# US 10,370,696 B2 · Method for cell recovery > Agents: this data is available as JSON from the patentgrep API and MCP server — see https://patentgrep.com/llms.txt - Number: US 10,370,696 B2 - Document ID: US10370696B2 - Status: Granted - Kind: B2 - Application number: 15/509,140 - Filed: 2015-09-09 - Published: 2017-09-07 - Granted: 2019-08-06 - Earliest claimed: 2014-09-09 - Assignees: Nissan Chemical Industries, Ltd. - Applicants: Nissan Chemical Industries, Ltd. - Inventors: Ayako Aihara; Taito Nishino - Web page: https://patentgrep.com/patent/US10370696B2 ## CPC - C12Q 1/24: Methods of sampling, or inoculating or spreading a sample; Methods of physically isolating an intact microorganisms - C12N 5/00: Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor; (plant reproduction by tissue culture techniques A01H4/00) - C12N 5/06: {Animal cells or tissues; Human cells or tissues (preservation of excised living parts A01N1/10)} - C12N 2500/34: Sugars - C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES - C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00) ## Abstract The present invention provides a method of isolating cells or tissues from a culture preparation of the cells or tissues in a medium composition which enables culture of the cells or tissues in suspension, which comprises at least one step selected from the group consisting of the following (A), (B) and (C): (A) passing the culture preparation through a filter having fine pores having a pore diameter of 5-500 μm, (B) adding a chelator to the culture preparation, and (C) diluting the culture preparation with a physiological aqueous solution. ## Claims ### Claim 1 A method of isolating cells or tissues, which method comprises (A1) passing a culture preparation of the cells or tissues in a medium composition, which comprises a polymer compound having an anionic functional group that enables culture of the cells or tissues in suspension, through a filter having fine pores with a diameter of 5-500 μm, (A2) isolating cells or tissues from the filtrate obtained in step (A1), and (B) adding a chelator to the culture preparation, wherein the chelator is citric acid or a salt thereof. ### Claim 2 (depends on 1) The method according to claim 1, wherein the polymer compound is an acidic polysaccharide having an anionic functional group. ### Claim 3 (depends on 1) The method according to claim 1, wherein the medium composition comprises deacylated gellan gum or a salt thereof. ### Claim 4 (depends on 1) The method according to claim 1, wherein the fine pores of the filter have a diameter of 20-100 μm. ### Claim 5 (depends on 1) The method according to claim 1, wherein the culture preparation is passed through the filter plural times. ### Claim 6 (depends on 1) The method according to claim 1, wherein, after addition of the chelator, the culture preparation is stirred for not less than 10 min. ### Claim 7 (depends on 1) The method according to claim 1, further comprising (C) diluting the culture preparation with a physiological aqueous solution, wherein the physiological aqueous solution is substantially free of calcium ion and magnesium ion. ### Claim 8 (depends on 7) The method according to claim 7, wherein the steps (A1), (A2), (B) and (C) are performed in the order of (A1), (A2), (B), (C). ### Claim 9 (depends on 7) The method according to claim 7, wherein the polymer compound is an acidic polysaccharide having an anionic functional group. ### Claim 10 (depends on 7) The method according to claim 7, wherein the medium composition comprises deacylated gellan gum or a salt thereof. ### Claim 11 (depends on 7) The method according to claim 7, wherein the fine pores of the filter have a diameter of 20-100 μm. ### Claim 12 (depends on 7) The method according to claim 7, wherein the culture preparation is passed through the filter plural times. ### Claim 13 (depends on 7) The method according to claim 7, wherein, after addition of the chelator, the culture preparation is stirred for not less than 10 min. ### Claim 14 (depends on 7) The method according to claim 7, wherein the physiological aqueous solution is phosphate buffered saline. ### Claim 15 A method of isolating cells or tissues, which method comprises (A1) passing a culture preparation of the cells or tissues in a medium composition, which comprises a polymer compound having an anionic functional group that enables culture of the cells or tissues in suspension, through a filter having fine pores with a diameter of 5-500 μm, (A2) isolating cells or tissues from the filtrate obtained in step (A1), and (C) diluting the culture preparation with a physiological aqueous solution, wherein the physiological aqueous solution is substantially free of calcium ion and magnesium ion. ### Claim 16 (depends on 15) The method according to claim 15, wherein the polymer compound is an acidic polysaccharide having an anionic functional group. ### Claim 17 (depends on 15) The method according to claim 15, wherein the medium composition comprises deacylated gellan gum or a salt thereof. ### Claim 18 (depends on 15) The method according to claim 15, wherein the culture preparation is passed through the filter plural times. ### Claim 19 (depends on 15) The method according to claim 15, wherein the fine pores of the filter have a diameter of 20-100 μm. ### Claim 20 (depends on 15) The method according to claim 15, wherein the physiological aqueous solution is phosphate buffered saline.