JP2018533926A - 脂肪間質細胞を急速分離する組成物 - Google Patents
脂肪間質細胞を急速分離する組成物 Download PDFInfo
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Abstract
【選択図】図2
Description
1.細胞が細胞培養皿に付着可能であること。
2.表面抗原として、CD45、CD34、CD14又はCD11b、CD79a又はCD19、或いはHLA−DRを発現することなく、CD105、CD73又はCD90を発現すること。
3.誘導によって脂肪細胞、骨芽細胞及び軟骨細胞へ分化する特性を有すること。
1.2.5%(v/v)のコラゲナーゼタイプI、0.1%(v/v)のトリプシン(Trypsin)、及び0.02%(v/v)のEDTAからなる酵素配合。
2.3%(v/v)のコラゲナーゼタイプI、0.1%(v/v)のトリプシン(Trypsin)、及び0.02%(v/v)のEDTAからなる酵素配合。
3.4%(v/v)のコラゲナーゼタイプI、0.1%(v/v)のトリプシン(Trypsin)、及び0.02%(v/v)のEDTAからなる酵素配合。
B 0.5%のコラゲナーゼタイプI+0.1%のトリプシン+0.02%のEDTA
C 0.5%のコラゲナーゼタイプI+0.2%のトリプシン+0.02%のEDTA
D 1%のコラゲナーゼタイプI+0.1%のトリプシン+0.02%のEDTA
E 2%のコラゲナーゼタイプI+0.1%のトリプシン+0.02%のEDTA
F 2.5%のコラゲナーゼタイプI+0.1%のトリプシン+0.02%のEDTA
G 3%のコラゲナーゼタイプI+0.1%のトリプシン+0.02%のEDTA
H 4%のコラゲナーゼタイプI+0.1%のトリプシン+0.02%のEDTA
E’ 2%のコラゲナーゼタイプIV+0.1%のトリプシン+0.02%のEDTA
F’ 2.5%のコラゲナーゼタイプIV+0.1%のトリプシン+0.02%のEDTA
G’ 3%のコラゲナーゼタイプIV+0.1%のトリプシン+0.02%のEDTA
I’ 3.5%のコラゲナーゼタイプIV+0.1%のトリプシン+0.02%のEDTA
H’ 4%のコラゲナーゼタイプIV+0.1%のトリプシン+0.02%のEDTA
Claims (9)
- 0.5〜8%(v/v)のコラゲナーゼタイプI、0.1〜0.6%(v/v)のトリプシン、及び0.01〜0.2%(v/v)の金属イオンキレート剤を含む脂肪間質細胞を分離する組成物。
- 当該金属イオンキレート剤は、エチレンジアミン四酢酸又はそのナトリウム塩、グリコールエーテルジアミン四酢酸又はそのナトリウム塩、ジエチレントリアミン五酢酸又はそのナトリウム塩、ポリリン酸塩、有機リン酸塩、リン酸エステル、ポリアクリル酸塩、有機リン酸塩、グルコン酸ナトリウム、又はこれらの混合から選択される請求項1に記載の組成物。
- 当該金属イオンキレート剤はEDTAである請求項2に記載の組成物。
- (a)脂肪組織を採取するステップ、
(b)請求項1に記載の組成物を加え、均質化して反応させることで、消化された組織の混合物を取得するステップであって、当該組成物は、0.5〜8%(v/v)のコラゲナーゼタイプI、0.1〜0.6%(v/v)のトリプシン、及び0.01〜0.2%(v/v)の金属イオンキレート剤を含むステップ、
(c)ステップ(b)の消化された組織の混合物を遠心分離し、異物を除去して、脂肪間質細胞を含む第1濾液を取得するステップ、
(d)低張液をステップ(c)の第1濾液に加えることで、血球細胞が除去された脂肪間質細胞の第2濾液を取得するステップ、及び
(e)ステップ(d)の第2濾液を中和して遠心分離するステップ、を含む脂肪間質細胞を分離する方法。 - 当該金属イオンキレート剤は、エチレンジアミン四酢酸又はそのナトリウム塩、グリコールエーテルジアミン四酢酸又はそのナトリウム塩、ジエチレントリアミン五酢酸又はそのナトリウム塩、ポリリン酸塩、有機リン酸塩、リン酸エステル、ポリアクリル酸塩、有機リン酸塩、グルコン酸ナトリウム、又はこれらの混合から選択される請求項4に記載の方法。
- 当該金属イオンキレート剤はEDTAである請求項5に記載の方法。
- 更に、総反応時間を1時間以内として脂肪組織を消化し、当該脂肪間質細胞を取得することを特徴とする請求項4に記載の方法。
- 5グラムの脂肪組織から少なくとも100万個の脂肪間質細胞を分離する効果を有することを特徴とする請求項4に記載の方法。
- 当該脂肪間質細胞は、脂肪細胞、造血細胞、血管内皮細胞、骨芽細胞、軟骨細胞、神経細胞又は上皮細胞に分化可能である請求項4に記載の方法。
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PCT/CN2015/091449 WO2017059566A1 (zh) | 2015-10-08 | 2015-10-08 | 一种快速分离脂肪间质细胞的组合物 |
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CN111621473A (zh) * | 2019-04-03 | 2020-09-04 | 成熙(上海)生物科技有限公司 | 一种新型人源脂肪干细胞制剂的制备方法 |
CN110628713A (zh) * | 2019-11-07 | 2019-12-31 | 安徽科门生物科技有限公司 | 一种脂肪源干细胞提取制备方法 |
CN111088222B (zh) * | 2019-11-11 | 2023-08-29 | 广东国科细胞科技有限公司 | 一种脂肪组织的单细胞悬液的制备方法 |
CN112063583B (zh) * | 2020-11-16 | 2021-03-23 | 广州杜德生物科技有限公司 | 一种从脂肪组织中高效分离提取脂肪间充质干细胞的方法 |
CN113717936A (zh) * | 2021-09-08 | 2021-11-30 | 四川中科博瑞生物科技有限公司 | 一种从脂肪中分离提取冻存脂肪干细胞的方法 |
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JP2010529851A (ja) * | 2007-06-15 | 2010-09-02 | アイピエリアン,インコーポレイティド | 多重分化能性/多能性細胞及び方法 |
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KR20080097190A (ko) * | 2005-12-29 | 2008-11-04 | 안트로제네시스 코포레이션 | 태반 줄기세포의 수집과 보존을 위한 개선된 조성물과 이조성물의 이용 방법 |
WO2009115581A2 (en) * | 2008-03-19 | 2009-09-24 | Cryo-Save Ag | Improved cryopreservation of adipose tissue for the isolation of mesenchymal stem cells |
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- 2015-10-08 CN CN201580083577.6A patent/CN108138140A/zh active Pending
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- 2015-10-08 WO PCT/CN2015/091449 patent/WO2017059566A1/zh active Application Filing
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JP2009537137A (ja) * | 2006-05-17 | 2009-10-29 | コグネート セラピューティクス,インコーポレーテッド | 脂肪吸引後の脂肪吸引物からの造血幹細胞の単離および精製 |
JP2010529851A (ja) * | 2007-06-15 | 2010-09-02 | アイピエリアン,インコーポレイティド | 多重分化能性/多能性細胞及び方法 |
JP2012505665A (ja) * | 2008-10-17 | 2012-03-08 | バクスター・インターナショナル・インコーポレイテッド | 脂肪組織から細胞集団を得る方法 |
CN102274491A (zh) * | 2011-07-13 | 2011-12-14 | 中国人民解放军第四军医大学 | 用于促进缺血组织血管生成的干细胞联合生长因子注射液及其制备方法和使用方法 |
CN104164403A (zh) * | 2014-07-25 | 2014-11-26 | 大连大学 | 一种提取及培养脂肪干细胞的方法 |
CN104560868A (zh) * | 2014-11-27 | 2015-04-29 | 广州赛莱拉干细胞科技股份有限公司 | 一种脂肪干细胞的原代分离培养方法 |
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METHODS, vol. 45, no. 2, JPN6019011657, 2008, pages 115 - 120, ISSN: 0004008166 * |
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