is 5 or more. ### Claim 5 (depends on 1) The cell culture substrate according to claim 1, further comprising at least one protein selected from gelatin, collagen, and/or albumin on the cell culture substrate. ### Claim 6 (depends on 5) The cell culture substrate according to claim 5, wherein the block polymer, at least one protein selected from gelatin, collagen, and/or albumin, and the adhesive matrix are sequentially laminated. ### Claim 7 (depends on 1) The cell culture substrate according to claim 1, which is a dry cell culture substrate. ### Claim 8 (depends on 1) The cell culture substrate according to claim 1, which is laminated on a supporting medium. ### Claim 9 (depends on 8) The cell culture substrate according to claim 8, which has an average thickness of 1,000 nm or less. ### Claim 10 (depends on 1) A method for producing a dry cell culture substrate, the method comprising: a step of coating a solution including at least one protein selected from gelatin, collagen, and/or albumin on the block polymer according to claim 1; a step of further coating a solution including the adhesive matrix according to claim 1 thereon to obtain a cell culture substrate; and a step of drying the obtained cell culture substrate. ### Claim 11 (depends on 1) A cell culture equipment comprising a supporting medium and the cell culture substrate according to claim 1. ### Claim 12 (depends on 2) The cell culture substrate according to claim 2, wherein the adhesive synthetic matrix is poly[2-(methacryloyloxy)ethyl dimethyl-(3-sulfopropyl) ammonium hydroxide] or an oligopeptide-carrying polymer. ### Claim 13 (depends on 2) The cell culture substrate according to claim 2, wherein the hydrophobic segment is obtainable by polymerizing a monomer represented by the following Formula (1): wherein in Formula (1), R1 represents a hydrogen atom or a methyl group; and R2 represents any one of a phenyl group, a carboxyalkyl group having an alkyl with 1 to 8 carbon atoms, a carboxyaralkyl group having an aralkyl with 7 or 8 carbon atoms, a group represented by the following Formula (2), or a group represented by the following Formula (3): wherein in Formula (2), n represents 2 or 3; and R3 represents an alkyl group having 1 to 3 carbon atoms, wherein in Formula (3), R4 and R5 each independently represent a hydrogen atom or an alkyl group having 1 to 6 carbon atoms; and the total number of carbon atoms of R4 and R5 is 5 or more. ### Claim 14 (depends on 2) The cell culture substrate according to claim 2, further comprising at least one protein selected from gelatin, collagen, and/or albumin on the cell culture substrate. ### Claim 15 (depends on 2) The cell culture substrate according to claim 2, which is a dry cell culture substrate. ### Claim 16 (depends on 2) The cell culture substrate according to claim 2, which is laminated on a supporting medium. ### Claim 17 (depends on 2) A cell culture equipment comprising a supporting medium and the cell culture substrate according to claim 2. ### Claim 18 (depends on 3) The cell culture substrate according to claim 3, wherein the hydrophobic segment is obtainable by polymerizing a monomer represented by the following Formula (1): wherein in Formula (1), R1 represents a hydrogen atom or a methyl group; and R2 represents any one of a phenyl group, a carboxyalkyl group having an alkyl with 1 to 8 carbon atoms, a carboxyaralkyl group having an aralkyl with 7 or 8 carbon atoms, a group represented by the following Formula (2), or a group represented by the following Formula (3): wherein in Formula (2), n represents 2 or 3; and R3 represents an alkyl group having 1 to 3 carbon atoms, wherein in Formula (3), R4 and R5 each independently represent a hydrogen atom or an alkyl group having 1 to 6 carbon atoms; and the total number of carbon atoms of R4 and R5 is 5 or more. ### Claim 19 (depends on 3) The cell culture substrate according to claim 3, further comprising at least one protein selected from gelatin, collagen, and/or albumin on the cell culture substrate. ### Claim 20 (depends on 3) The cell culture substrate according to claim 3, which is a dry cell culture substrate.