JP2012511929A - 攪拌タンク反応器及び方法 - Google Patents
攪拌タンク反応器及び方法 Download PDFInfo
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- C12M23/00—Constructional details, e.g. recesses, hinges
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- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M23/00—Constructional details, e.g. recesses, hinges
- C12M23/28—Constructional details, e.g. recesses, hinges disposable or single use
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M47/00—Means for after-treatment of the produced biomass or of the fermentation or metabolic products, e.g. storage of biomass
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Abstract
Description
本発明は攪拌タンク及び方法に関する。
あるいは、未浄化細胞培養液に可溶性ポリマー電解質を加え、関心生体分子を捕捉及び凝集物形成させ、この凝集物を沈殿させて残余の溶液から分離させる。凝集物に緩く接着している不純物を代表的には洗浄により除去する。次いで、溶液のイオン強度が増大してポリマー電解質から標的タンパク質が解離され、ポリマー電解質はタンパク質含有溶液中に再溶解する。
誘導磁性粒子を使用する現場製品回収法はタンパク質精製法の1例であるが、この方法では関心生体分子を未浄化細胞培養液から直接的に精製させ得る。
この方法の欠点は、設計、構造、並びに高勾配磁気分離の検証に相当の設備投資を要することである。しかもこの方法は、今後、バイオプロセス業界でのタンパク質精製の規準になると見込まれる使い捨て性用途には向かない。
然し乍ら、析出物は典型的には大きなスラッジ塊形態であるため、その除去が問題となる。
胴部22は成型プラスチックまたはガラスの単一部品として作製することが好ましい。あるいは胴部22は、熱、接着剤、またはガスケット(図示せず)で相互にシールした2つ以上のプラスチックまたはガラス部品として作製し得る。頂部及び胴部形成用に使用し得る好適なポリマーには、これに限定しないが、ポリカーボネート、ポリエステル、ナイロン、PTFEレジン、及びその他のフルオロポリマー、アクリリック及びメタアクリリックレジン及びそのコポリマー、ポリスルホン、ポリエーテルスルホン、ポリアリルスルホン、ポリスチレン、ポリエーテルイミド、ナイロン、ポリエステル、ポリエチレンテレフタレート(PET)、ポリビニルクロリド、クロリネーテッドポリビニルクロリド、ABS及びその合金及び混合物、ポリオレフィン、好ましくは、直鎖状低密度ポリエチレン、低密度ポリエチレン、高密度ポリエチレン、及び超ポリマー重量ポリエチレン及びそのコポリマー、ポリプロピレン及びそのコポリマー、及び、メタロセン発生ポリオレフィン、が含まれる。好ましいポリマーは、ポリオレフィン、特にはポリエチレン及びそのコポリマー、ポリスチレン、及びポリカーボネート、である。頂部及び胴部は同一ポリマーまたは、所望であれば異なるポリマーから作製し得る。再使用性の実施例では、胴部をガラス、アクリリックあるいはその他の、プロセス上悪影響を及ぼさない材料から作製し得る。斯界に既知の如く、胴部22もまた廃棄性のプラスチック製袋であり得る。
所望であれば、図1の1つ以上のポート32aまたは32bを用いて胴部内にガスを提供させ得る。POREX(商標名)多孔質材料等のプラスチック製のフリット、微孔質膜またはセラミックストーンまたは燒結金属フィルタを胴部内でポートの内側に装着し、かくして所望サイズ化したガス泡沫を提供させ得る。あるいは、頂部16のポート30aを用いて、胴部内を下方に伸延するガス供給管を保持させ得る。この場合でも、胴部内でポートの内側にフリット、微孔質膜またはセラミックストーンまたは燒結金属フィルタを装着し、所望サイズ化したガス泡沫を提供させ得る。あるいは、攪拌用アセンブリ内の多孔質フィルタ/膜110を通して胴部内にガスを提供させ、このガスをポート32bを通して提供させ得る。
膜あるいはその他としてのフィルタは、マサチューセッツ州ビレリカのMillipore社から入手可能DURAPORE(商標名)PVDF膜における如く、その深さ方向全体において孔寸法が対称性を有し得、または、同社より入手可能なMILLIPORE EXPRESS(商標名)及びMILLIPORE EXPRESS(商標名)PLUSまたはSH PES膜における如くその厚さ方向を通して孔寸法が非対称性を有し得る。フィルタは所望であれば、別個の上流側の層としてか、または膜自体の上流側の一体部分としてのプレフィルタ層を含み得る。
バイオリアクターの、円筒管の如き胴部22が、図5に示す如く、フィルタベース部100とシール関係下に配置される。フィルタベース部100から下方に伸延してフィルタベース部100を支持する複数の脚部6’を設け得る。
他の実施例では、胴部22’の出口がフィルタベース部100の出口としてのポート32と流体連通されるが、胴部22’にはフィルタまたは膜は格納されない。それらに代えて、出口としてのポート32’が、図示されない管またはその他導管を介して、関心生体分子を殺菌濾過するところのMillex(商標名)フィルタまたはOptiscale(商標名)またOpticap(商標名)フィルタ等の内蔵型フィルタ装置(図示せず)と流体連通する。このフィルタ装置の出口は、磁路精製ステップ(例えば、クロマトグラフィープロセストレイン)等の好適使用ポイントに接続される。
作動に際し、スタンド内に殺菌装置を配置し、この装置の適宜の各ポート位置に空気、液体、プローブ、サンプリング、等に使用する種々の連結部を装着する。装置の、頂部16を胴部22に装着する位置よりも幾分下方で且つ、液体/空気界面を形成する所望の高さまで媒体を充填し、この種の装置に一般的なガスのヘッドスペースを残す。ポート32の少なくとも一つは前記界面高さ以下に位置付けられる。
特定のプロセス条件設定下に可溶性となるポリマーを追加し、条件(例えば、温度、塩濃度、光、電界、またはpH)の変化に際して不溶化させて溶液から析出させる。あるいは、関心生体分子または可溶性不純物に結合させるための、生体分子を精製させ得る任意のリガンドまたは機能を持つアフィニティまたは単数または複数のイオン交換ビードを添加し得る。攪拌を継続して固形物の沈降を抑制するが、前記固形物は本実施例では、ポリマー、細胞や細胞破片、宿主細胞タンパク質、DNA等、及び所望の生体分子、等の不純物を含有する析出物が含まれ、液体中のまたはポリマーナイム他はポリマー上に取り込まれた不純物が確実に除去されるよう1回以上(好適なバッファを使用する等により)洗浄され得る。1回または複数回の洗浄ステップは、フィルタベース部100内の1つ以上の膜を通す濾過により実施され得、上澄み液がポート32bを介して送られて廃棄される。
4 スタンド
8 支持リム
10 支持部材
12 駆動機構
14 攪拌用アセンブリ
16 頂部
18 アーム
20 軸
22、22’ 胴部
24 クリップ
26 側壁
30a ポート
30b ポート
30c ポート
32a ポート
32b ポート
40 軸
42 パドル
50 入口
51 配管
100、100’ フィルタベース部
101 支持表面
102 溝
103 孔
106 O−リング
107 支持リング
110 多孔質フィルタ/膜
Claims (8)
- サンプルの培養または処理用アセンブリであって、
内側空間を有する第1容器と、
該第1容器にシール自在に固着するようになっており且つ少なくとも1つの膜を支持する第1ベース部と、
該第1ベース部の出口と、
前記第1ベース部の前記出口と流体連通するようになっている第2容器と、
該第2容器にシール自在に固着するようになっている第2ベース部と、
含むアセンブリ。 - 前記第2容器が少なくとも1つの膜を支持する請求項1のアセンブリ。
- 前記第1容器がバイオリアクターである請求項1のアセンブリ。
- 前記第1容器の同部内部に、サンプル攪拌手段を更に含む請求項1のアセンブリ。
- 不純物を含有する混合物から得た生体分子の精製方法であって、
所定条件下の混合物を提供すること、
前記所定条件下に前記混合物に可溶であり且つ前記生体分子に可逆的及び選択的に結合可能な1つ以上のポリマーを添加すること、
可溶化した前記1つ以上のポリマーを前記混合物全体に混入させること、
前記混合物における前記所定条件を変化させることにより、前記1つ以上のポリマー及び結合した生体分子を溶液から析出物として析出させること、
前記析出物を洗浄液と接触させて洗浄すると共に上澄み液を第1膜を通して濾過すること、
結合した生体分子を前記ポリマーから回収すると共に、外生体分子を第2膜を通して濾過すること、
を含む方法。 - 前記上澄み液の濾過及び前記生体分子の濾過が、同一装置で実施される請求項4の方法。
- 前記生体分子が、遺伝子組み換え抗体、遺伝子組み換えモノクロナール抗体、ポリクロナール抗体、ヒト化抗体、抗体断片、からなる群から選択した抗体である請求項5の方法。
- 前記生体分子がタンパク質である請求項5の方法。
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US20186508P | 2008-12-16 | 2008-12-16 | |
US61/201,865 | 2008-12-16 | ||
PCT/US2009/067097 WO2010074953A1 (en) | 2008-12-16 | 2009-12-08 | Stirred tank reactor and method |
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JP2013083213A Division JP5863699B2 (ja) | 2008-12-16 | 2013-04-11 | 不純物を含有する混合物から得た生体分子の精製方法 |
JP2014146868A Division JP5906284B2 (ja) | 2008-12-16 | 2014-07-17 | サンプルの培養または処理用アセンブリ |
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JP2011542238A Pending JP2012511929A (ja) | 2008-12-16 | 2009-12-08 | 攪拌タンク反応器及び方法 |
JP2013083213A Expired - Fee Related JP5863699B2 (ja) | 2008-12-16 | 2013-04-11 | 不純物を含有する混合物から得た生体分子の精製方法 |
JP2014146868A Expired - Fee Related JP5906284B2 (ja) | 2008-12-16 | 2014-07-17 | サンプルの培養または処理用アセンブリ |
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JP2014146868A Expired - Fee Related JP5906284B2 (ja) | 2008-12-16 | 2014-07-17 | サンプルの培養または処理用アセンブリ |
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JP (3) | JP2012511929A (ja) |
CN (2) | CN102257122B (ja) |
DK (1) | DK2370561T3 (ja) |
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- 2009-12-08 CN CN201510375479.1A patent/CN105037535A/zh active Pending
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Also Published As
Publication number | Publication date |
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JP5863699B2 (ja) | 2016-02-17 |
JP5906284B2 (ja) | 2016-04-20 |
EP2370561A4 (en) | 2013-07-24 |
SG171446A1 (en) | 2011-07-28 |
ES2749232T3 (es) | 2020-03-19 |
CN102257122B (zh) | 2015-07-29 |
JP2014195476A (ja) | 2014-10-16 |
WO2010074953A1 (en) | 2010-07-01 |
JP2013155187A (ja) | 2013-08-15 |
DK2370561T3 (da) | 2019-10-21 |
US20100190963A1 (en) | 2010-07-29 |
EP2370561A1 (en) | 2011-10-05 |
EP2370561B1 (en) | 2019-08-07 |
CN105037535A (zh) | 2015-11-11 |
US20130005950A1 (en) | 2013-01-03 |
US9803165B2 (en) | 2017-10-31 |
CN102257122A (zh) | 2011-11-23 |
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