JPWO2019054468A1 - 細胞含有試料からの細胞の遊離方法 - Google Patents
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Abstract
Description
細胞含有試料と酵素とを含む反応混合液及び気相を、反応混合液と気相との合計体積に対して気相の体積が7体積%以上となる割合で収容し閉鎖した袋状容器を振盪させる工程を含むことを特徴とする方法。
(2)細胞含有試料が脂肪組織である、(1)に記載の方法
(3)袋状容器の反応混合液及び気相を収容する収容部の、平面視での、短手方向幅をAとし長手方向幅をBとしたとき、A/Bが0.1以上1以下である、(1)又は(2)に記載の方法。
本明細書は本願の優先権の基礎となる日本国特許出願番号2017−178267の開示内容を包含する。
<用語、材料>
細胞含有試料としては生体組織、in vitroで調製した細胞培養物等が例示できる。生体組織は典型的には動物より採取した生体組織であり、例えば、脂肪、皮膚、血管、角膜、口腔、腎臓、肝臓、膵臓、心臓、神経、筋肉、前立腺、腸、羊膜、胎盤、臍帯などに由来する生体組織が挙げられる。本明細書で開示する方法は特に脂肪組織から間質血管画分(SVF)細胞を取り出すのに有用である。
有核細胞とは、細胞内に核を有する細胞である。
本発明で用いる袋状容器は、柔軟性(可撓性ともいう)を有する材料により形成された袋状の容器である。袋状容器は、少なくとも一部を透明または半透明の材料からなるものとすると、内部の細胞懸濁液の様子を目視で観察できるため好ましい。
本実施形態の容器1は、容器本体10、入口ポート41及び出口ポート51を備える。
本実施形態の容器1は、容器本体10及び入口ポート兼出口ポート61を備える。
本実施形態の容器1は、容器本体10及び入口ポート兼出口ポート61を備える。
本発明は、
細胞含有試料を酵素処理して細胞含有試料から細胞を遊離させる方法であって、
細胞含有試料と酵素とを含む反応混合液及び気相を、反応混合液と気相との合計体積に対して気相の体積が7体積%以上となる割合で収容し閉鎖した袋状容器を振盪させる工程を含むことを特徴とする。
第1又は第2の酵素処理用バッグ1の第1出口ポート31及び第2出口ポート32を蓋313、323で封鎖し、収容部14内に、10〜50mLの脂肪組織と、10〜50mLのコラゲナーゼ液(コラゲナーゼ濃度0.1w/v%となるようにコラゲナーゼを生理食塩液で溶解して調製)とを、脂肪組織とコラゲナーゼ液との体積比が1:1〜1:3となるように収容して反応混合液201とし、収容部14内の空気相202の体積が、収容部14の体積(すなわち反応混合液201と空気相202との合計体積)に対して表1に示す所定の割合(%)となるように空気を加えて、第1又は第2の酵素処理用バッグ1の第1入口ポート21、第2入口ポート22、第3入口ポート24を蓋213、223、233で封鎖して密封した。
結果を表1に示す。
Claims (3)
- 細胞含有試料を酵素処理して、細胞含有試料から細胞を遊離させる方法であって、
細胞含有試料と酵素とを含む反応混合液及び気相を、反応混合液と気相との合計体積に対して気相の体積が7体積%以上となる割合で収容し閉鎖した袋状容器を振盪させる工程を含むことを特徴とする方法。 - 細胞含有試料が脂肪組織である、請求項1に記載の方法。
- 袋状容器の反応混合液及び気相を収容する収容部の、平面視での、短手方向幅をAとし長手方向幅をBとしたとき、A/Bが0.1以上1以下である、請求項1又は2に記載の方法。
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JP5021936B2 (ja) * | 2006-01-16 | 2012-09-12 | 株式会社カネカ | 脂肪組織の酵素処理液から有核細胞を採取する方法 |
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US6068775A (en) * | 1998-04-13 | 2000-05-30 | Circe Biomedical, Inc. | Removal of agent from cell suspension |
WO2005012480A2 (en) * | 2003-06-25 | 2005-02-10 | Macropore Biosurgery Inc. | Systems and methods for separating and concentrating regenerative cells from tissue |
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