JP2016010359A - 細胞又は組織のガラス化凍結保存用治具 - Google Patents
細胞又は組織のガラス化凍結保存用治具 Download PDFInfo
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- JP2016010359A JP2016010359A JP2014134084A JP2014134084A JP2016010359A JP 2016010359 A JP2016010359 A JP 2016010359A JP 2014134084 A JP2014134084 A JP 2014134084A JP 2014134084 A JP2014134084 A JP 2014134084A JP 2016010359 A JP2016010359 A JP 2016010359A
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Abstract
【解決手段】支持体上に、接着層とガラス化液吸収層を少なくともこの順に有するガラス化液吸収体を有し、該ガラス化液吸収層の細胞又は組織を載置する部分と支持体の間に接着層が存在しない部分を有する細胞又は組織のガラス化凍結保存用治具。
【選択図】図2
Description
(2)該支持体が光透過性支持体であり、該ガラス化液吸収層が、多孔性樹脂シート、多孔性金属酸化物シート、および、密度が0.12〜0.3g/cm3であり坪量が10〜100g/m2である紙または不織布の中から選ばれる何れか一種であることを特徴とする(1)に記載の細胞又は組織のガラス化凍結保存用治具。
P=(V/T)×100(%)
P:空隙率(%)
V:空隙容量(ml/m2)
T:厚み(μm)
繊度0.06dtex(繊維径約2μm)、カット長3mmの延伸結晶化ポリエチレンテレフタレートステープル40質量部、繊度0.1dtex(繊維径約3μm)、カット長3mmの延伸結晶化ポリエチレンテレフタレートステープル20質量部および繊度0.2dtex(繊維径約4μm)、カット長3mmの非延伸ポリエチレンテレフタレートステープル40質量部を湿式抄造し、更に、表面温度200℃の熱カレンダーにより繊維間を融着させつつ厚み調整を行い、平均繊維径が3.0μm、坪量25g/m2、密度0.5g/cm3の不織布(ガラス化液吸収層)を得た。
実施例1のガラス化凍結保存用治具の作製において、易接着処理された透明PETフィルム支持体上に、接着層として、ヘンケルジャパン社製のホットメルトウレタン樹脂Purmelt QR 170−7141Pを乾燥時の固形分量30g/m2となるように塗布した以外は同様にして、実施例2のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、アドバンテック社製の濾紙 No.5C(坪量120g/m2、密度0.57g/cm3、厚み210μm)を当該接着層に重ね合わせた以外は同様にして、実施例3のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層の無塗布部分を設けず、透明PETフィルム支持体上の全面に塗布した接着層が乾燥する前に、ガラス化液吸収層を重ね合わせた以外は同様にして、比較例1のガラス化凍結保存用治具を作製した。
実施例3のガラス化凍結保存用治具の作製において、接着層の無塗布部分を設けず、透明PETフィルム支持体上の全面に塗布した接着層が乾燥する前に、ガラス化液吸収層を重ね合わせた以外は同様にして、比較例2のガラス化凍結保存用治具を作製した。
実施例1〜3、および比較例1〜2の各ガラス化凍結保存用治具の、ガラス化液吸収層と支持体との間に接着層が存在しない部分(2mm×5mm)のガラス化液吸収層上に、ガラスビーズ(直径100μm)を疑似細胞として複数個含むガラス化液1μLを滴下付着させた。また、比較例1〜2の各ガラス化凍結保存用治具のガラス化液吸収層上に、同様に、ガラスビーズ(直径100μm)を疑似細胞として複数個含むガラス化液1μLを滴下付着させた。なお、ガラス化液は、シグマアルドリッチ社製 修正TCM199培地に、20容積%血清、15容積%DMSO、15容積%エチレングリコール、0.2容積%スクロースが含まれる組成のものを用いた。滴下付着後、落射型正立光学顕微鏡(オムロン(株)製、VC4500−S1)にて、ガラス化液吸収層上に載置された疑似細胞周辺のガラス化液が吸収される様子を観察し、ガラス化液の吸収性を以下の基準で評価した。この結果を表1に示す。
×:ガラス化液滴下付着後、経過時間が10秒を超えても、疑似細胞周辺の余分なガラス化液の残存が確認された。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、キュプラ繊維からなる不織布である旭化成せんい社製Bemliese(登録商標) SE14G(坪量14g/m2、密度0.19g/cm3、厚み70μm)を当該接着層に重ね合わせた以外は同様にして、実施例4のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、紙である大王製紙(株)製 エリエール(登録商標) プロワイプ ソフトマイクロワイパーS220(坪量22g/m2、密度0.21g/cm3)を当該接着層に重ね合わせた以外は同様にして、実施例5のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、ポリエチレン樹脂からなる多孔性樹脂シートである旭化成社製のサンファインAQ800(細孔径35μm、空隙率43%、厚み500μm)を当該接着層に重ね合わせた以外は同様にして、実施例6のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、多孔性金属酸化物シートであるコバレントマテリアル社製の石英ガラス多孔体(細孔径3.5μm、空隙率33%、厚み2mm)を当該接着層に重ね合わせた以外は同様にして、実施例7のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、ポリテトラフルオロエチレン樹脂からなる多孔性樹脂シートであるメルクミリポア社製の親水化処理されたポリテトラフルオロエチレン多孔体(細孔径10μm、空隙率80%、厚み85μm)を当該接着層に重ね合わせた以外は同様にして、実施例8のガラス化凍結保存用治具を作製した。
実施例1のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、ポリテトラフルオロエチレン樹脂からなる多孔性樹脂シートであるメルクミリポア社製の親水化処理されたポリテトラフルオロエチレン多孔体(細孔径10μm、空隙率80%、厚み85μm)を当該接着層に重ね合わせた以外は同様にして、実施例9のガラス化凍結保存用治具を作製した。
実施例2のガラス化凍結保存用治具の作製において、接着層を塗布後、ドライヤーで加熱しながら、ガラス化液吸収層として、ポリビニリデンジフロライド樹脂からなる多孔性樹脂シートであるメルクミリポア社製のポリビニリデンジフロライド多孔体(細孔径0.5μm、空隙率70%、厚み125μm)を当該接着層に重ね合わせた以外は同様にして、実施例10のガラス化凍結保存用治具を作製した。
上記のようにして得られた実施例4〜10の各ガラス化凍結保存用治具の、ガラス化液の吸収性の評価を、先の実施例1〜3および比較例1〜2の各ガラス化凍結保存用治具と同様の方法にて評価した。この結果を表2に示す。
実施例4〜10の各ガラス化凍結保存用治具について、先の「ガラス化液の吸収性の評価」に記載の方法で、ガラス化液吸収層と支持体との間に接着層が存在しない部分(2mm×5mm)のガラス化液吸収層上に、疑似細胞をガラス化液と共に滴下付着させた。滴下付着後、透過型光学顕微鏡(オリンパス(株)製、SZH−121)にて、ガラス化液吸収層上に載置した疑似細胞を観察できるかどうかを以下の基準で評価した。この結果を表2に示す。
○:疑似細胞の形態を観察することができる。
△:疑似細胞の存在を確認することができるが、形態を観察することができない。
×:疑似細胞の存在を確認することが困難である。
2、2a〜2d ガラス化液吸収体
3 ガラス化液吸収層
4 支持体
5 接着層
6 ガラス化液吸収層と支持体との間に接着層が存在しない部分
7 ガラス化液吸収層と支持体との間に接着層が存在する部分
8 細胞又は組織を載置する部分
9 ガラス化凍結保存用治具
Claims (2)
- 支持体上に、接着層とガラス化液吸収層を少なくともこの順に有するガラス化液吸収体を有し、該ガラス化液吸収層の細胞又は組織を載置する部分と支持体の間に、接着層が存在しない部分を有することを特徴とする細胞又は組織のガラス化凍結保存用治具。
- 該支持体が光透過性支持体であり、該ガラス化液吸収層が、多孔性樹脂シート、多孔性金属酸化物シート、および、密度が0.12〜0.3g/cm3であり坪量が10〜100g/m2である紙または不織布の中から選ばれる何れか一種であることを特徴とする請求項1に記載の細胞又は組織のガラス化凍結保存用治具。
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JP6124845B2 (ja) | 2017-05-10 |
CN106459862A (zh) | 2017-02-22 |
US20170135335A1 (en) | 2017-05-18 |
KR20170018035A (ko) | 2017-02-15 |
KR101988140B1 (ko) | 2019-06-11 |
EP3162883B1 (en) | 2018-10-31 |
EP3162883A1 (en) | 2017-05-03 |
EP3162883A4 (en) | 2018-03-14 |
WO2016002410A1 (ja) | 2016-01-07 |
US10039278B2 (en) | 2018-08-07 |
CN106459862B (zh) | 2018-12-18 |
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