JP4135892B2 - Residual blood recovery method and residual blood recovery device in leukocyte removal filter unit - Google Patents
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Description
【０００１】
【発明の属する技術分野】
本発明は、白血球除去フィルターユニットにおいてそこに残留する白血球が除去された血液をこのユニットから回収する方法に関する。本発明の方法によると、捕捉した白血球や血小板からのサイトカイン、メディエーターの放出を抑制し、白血球が除去された血液、少なくとも赤血球及び血漿を回収することができる。
さらに、本発明はこのような血液回収方法に用いる血液回収装置、及び血液回収システムに関する。
【０００２】
【従来の技術】
近年、輸血分野に於いて、血液製剤中の白血球混入による副作用が数多く知られている．この副作用予防のため、ポリエステル製の不織布やコットン綿などの材料を用いた白血球除去フィルターにより輸血用血液製剤からの白血球除去が行われている。
【０００３】
一方、全身性エリトマトーデス、悪性関節リウマチ、多発性硬化症、潰瘍性大腸炎、クローン病等の自己免疫性疾患、白血病、癌などの治療、或いは移植前の免疫抑制の目的で白血球を白血球除去フィルターを用いて選択的に除去する技術が進歩してきた．現在、不織布等をフィルターとして用いた白血球除去器が開発されており、これら白血球除去器に於いては高い白血球の除去能力が要求されている。
【０００４】
従来こうした白血球除去フィルターを用いる場合、白血球除去処理をした後に、白血球除去フィルター内に残った少なくとも赤血球及び血漿を生理食塩液で回収することが多かった。しかしながら、この回収方法は、白血球や血小板が白血球除去フィルター内の濾材である多孔質体、平膜、不織布、織布、粒子等に吸着することで活性化されているため、生理食塩液による温度変化、粘度変化、ｐH変化および生理食塩液の注入による流速変化などに例示される様々な外部からの物理的、化学的刺激により白血球や血小板からサイトカインやメディエーターが放出され、捕捉後の血液にサイトカインやメディエーターが高濃度に混入する可能性があった。
【０００５】
特許文献１には、セルロースジアセテートからなるビーズ状の白血球除去材を用いて、リンパ球に比し 炎症性細胞への親和性が高い担体と接触させ、かくして 得られる炎症性細胞が吸着除去された処理血液を当該患者に戻すことを特徴とする炎症性疾患の処置方法が記載されているが、気体を用いかつ生理的溶液を用いて白血球除去フィルターユニット内の残留血液を回収することは開示されていない。
【０００６】
一方、従来より、空気による回収も同時に行われてきたが、空気で回収する場合、白血球除去フィルター中の血液が空気で置換されがたい部分に空気が達すると、急激にカラム圧が上昇し、その後の血液を十分回収することが困難であった。
特許文献２には、白血球除去フィルターを挟んだバイパス回路を用いたフィルターユニットが開示されているが、生理的溶液を用いて白血球除去フィルター下流側の赤血球及び血漿を更に回収し、メディエアーターの濃度上昇なく回収することについて開示されていない。更に白血球除去フィルター下流の回路が複雑な場合十分な回収が困難であるため、十分とはいえない。
【０００７】
【特許文献１】
特開平7-178167号公報
【特許文献２】
特開2000-51347号公報
【０００８】
【発明が解決しようとする課題】
本発明は、白血球除去フィルターユニットにおいて白血球が捕捉されているが、まだ白血球除去フィルターユニット内に残留する白血球が除去された血液をこのユニットから回収するときに、カラムの差圧を上昇させることなく、サイトカイン、メディエーターの濃度上昇のリスクを抑制することができる、白血球除去フィルターユニットから残留する血液を回収する方法を提供することを目的とする。さらに、本発明は、この方法に用いる残留血液回収装置、血液回収システムを提供することを目的とする。
【０００９】
【課題を解決するための手段】
本発明者らは、上記課題を解決するために鋭意研究を重ねた結果、血液を回収する工程で、直接生理的溶液を用いず空気を用いることで、サイトカイン、メディエーターの濃度上昇のリスクを抑制することができることを見出し、さらにこの処理を行った後、白血球除去フィルター下流側の回路に残留する血液を生理的溶液で回収できることを確認し、本発明を完成するに至った。
【００１０】
すなわち、本発明は、少なくとも白血球除去フィルター及び該白血球除去フィルターの出口側に液密に接続された下流側回路とからなる白血球除去フィルターユニットの白血球除去フィルターに血液を流して白血球を捕捉させた後、白血球除去フィルターに残留する白血球が除去された血液を回収する方法において、白血球除去フィルター内に残留する白血球が除去された血液を気体により白血球除去フィルターの出口を過ぎるまで押し出す工程、及び白血球除去フィルターの出口と下流側回路の接続部位から無菌的に生理的溶液を流し込み下流側回路に残留する白血球が除去された血液を下流側回路外に押し出す工程とを有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球由来のメディエーターの濃度上昇を抑える、または捕捉された前記白血球中の顆粒球由来のメディエーターの濃度上昇を抑える、または捕捉された血小板由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液回収方法に関する。
【００１１】
また、本発明は、少なくとも白血球除去フィルター及び該白血球除去フィルターの出口側に液密に接続された下流側回路とからなる白血球除去フィルターユニットの白血球除去フィルターに血液を流して白血球を捕捉させた後、白血球除去フィルターに残留する白血球が除去された血液を回収する装置であって、少なくとも白血球を捕捉した白血球除去フィルター入口に気体を送気する手段、気体が白血球除去フィルター出口に達したとき気体を検出する手段、白血球除去フィルター出口と下流側回路の接続部位に生理的溶液を注入する手段を有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液を回収する残留血液回収装置に関する。
【００１２】
更に、本発明は、少なくとも白血球除去フィルター及び該白血球除去フィルターの出口側に液密に接続された下流側回路とからなる白血球除去フィルターユニットの白血球除去フィルターに血液を流して白血球を捕捉させた後、白血球除去フィルターに残留する白血球が除去された血液を回収する回路システムであって、少なくとも白血球を捕捉した白血球除去フィルターの上流側に気体を送気する回路、白血球除去フィルターの出口と下流側回路の接続部位に生理的溶液を注入する回路を有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液を回収する血液回収システムに関する。
【００１３】
【発明の実施の形態】
以下に、本発明を詳細に説明する。
本発明の白血球除去フィルターユニットとは、少なくとも白血球除去フィルター及び該白血球除去フィルターの出口側に液密に接続された下流側回路とからなるものをいう。
【００１４】
本発明の白血球除去フィルターとは、入口と出口のある容器に白血球に親和性のある白血球除去材を充填したものをいう。白血球除去材は、白血球を除去できるものであれば特に限定はなく各種のものを用いることができ、多孔質体、平膜、不織布、織布、粒子等が例示できる。白血球除去の観点より好ましい形状は、多孔質体、不織布、織布、粒子が挙げられ、最も好ましくは不織布が良好に用いられる。
【００１５】
白血球除去材の形状が不織布の場合、フィラメントは、モノフィラメントでもマルチフィラメントでも構わないし、多孔質フィラメントでも異型フィラメントでも構わない。
【００１６】
また不織布では、平均繊維直径が 0.8μm以上50μm未満の不織布が好ましい。繊維径が大きくなると基材の表面積確保が困難となり、白血球の吸着面積が減少し、白血球の除去性能が低下するため好ましくない。
【００１７】
一方、繊維径が小さくなると白血球除去材の目詰まりが発生するリスクが高まり好ましくない。以上の観点より、更に好ましい平均繊維直径は1.0μm以上30μm未満、最も好ましくは、1.5μm以上25μm未満である。
また、白血球除去フィルターには、アグリゲート等の除去を目的に白血球除去材の血液の導入部側に平均繊維直径25μm以上100μm未満のプレフィルターを配していても良好に用いられる。
【００１８】
更に、不織布の場合、白血球の除去率を上げる上でその嵩密度が0.10g/cm3以上0.45g/cm3未満であることも重要である。嵩密度が0.10g/cm3未満の場合白血球の除去性能が低下するため好ましくない。一方嵩密度が0.45g/cm3以上の場合、極端に目詰まりしやすくなるため好ましくない。更に、上記の観点より好ましくは0.12g/cm3以上0.45g/cm3未満、最も好ましくは0.13g/cm3以上0.40g/cm3未満である。
【００１９】
白血球除去材の形状が粒状の場合、球状、多角球状等いかなる形状であっても有用に用いられる。また表面は平滑であっても、凹凸があっても白血球を除去できる表面であれば有用に用いられる。また、粒径は、50μm以上10mm以下であれば有用に用いられる。粒径は50μm未満であると粒子の流出防止が困難であり好ましくない。粒径が10mmより大きい場合十分な表面積が得られず好ましくない。従って、好ましくは、80μm以上8mm未満、最も好ましくは100μm以上６ｍｍ未満である。
【００２０】
本発明の白血球除去材の材質としては、血球にダメージを与えにくいものであれば特に限定はなく各種のものを用いることができ、有機高分子、無機高分子、金属等が例示できる。その中でも有機高分子は切断などの加工性に優れるため好ましい素材である。有機高分子等の材質は、セルロース、セルロースモノアセテート、セルロースジアセテート、セルローストリアセテート等のセルロース及び／またはその誘導体等の天然高分子、ポリエチレンテレフタレート、ポリブチレンテレフタレート等のポリエステル、ポリエチレン、ポリプロピレン等のポリオレフィン、ポリフッ化ビニリデン、ポリアミド、ポリイミド、ポリウレタン、ポリスルホン、ポリアクリロニトリル等の高分子材料を例示できる。またこれらに親水性を付与する目的で表面をコーティング、放射線グラフト等の方法で高分子材料で修飾した材料も有用に用いられる。
【００２１】
これらの材料が表面修飾なしで白血球に対して親和性を有する場合は、そのまま用いることも可能である。また表面修飾なしでそれら機能を発揮できない場合は、機能発揮を目的としてコーティング等の表面改質をして用いることが好ましい。
特に、白血球を除去する目的で、表面にコーティング等の処理によって改質されているものが好ましく用いられる。
白血球除去材として、血小板の吸着性が低い材料を表面にコーティング等の処理することで、血小板の通過性を付与した白血球選択除去材を使用することも可能である。
一方、血小板の吸着性を高める材料の表面コーティング等の処理により、血小板の除去性を付与することも可能であり、いずれの白血球除去材も有用に用いられる。
【００２２】
以上、白血球除去材の形状と材質について述べたが、これらを総合すると、白血球除去材の好ましい実例として、ポリエチレンテレフタレート、ポリブチレンテレフタレート等のポリエステル不織布、セルロース、セルロースモノアセテート、セルロースジアセテート、セルローストリアセテート等のセルロース系ビーズあるいはこれら表面をコーティング、放射線グラフト等の方法で表面修飾した材料が好ましく用いられるが、これに限定されるものではない。
【００２３】
また、本発明の白血球除去フィルターは、白血球除去率が30％以上であるときに有用に用いられる。白血球除去率が高い場合は、白血球除去フィルターに捕捉される白血球が多いことより、白血球が除去された血液を回収する際に、メディエーター等の上昇のリスクがより高まり、本発明のメディエーター抑制効果が発揮されるため好ましい。
従って、更に好ましい白血球除去率は50％以上、最も好ましくは70％以上のときに有用に用いられる。
また、リンパ球、顆粒球、単球等の特定の白血球分画を除去する場合は、特定の白血球分画での白血球除去率を用いることができる。
白血球除去率は、処理中あるいは目標血液処理時の白血球除去フィルター前後の血液をサンプリングし求めることができるし、また複数回サンプリング可能な場合はそれらの平均値を用いて求めても好ましく用いられる。
【００２４】
本発明の白血球除去フィルターの出口側に液密に接続された下流側回路は、白血球除去フィルターの出口に液密に接続された回路であれば、何れの形状であっても有効に用いられる。特に、生理的溶液を注入できる回路が接続されている場合最も有効に用いられる。回路内断面の形状は円状、楕円状、多角状等のいずれも有用に用いられる。残留血液の回収の面より円状、楕円状が特に好ましく用いられる。回路材質は、血液成分に影響を与えない材料であれば何れも有用にもちいることができる。
【００２５】
また、下流側回路はチューブに加えて、ドリップチャンバー、加温バッグ、採血ポート等何れの付属部品を有していても有用に用いられる。特に、本発明では、これら付属部品が付属して、気体での回収が困難な場合においても、生理的溶液を用いて残留する血液を回収できるため非常に有効に用いられる。
【００２６】
本発明において、血液とは、全血、顆粒球含有液、リンパ球含有液、単球含有液又はこれら全てを含む等の白血球含有液、血小板含有液などの血球成分を含有する血液も含む。また、アレルギー、炎症性疾患等の活性化した血液について用いた場合、特に有用に用いられる。
【００２７】
これら血液を処理するときは、血液の抗凝固目的で抗凝固剤を血液中に加えることができる。抗凝固剤を例示すると、抗凝固活性を有する化合物であれば、特に限定されないが、ヘパリン、低分子ヘパリン、メシル酸ナファモスタット、メシル酸ガベキサート等の蛋白分解酵素阻害剤、ACD-A、CPD等のクエン酸等が好適な例として挙げられ、最も好ましくは、ヘパリン、メシル酸ナファモスタット、クエン酸が良好に用いられる。
【００２８】
本発明の回収とは、気体により白血球除去フィルター内に残存する白血球が除去された血液をフィルターの下流側回路に押出す工程、及び続いて生理的溶液により白血球除去フィルター下流側回路内に残留した白血球が除去された血液を上記回路外に押出す工程を言う。また、この場合、回収方向は、白血球除去フィルターの血液の入口側より出口側へ回収する。
【００２９】
本発明の白血球除去フィルターユニット内の残留血液回収方法は白血球のみを選択的に除去し、血小板を回収する場合、白血球由来のメディエーターの濃度上昇を抑制するため好ましく用いられる。
特に、白血球中の顆粒球を効率的に除去する場合において、顆粒球から即座に放出されるヒスタミン、ロイコトリエン等のメディエーターの放出を抑制できるため好ましく用いられる。
【００３０】
また、本発明の白血球除去フィルターユニット内の残留血液回収方法は白血球のみならず、血小板を同時に除去する場合、白血球由来のメディエーターに加えて、セロトニンやトロンボキサン等の血小板由来のメディエーターを抑制するため更に有用に用いられる。
【００３１】
本発明の気体とは、血液に悪影響を与えない気体であり、空気、窒素ガス、酸素ガス、炭酸ガスやこれら混合ガスが有用に用いられる。好ましくは、使用の簡便さより空気が有用に用いられる。
【００３２】
本発明で用いる生理的溶液は、血液及び生体に対して悪影響を与えない溶液をいう。例示すると生理食塩液、リンゲル液、乳酸リンゲル液、酢酸リンゲル液、タイロード液等の生理的塩類溶液、ブドウ糖、デキストラン等の糖含有液、クエン酸、ヘパリン、メシル酸ナファモスタット、メシル酸ガベキサート等の蛋白分解酵素阻害剤等の抗凝固剤含有液を言う。抗凝固剤含有液は他の生理的溶液により希釈されても有用に用いられる。
【００３３】
また、上記に例示した生理的塩類溶液、糖含有液、抗凝固剤含有液を任意の割合で混合した混合液であっても良好に用いられる。更に、他の生体に対して悪影響を与えない溶液で、上記に例示した生理的塩類溶液、糖含有液や抗凝固剤含有液を少なくとも１つ以上加えた混合液を希釈した液も好ましく用いられる。取扱い性の面より、好ましくは生理的塩類溶液、糖含有液や抗凝固剤含有液はそのまま、あるいは任意の割合で混合した混合液が良好に用いられ、より好ましくは、生理的塩類溶液、糖含有液や抗凝固剤含有液をそのまま用いることである。
【００３４】
以下に本発明の血液回収装置について詳細に説明する。
本発明の血液回収装置は、少なくとも白血球除去フィルターと白血球除去フィルターの出口側に液密に接続された下流側回路からなる白血球除去フィルターユニットの白血球除去フィルターに血液を流し、該白血球除去フィルターに白血球を捕捉させた後、白血球を捕捉した白血球除去フィルターに残留する少なくとも赤血球及び血漿を回収する血液回収装置であって、少なくとも白血球を捕捉した白血球除去フィルター入口に気体を送気する手段、気体が白血球除去フィルター出口に達したとき気体を検出する手段、白血球除去フィルター出口と下流側回路の接続部位に生理的溶液を注入する手段を有することを特徴とする白血球除去フィルターユニット内の残留血液を回収する残留血液回収装置に関する。
【００３５】
本発明の白血球を捕捉した白血球除去フィルター入口に気体を送気する手段とは、白血球を捕捉させた後の白血球除去フィルター入口に気体を一定あるいは可変の流速にて送気できる手段であればいかなる手段も用いることができる。例えば、白血球を捕捉する際に用いる血液送液手段、送風ポンプ、圧縮空気、及びエアーコンプレッサー等が有用に用いられる。特に、白血球を除去する際に用いる血液送液手段が、簡便に用いることができ、例示すると、血液ポンプ、ローラーポンプ、フィンガーポンプ、シリンジポンプ、落差による送液等が挙げられる。流速を一定にする観点より血液ポンプ、ローラーポンプが有用に用いられる。一方、簡便さの観点より落差が有用に用いられる。
【００３６】
本発明の送気手段は、流速10ml/min以上100ml/min未満が好ましくもちいられる。流速が10ml/min未満の場合、送気に時間を要し好ましくない。一方100ml/min以上の場合、送気時に圧がかかり白血球除去フィルター内の圧上昇により赤血球の溶血のリスクも高まり好ましくない。以上の観点より、より好ましい流速は12ml/min以上90ml/min未満、最も好ましくは15ml/min以上80ml/min未満である。
【００３７】
本発明の気体が白血球除去フィルター出口に達したとき気体を検出する手段は、白血球除去フィルター出口より気体が排出されたことを検知できる手段であれば何れの検出器でも有用に用いられる。好ましい例を例示すると、気泡検知器、圧力センサー等が挙げられる。
【００３８】
圧力センサーや圧力計による気体排出の検知は、白血球除去フィルター内に空気が送気され、内部の液体がほとんど回収されるとフィルターの内部が血液から気体に置換される場合、バブルポイント様の排出圧力が発生し、この圧力を検出することで容易に気体排出が検出できる。不織布状の白血球除去フィルター材を用いた場合、白血球除去フィルターの入口と出口の差圧が急激に上昇するため特に有用に用いることができる。この場合、差圧が100mmHg以上となると血液の排出が終了し気体のでる直前と判断できる。
【００３９】
本発明の白血球除去フィルター出口以降に生理的溶液を注入する手段とは、白血球除去フィルター出口下流側回路に生理的溶液を注入できる手段であれば何れの注入方法も有用に用いられる。注入する手段としては、上記フィルター下流側回路でフィルター出口近傍に設けられた分岐回路より、落差、血液ポンプ、ローラーポンプ、フィンガーポンプ、シリンジポンプ、ペリスタポンプ等の手段で一定あるいは可変の流量で生理的溶液を注入する手段が有用に用いられる。
本発明の白血球除去フィルターの出口と下流側回路の接続部位とは、白血球除去フィルターの出口とこれに接続する下流側回路の間に設けられていても良いし、下流側回路の白血球除去フィルターの出口に近い位置に設けられていても好ましく用いられる。
【００４０】
本血液回収装置には、ドリップチャンバー、ポンプ用チューブ、圧力測定チューブ、圧力測定血液回収装置、ポンプ等を適宜加えて用いることができる。
【００４１】
以下に、本発明の血液回収システムについて説明する。
本発明の回路システムは、少なくとも白血球除去フィルター及び該白血球除去フィルターの出口側に液密に接続された下流側回路とからなる白血球除去フィルターユニットの白血球除去フィルターに血液を流し、該白血球除去フィルターに白血球を捕捉させた後、白血球を捕捉した白血球除去フィルターに残留する白血球が除去された血液を回収する回路システムであって、少なくとも白血球を捕捉した白血球除去フィルターの上流側に気体を送気する回路、白血球除去フィルターの出口と下流側回路の接続部位に生理的溶液を注入する回路を有することを特徴とする白血球を捕捉した白血球除去フィルター内の残留血液を回収する血液回収システムである。
本発明における白血球を捕捉した白血球除去フィルターに残留する白血球が除去された血液には、少なくとも赤血球及び血漿が含まれる。
【００４２】
本発明の白血球を捕捉した白血球除去フィルター上流側に気体を送気する回路とは、気体を送気できる気密な回路であり、気体の入口を有している必要がある。気体を送気する回路は白血球除去フィルターの上流側に配されている必要がある。気体の入口には必要に応じて、ウイルスや細菌、微粒子を通さないフィルターを配していることが好ましい。白血球除去フィルター上流側に気体を送気する回路を新たに設けずに、血液の流入口から血液入口側回路をはずすことで代用しても有用に用いられる。
【００４３】
白血球除去フィルターの出口と下流側回路の接続部位に生理的溶液を注入する回路とは、生理的溶液を送液できる液密な回路であり、生理的溶液の入口を有している必要がある。生理的溶液を送液する回路は白血球除去フィルターの直後に配されていることが好ましい。生理的溶液の入口には必要に応じて、スパイク針、注射針等を配しても良く、この場合、生理的溶液等の入った溶液バッグに液密に接合することが可能となる。また、ルアーロック等によりシリンジポンプに接合することも可能である。更に、あらかじめ回路末端に、生理的溶液の充填された容器を配していても有用に用いられる。また、白血球除去フィルター直後に針刺し用ポートを配して、このポートから注入することも可能である。空気の血液への混入を抑制するため生理的溶液の入口と血液の出口との間にはドリップチャンバーを配することが好ましい。
【００４４】
【実施例】
以下に、本発明の実施例を示すが、本発明はこれに限定されるものではない。
【実施例１】
平均繊維直径2.7μmのポリエステル繊維よりなる不織布（目付90g/m2、厚さ0.42mm）を75mmの円形に切断し、上下にポアサイズ90μmのポリエチレンフィルタで挟み、10枚をカラムに充填した。
抗凝固剤としてメシル酸ナファモスタットを添加したヒト新鮮血液（白血球数：8.200/μL、血小板数：300.000/μL）(メシル酸ナファモスタット濃度:0.012mg/ml)400mLを血液ポンプにて20mL/分の一定流速で室温にて流し、白血球を除去した。
【００４５】
血液流入口を開放し、血液ポンプにて20ml/minでカラム出口まで空気を流した。気泡検知器により空気の流出を確認後、白血球除去フィルター出口直後の回路より生理食塩液をフィンガーポンプにて流速20ml/minで30ml流し、白血球除去フィルター下流側の白血球除去された血液を回収した。処理前の血液と気体回収液20ml及び生理食塩液回収液30mlをあわせた50mlの回収液のヒスタミン濃度及びセロトニン濃度をＥＩＡキット（イムノテック社製）により測定した。ヒスタミン及びセロトニン濃度より、それぞれの濃度上昇率を下式(1)、(2)により求めた。結果を表１に示す。
また、白血球除去フィルター処理前の血液と処理後の血液中の白血球数を多項目自動血球分析装置(Sysmex社製)により測定し、白血球除去率を下式(3)より求めた。結果を表１に示す。
【００４６】
ヒスタミン濃度上昇率＝回収液のヒスタミン濃度／処理前血液のヒスタミン濃度・・・・・式(1)
セロトニン濃度上昇率＝回収液のセロトニン濃度／処理前血液のセロトニン濃度・・・・・式(2)
白血球除去率＝{１−（白血球除去フィルター処理後の白血球数/処理前血液の白血球数）}×100・・・・・式(3)
【００４７】
【比較例１】
白血球除去フィルターから空気を用いる代りに生理食塩液50mlを用いて白血球除去された血液50mlを回収し、回収液とした。処理前の血液と回収液のヒスタミン及びセロトニン濃度、及び白血球除去率は実施例１と全く同じ操作を行い、実施例1記載の式(1)、(2)及び(3)を用いてそれぞれヒスタミン濃度上昇率、セロトニン濃度上昇率及び白血球除去率を求めた。結果を表１に示す。
【００４８】
【表１】
【実施例２】
平均繊維直径2.7μmのポリエステルの繊維よりなる不織布（90g/m2目付）を幅150mm、長さ760mmに切断し、直径3.4mmのポリエチレン製の円筒状メッシュの周囲に巻いた。次いで平均繊維直径12μmのポリエステル繊維からなる不織布(30g/m2目付)を第1プレフィルターとし、幅150mm、長さ１、300mmで巻いた。更に平均繊維直径33μmのポリエステル繊維からなる不織布(50g/m2目付)を第２プレフィルターとし、幅150mm、長さ1.300mmで巻いた。外側にポリエチレン製のメッシュを幅150mm、長さ150mm巻いた。この円筒直径は39mmであった。この円筒の両端をウレタンで閉塞し、天井部と底部にそれぞれ血液の入口と出口を有する内径41mmの円筒状ポリカーボネート容器に、円筒の外周面が容器の血液入口に、内周面が血液の出口にそれぞれ通じるように納め、白血球除去フィルターとした。
【００５０】
抗凝固剤としてヘパリンを添加した牛新鮮血液（白血球数：5.300/μL、血小板数：170.000/μL）(ヘパリン濃度:1000IU/L)3000mLを血液ポンプにて50mL/分の一定流速で室温にて流し、白血球を除去した。血液流入口を開放し、50ml/minでカラム出口まで空気を流した。圧力検知器により空気混入による圧上昇を検知後、白血球除去フィルター出口直後の回路より生理食塩液200mlを流速50ml/minで流し、白血球除去フィルター以後の回路に残留する白血球が除去された血液200mlを回収した。処理前の血液と気体回収液200ml及び生理食塩液回収液180mlをあせた380mlの回収液のヒスタミン濃度及びセロトニン濃度をEIAキット（イムノテック社製）により測定した。実施例１記載の式(1)及び(2)を用いてそれぞれヒスタミン濃度上昇率、セロトニン濃度上昇率を求めた。また、実施例１と同様の方法で、白血球除去フィルター処理後血液の白血球数及び処理前血液の白血球数を1000ml処理時、2000ml処理時、3000ml処理時それぞれで求め、これら値を平均し、それぞれを白血球除去フィルター処理後血液の白血球数及び処理前血液の白血球数とし、実施例１記載の式(3)を用いて白血球除去率を用いた。
【００５１】
【比較例２】
実施例２と同じ白血球除去フィルターを用い、白血球除去フィルター前から空気を用いる代りに5％ブドウ糖液380mlを用いて白血球除去された血液を回収し、回収液とした。ヒスタミン及びセロトニン濃度、及び白血球数のサンプリングは実施例２と全く同じ操作を行い、実施例１記載の式(1)、(2)、及び(3)を用いてそれぞれヒスタミン濃度上昇率、セロトニン濃度上昇率及び白血球除去率を求めた。結果を表２に示す。
【００５２】
【表２】
【発明の効果】
本発明によると、白血球除去フィルターユニット内に残留する白血球を除去された血液(赤血球、血漿等)をサイトカイン、メディエーターの放出を抑制して回収することができる。[0001]
BACKGROUND OF THE INVENTION
The present invention relates to a method for recovering blood from which leukocytes remaining in the leukocyte removal filter unit have been removed. According to the method of the present invention, release of cytokines and mediators from captured leukocytes and platelets can be suppressed, and blood from which leukocytes have been removed, at least erythrocytes and plasma, can be collected.
Furthermore, the present invention relates to a blood collection device and a blood collection system used for such a blood collection method.
[0002]
[Prior art]
In recent years, in the field of blood transfusion, many side effects due to leukocyte contamination in blood products have been known. In order to prevent this side effect, leukocyte removal from blood products for blood transfusion is performed by a leukocyte removal filter using a material such as polyester nonwoven fabric or cotton cotton.
[0003]
On the other hand, leukocyte removal filter for the treatment of systemic lupus erythematosus, malignant rheumatoid arthritis, multiple sclerosis, ulcerative colitis, autoimmune diseases such as Crohn's disease, leukemia, cancer, etc. or immunosuppression before transplantation The technology for selective removal by using sapphire has progressed. Currently, leukocyte removers using a nonwoven fabric or the like as a filter have been developed, and these leukocyte removers are required to have high leukocyte removal ability.
[0004]
Conventionally, when such a leukocyte removal filter is used, at least red blood cells and plasma remaining in the leukocyte removal filter are often collected with a physiological saline solution after the leukocyte removal treatment. However, this recovery method is activated by adsorbing leukocytes and platelets to porous bodies, flat membranes, non-woven fabrics, woven fabrics, particles, etc., which are filter media in the leukocyte removal filter. Cytokines and mediators are released from leukocytes and platelets by various external physical and chemical stimuli exemplified by changes, viscosity changes, pH changes and flow rate changes by injection of physiological saline, etc., and cytokines are captured in the blood after capture And mediators could be mixed in at high concentrations.
[0005]
In Patent Document 1, a bead-shaped leukocyte-removing material made of cellulose diacetate is used to contact a carrier having a higher affinity for inflammatory cells than lymphocytes, and the inflammatory cells thus obtained are adsorbed and removed. A method for treating inflammatory diseases characterized by returning treated blood to the patient is disclosed, but it is disclosed to collect residual blood in a leukocyte removal filter unit using a gas and using a physiological solution It has not been.
[0006]
On the other hand, conventionally, recovery by air has also been performed at the same time, but when recovering with air, if the air reaches the part where the blood in the leukocyte removal filter is difficult to be replaced with air, the column pressure suddenly increases, It was difficult to sufficiently collect the subsequent blood.
Patent Document 2 discloses a filter unit using a bypass circuit with a leukocyte removal filter sandwiched between them, but further recovers red blood cells and plasma downstream of the leukocyte removal filter using a physiological solution, There is no disclosure about recovery without increasing the concentration. Furthermore, if the circuit downstream of the leukocyte removal filter is complicated, it cannot be said that it is sufficient because sufficient recovery is difficult.
[0007]
[Patent Document 1]
JP-A-7-178167
[Patent Document 2]
JP 2000-51347 A
[0008]
[Problems to be solved by the invention]
In the present invention, leukocytes are captured in the leukocyte removal filter unit, but when the blood from which leukocytes still remaining in the leukocyte removal filter unit are removed is collected from this unit, the differential pressure of the column is not increased. An object of the present invention is to provide a method for recovering residual blood from a leukocyte removal filter unit, which can suppress the risk of an increase in the concentration of cytokines and mediators. Furthermore, this invention aims at providing the residual blood collection | recovery apparatus and blood collection system which are used for this method.
[0009]
[Means for Solving the Problems]
As a result of intensive studies to solve the above problems, the present inventors have suppressed the risk of increased concentrations of cytokines and mediators by using air instead of a direct physiological solution in the process of collecting blood. After performing this process, it was confirmed that the blood remaining in the circuit on the downstream side of the leukocyte removal filter can be collected with a physiological solution, and the present invention has been completed.
[0010]
That is, the present invention captures leukocytes by flowing blood through a leukocyte removal filter of a leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit liquid-tightly connected to the outlet side of the leukocyte removal filter. In the method of recovering blood from which leukocytes remaining on the leukocyte removal filter are collected, the step of pushing the blood from which leukocytes remaining in the leukocyte removal filter have been removed by gas until the outlet of the leukocyte removal filter is passed, and the leukocyte removal filter A process in which a physiological solution is aseptically poured from the connection portion of the outlet of the downstream circuit and the downstream circuit, and the blood from which the white blood cells remaining in the downstream circuit have been removed is pushed out of the downstream circuit. And, when collecting blood remaining in the leukocyte removal filter, suppresses an increase in the concentration of the mediator derived from the captured leukocyte, suppresses an increase in the concentration of the mediator derived from granulocyte in the captured leukocyte, or Suppresses the concentration increase of trapped platelet-derived mediators The present invention relates to a method for collecting residual blood in a leukocyte removal filter unit.
[0011]
In addition, the present invention allows blood to flow through a leukocyte removal filter of a leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit that is liquid-tightly connected to the outlet side of the leukocyte removal filter to capture leukocytes. , A device for collecting blood from which leukocytes remaining on the leukocyte removal filter have been removed, at least means for sending gas to the leukocyte removal filter inlet that has captured leukocytes, and gas when the gas reaches the leukocyte removal filter outlet Means for detection, means for injecting physiological solution into the connection site between the leukocyte removal filter outlet and the downstream circuit When collecting blood remaining on the leukocyte removal filter, the increase in the concentration of the captured leukocyte-derived mediator is suppressed. The present invention relates to a residual blood collecting apparatus for collecting residual blood in a leukocyte removal filter unit.
[0012]
Furthermore, the present invention provides a method in which blood is passed through a leukocyte removal filter of a leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit connected in a liquid-tight manner to the outlet side of the leukocyte removal filter to capture leukocytes. , A circuit system for collecting blood from which leukocytes remaining on the leukocyte removal filter have been removed, at least a circuit for supplying gas to the upstream side of the leukocyte removal filter that has captured leukocytes, and an outlet and downstream circuit for the leukocyte removal filter A circuit for injecting physiological solution into the connection site of When collecting blood remaining on the leukocyte removal filter, the increase in the concentration of the captured leukocyte-derived mediator is suppressed. The present invention relates to a blood collection system for collecting residual blood in a leukocyte removal filter unit.
[0013]
DETAILED DESCRIPTION OF THE INVENTION
The present invention is described in detail below.
The leukocyte removal filter unit of the present invention refers to a unit comprising at least a leukocyte removal filter and a downstream circuit connected fluid-tightly to the outlet side of the leukocyte removal filter.
[0014]
The leukocyte removal filter of the present invention is a filter in which a container having an inlet and an outlet is filled with a leukocyte removing material having affinity for leukocytes. The leukocyte removing material is not particularly limited as long as it can remove leukocytes, and various materials can be used, and examples thereof include a porous body, a flat membrane, a nonwoven fabric, a woven fabric, and particles. Preferred shapes from the viewpoint of leukocyte removal include porous bodies, non-woven fabrics, woven fabrics, and particles, and most preferred are non-woven fabrics.
[0015]
When the shape of the leukocyte removing material is a non-woven fabric, the filament may be a monofilament or a multifilament, and may be a porous filament or an irregular filament.
[0016]
The nonwoven fabric is preferably a nonwoven fabric having an average fiber diameter of 0.8 μm or more and less than 50 μm. When the fiber diameter is increased, it is difficult to ensure the surface area of the substrate, the leukocyte adsorption area is decreased, and the leukocyte removal performance is lowered, which is not preferable.
[0017]
On the other hand, if the fiber diameter is small, the risk of clogging of the leukocyte removal material increases, which is not preferable. From the above viewpoint, the more preferable average fiber diameter is 1.0 μm or more and less than 30 μm, and most preferably 1.5 μm or more and less than 25 μm.
Further, the leukocyte removal filter can be used satisfactorily even if a prefilter having an average fiber diameter of 25 μm or more and less than 100 μm is disposed on the blood introduction side of the leukocyte removal material for the purpose of removing aggregates and the like.
[0018]
Furthermore, in the case of non-woven fabric, the bulk density is 0.10 g / cm to increase the leukocyte removal rate. Three 0.45g / cm Three It is also important to be less than. Bulk density is 0.10g / cm Three If the ratio is less than 1, the leukocyte removal performance is lowered, which is not preferable. On the other hand, the bulk density is 0.45 g / cm Three In the above case, clogging is extremely easy, which is not preferable. Further, from the above viewpoint, preferably 0.12 g / cm. Three 0.45g / cm Three Less than, most preferably 0.13 g / cm Three 0.40 g / cm Three Is less than.
[0019]
When the shape of the leukocyte removing material is granular, any shape such as a spherical shape or a polygonal spherical shape is usefully used. Also, any surface that can remove leukocytes even if the surface is smooth or uneven is useful. Further, the particle diameter is useful if it is 50 μm or more and 10 mm or less. If the particle size is less than 50 μm, it is difficult to prevent the particles from flowing out, which is not preferable. When the particle size is larger than 10 mm, a sufficient surface area cannot be obtained, which is not preferable. Therefore, it is preferably 80 μm or more and less than 8 mm, and most preferably 100 μm or more and less than 6 mm.
[0020]
The material of the leukocyte-removing material of the present invention is not particularly limited as long as it does not easily damage blood cells, and various materials can be used, and examples thereof include organic polymers, inorganic polymers, and metals. Among them, an organic polymer is a preferable material because it has excellent processability such as cutting. Materials such as organic polymer include natural polymers such as cellulose, cellulose monoacetate, cellulose diacetate, cellulose triacetate and the like, and polyesters such as polyethylene terephthalate and polybutylene terephthalate, polyolefins such as polyethylene and polypropylene And polymer materials such as polyvinylidene fluoride, polyamide, polyimide, polyurethane, polysulfone, and polyacrylonitrile. For the purpose of imparting hydrophilicity to these, materials whose surfaces are modified with a polymer material by a method such as coating or radiation grafting are also useful.
[0021]
When these materials have affinity for leukocytes without surface modification, they can be used as they are. In addition, when these functions cannot be performed without surface modification, it is preferable to use after surface modification such as coating for the purpose of function display.
In particular, for the purpose of removing leukocytes, those whose surface is modified by a treatment such as coating are preferably used.
As the leukocyte removing material, a leukocyte selective removing material imparted with the ability to pass platelets can be used by coating the surface with a material having low platelet adsorptivity.
On the other hand, it is possible to impart platelet removability by a treatment such as surface coating of a material that enhances platelet adsorptivity, and any leukocyte-removing material is useful.
[0022]
As mentioned above, the shape and the material of the leukocyte-removing material have been described. When these are combined, preferred examples of the leukocyte-removing material include polyester nonwoven fabrics such as polyethylene terephthalate and polybutylene terephthalate, cellulose, cellulose monoacetate, cellulose diacetate, and cellulose triacetate. Cellulosic beads such as these or materials whose surfaces are modified by a method such as coating or radiation grafting are preferably used, but are not limited thereto.
[0023]
The leukocyte removal filter of the present invention is usefully used when the leukocyte removal rate is 30% or more. When the leukocyte removal rate is high, more leukocytes are captured by the leukocyte removal filter, so that when the blood from which leukocytes have been removed is collected, there is a higher risk of mediators and the like, and the mediator inhibitory effect of the present invention is increased. Since it is exhibited, it is preferable.
Therefore, it is useful when the leukocyte removal rate is more preferably 50% or more, and most preferably 70% or more.
Moreover, when removing specific leukocyte fractions such as lymphocytes, granulocytes, monocytes and the like, the leukocyte removal rate in the specific leukocyte fraction can be used.
The leukocyte removal rate can be obtained by sampling the blood before and after the leukocyte removal filter during the treatment or at the time of target blood treatment, and when it can be sampled a plurality of times, it is preferably used even if it is obtained using the average value thereof.
[0024]
The downstream circuit connected liquid-tightly to the outlet side of the leukocyte removal filter of the present invention can be effectively used in any shape as long as it is a circuit connected liquid-tightly to the outlet of the leukocyte removal filter. Particularly, it is most effectively used when a circuit capable of injecting a physiological solution is connected. Any of a circular shape, an elliptical shape, a polygonal shape, and the like is usefully used for the cross section in the circuit. A circular shape and an elliptical shape are particularly preferably used from the viewpoint of recovery of residual blood. As the circuit material, any material that does not affect blood components can be used.
[0025]
In addition to the tube, the downstream circuit is useful even if it has any accessory parts such as a drip chamber, a heating bag, and a blood collection port. In particular, in the present invention, even when these accessory parts are attached and recovery with gas is difficult, the remaining blood can be recovered using a physiological solution, so that it can be used very effectively.
[0026]
In the present invention, blood includes blood containing blood cell components such as whole blood, granulocyte-containing liquid, lymphocyte-containing liquid, monocyte-containing liquid, or a leukocyte-containing liquid such as those containing all of them, and a platelet-containing liquid. In addition, it is particularly useful when used for activated blood such as allergies and inflammatory diseases.
[0027]
When processing these blood, an anticoagulant can be added to the blood for the purpose of anticoagulation of the blood. Examples of anticoagulants include, but are not limited to, as long as they have anticoagulant activity, such as heparin, low molecular heparin, nafamostat mesylate, gabexate mesylate, ACD-A, CPD, etc. Citric acid and the like are preferable examples, and heparin, nafamostat mesylate, and citric acid are most preferably used.
[0028]
The recovery of the present invention refers to a step of extruding blood from which leukocytes remaining in the leukocyte removal filter have been removed by gas into the downstream circuit of the filter, and subsequently remaining in the downstream circuit of the leukocyte removal filter with a physiological solution. This refers to the process of extruding the blood from which white blood cells have been removed out of the circuit. In this case, the recovery direction is recovery from the blood inlet side to the outlet side of the leukocyte removal filter.
[0029]
The method for collecting residual blood in the leukocyte removal filter unit of the present invention is preferably used in the case of selectively removing only white blood cells and collecting platelets, in order to suppress an increase in the concentration of leukocyte-derived mediators.
Particularly, when granulocytes in leukocytes are efficiently removed, it is preferably used because release of mediators such as histamine and leukotriene that are immediately released from granulocytes can be suppressed.
[0030]
In addition, the method for collecting residual blood in the leukocyte removal filter unit of the present invention suppresses not only leukocytes but also platelet-derived mediators such as serotonin and thromboxane in addition to leukocyte-derived mediators when removing platelets simultaneously. Furthermore, it is usefully used.
[0031]
The gas of the present invention is a gas that does not adversely affect blood, and air, nitrogen gas, oxygen gas, carbon dioxide gas, or a mixed gas thereof is usefully used. Preferably, air is useful because of its ease of use.
[0032]
The physiological solution used in the present invention refers to a solution that does not adversely affect blood and a living body. Examples include physiological salt solutions such as physiological saline, Ringer's solution, Lactated Ringer's solution, Ringer's acetate, Tyrode's solution, sugar-containing solutions such as glucose and dextran, and proteolysis such as citric acid, heparin, nafamostat mesylate, and gabexate mesylate. An anticoagulant-containing solution such as an enzyme inhibitor. The anticoagulant-containing liquid is useful even if diluted with other physiological solutions.
[0033]
Moreover, even if it is the liquid mixture which mixed the physiological salt solution illustrated above, the sugar containing liquid, and the anticoagulant containing liquid in arbitrary ratios, it is used favorably. In addition, a solution obtained by diluting a mixed solution obtained by adding at least one physiological salt solution, sugar-containing solution or anticoagulant-containing solution exemplified above with a solution that does not adversely affect other living bodies is also preferably used. . From the viewpoint of handling, preferably a physiological salt solution, a sugar-containing solution or an anticoagulant-containing solution is used as it is or a mixed solution mixed in an arbitrary ratio is preferably used, and more preferably a physiological salt solution, a sugar solution It is to use the containing liquid or the anticoagulant-containing liquid as it is.
[0034]
The blood collection device of the present invention will be described in detail below.
The blood collection device of the present invention allows blood to flow through a leukocyte removal filter of a leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit connected fluid-tightly to the outlet side of the leukocyte removal filter, Is a blood collection device that collects at least red blood cells and plasma remaining in the leukocyte removal filter that has captured leukocytes, and means for supplying gas to the inlet of the leukocyte removal filter that has captured at least leukocytes, Residual blood in the leukocyte removal filter unit is collected, characterized by having means for detecting gas when it reaches the removal filter outlet and means for injecting a physiological solution into the connection site between the leukocyte removal filter outlet and the downstream circuit. The present invention relates to a residual blood collection device.
[0035]
The means for supplying gas to the leukocyte removal filter inlet capturing the leukocytes of the present invention is any means as long as the gas can be supplied to the leukocyte removal filter inlet after capturing leukocytes at a constant or variable flow rate. Means can also be used. For example, blood feeding means, a blower pump, compressed air, an air compressor and the like used for capturing leukocytes are useful. In particular, blood feeding means used for removing leukocytes can be used conveniently. Examples include blood pumps, roller pumps, finger pumps, syringe pumps, and liquid feeding by a drop. From the viewpoint of keeping the flow rate constant, a blood pump and a roller pump are usefully used. On the other hand, a head is usefully used from the viewpoint of simplicity.
[0036]
The air feeding means of the present invention is preferably used at a flow rate of 10 ml / min or more and less than 100 ml / min. When the flow rate is less than 10 ml / min, it takes time to supply air, which is not preferable. On the other hand, when the flow rate is 100 ml / min or more, pressure is applied during air supply, and the risk of hemolysis of red blood cells increases due to an increase in pressure in the leukocyte removal filter, which is not preferable. From the above viewpoint, a more preferable flow rate is 12 ml / min or more and less than 90 ml / min, and most preferably 15 ml / min or more and less than 80 ml / min.
[0037]
As the means for detecting the gas when the gas of the present invention reaches the leukocyte removal filter outlet, any detector can be used as long as it can detect that the gas has been discharged from the leukocyte removal filter outlet. Examples of preferred examples include a bubble detector and a pressure sensor.
[0038]
Detection of gas discharge with a pressure sensor or pressure gauge is a bubble point-like discharge when air is sent into the leukocyte removal filter and the inside of the filter is replaced with gas when the internal liquid is almost recovered. Pressure is generated, and gas discharge can be easily detected by detecting this pressure. When a non-woven leukocyte removal filter material is used, the differential pressure between the inlet and the outlet of the leukocyte removal filter increases rapidly, so that it can be used particularly useful. In this case, when the differential pressure is 100 mmHg or more, it can be determined that blood is completely discharged and immediately before gas is discharged.
[0039]
As the means for injecting a physiological solution after the leukocyte removal filter outlet of the present invention, any injection method can be used as long as it is a means capable of injecting the physiological solution into the downstream circuit of the leukocyte removal filter outlet. As a means for injecting, a physiological circuit with a constant or variable flow rate by means of a head, a blood pump, a roller pump, a finger pump, a syringe pump, a peristaltic pump, or the like from a branch circuit provided in the vicinity of the filter outlet in the downstream circuit of the filter. Means for injecting the solution are usefully used.
The outlet of the leukocyte removal filter of the present invention and the connection site of the downstream circuit may be provided between the outlet of the leukocyte removal filter and the downstream circuit connected thereto, or the leukocyte removal filter of the downstream circuit Even if it is provided at a position close to the outlet, it is preferably used.
[0040]
A drip chamber, a pump tube, a pressure measuring tube, a pressure measuring blood collecting device, a pump and the like can be appropriately added to the blood collecting device.
[0041]
Below, the blood collection system of this invention is demonstrated.
The circuit system of the present invention allows blood to flow through a leukocyte removal filter of a leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit liquid-tightly connected to the outlet side of the leukocyte removal filter. A circuit system for collecting blood from which leukocytes remaining after removing leukocytes after capturing leukocytes and collecting blood from which leukocytes have been removed, and supplying gas to the upstream side of the leukocyte removal filter that captured at least leukocytes A blood recovery system for recovering residual blood in a leukocyte removal filter that has captured leukocytes, comprising a circuit for injecting a physiological solution into a connection site between an outlet of the leukocyte removal filter and a downstream circuit.
The blood from which leukocytes remaining on the leukocyte removal filter that has captured leukocytes in the present invention are removed includes at least red blood cells and plasma.
[0042]
The circuit for supplying gas to the upstream side of the leukocyte removal filter that has captured leukocytes of the present invention is an airtight circuit that can supply gas and needs to have a gas inlet. The circuit for supplying the gas needs to be arranged upstream of the leukocyte removal filter. It is preferable that a filter that does not allow viruses, bacteria, and fine particles to pass therethrough is provided at the gas inlet, if necessary. Instead of providing a new circuit for supplying gas upstream of the leukocyte removal filter, the blood inlet side circuit is removed from the blood inlet and can be used instead.
[0043]
The circuit for injecting a physiological solution into the connection site between the outlet of the leukocyte removal filter and the downstream circuit is a liquid-tight circuit capable of feeding the physiological solution and needs to have an inlet for the physiological solution. . The circuit for feeding physiological solution is preferably arranged immediately after the leukocyte removal filter. If necessary, a spike needle, an injection needle, or the like may be provided at the physiological solution inlet. In this case, the physiological solution can be joined in a liquid-tight manner to a solution bag containing the physiological solution or the like. It is also possible to join the syringe pump with a luer lock or the like. Further, it is useful even if a container filled with a physiological solution is arranged in advance at the end of the circuit. It is also possible to place a needle puncture port immediately after the leukocyte removal filter and inject from this port. A drip chamber is preferably disposed between the physiological solution inlet and the blood outlet in order to prevent air from entering the blood.
[0044]
【Example】
Examples of the present invention are shown below, but the present invention is not limited thereto.
[Example 1]
Nonwoven fabric made of polyester fiber with an average fiber diameter of 2.7μm (weight per unit: 90g / m 2 , 0.42 mm thick) was cut into a 75 mm circle, sandwiched between upper and lower polyethylene filters with a pore size of 90 μm, and 10 sheets were packed into a column.
Human fresh blood with added nafamostat mesylate as anticoagulant (white blood cell count: 8.200 / μL, platelet count: 300.000 / μL) (nafamostat mesylate concentration: 0.012 mg / ml) 400 mL with a blood pump at 20 mL / min At a constant flow rate at room temperature to remove leukocytes.
[0045]
The blood inlet was opened, and air was allowed to flow to the column outlet at 20 ml / min with a blood pump. After confirming the outflow of air with the bubble detector, 30 ml of physiological saline was flowed from the circuit immediately after the leukocyte removal filter outlet with a finger pump at a flow rate of 20 ml / min, and the leukocyte-removed blood downstream of the leukocyte removal filter was collected. The histamine concentration and serotonin concentration of 50 ml of the combined solution of blood before treatment and 20 ml of gas recovery solution and 30 ml of physiological saline solution were measured with an EIA kit (manufactured by Immunotech). From the histamine and serotonin concentrations, the respective concentration increase rates were determined by the following equations (1) and (2). The results are shown in Table 1.
In addition, the number of leukocytes in the blood before and after the leukocyte removal filter treatment was measured with a multi-item automatic blood cell analyzer (manufactured by Sysmex), and the leukocyte removal rate was determined from the following equation (3). The results are shown in Table 1.
[0046]
Increase rate of histamine concentration = histamine concentration in recovered solution / histamine concentration in blood before treatment: Formula (1)
Serotonin concentration increase rate = recovered solution serotonin concentration / pre-treatment blood serotonin concentration formula (2)
Leukocyte removal rate = {1− (number of leukocytes after leukocyte removal filter treatment / number of leukocytes in blood before treatment)} × 100 (3)
[0047]
[Comparative Example 1]
Instead of using air from the leukocyte removal filter, 50 ml of blood from which leukocytes were removed using 50 ml of physiological saline was collected and used as a collected solution. The histamine and serotonin concentrations in the blood and the collected liquid before treatment, and the leukocyte removal rate were exactly the same as in Example 1, and histamine was used using the formulas (1), (2) and (3) described in Example 1, respectively. The concentration increase rate, serotonin concentration increase rate and leukocyte removal rate were determined. The results are shown in Table 1.
[0048]
[Table 1]
[Example 2]
Nonwoven fabric made of polyester fibers with an average fiber diameter of 2.7μm (90g / m 2 Was cut into a width of 150 mm and a length of 760 mm and wound around a polyethylene cylindrical mesh having a diameter of 3.4 mm. Next, a nonwoven fabric (30 g / m) made of polyester fibers having an average fiber diameter of 12 μm 2 The basis weight was the first prefilter, and it was wound with a width of 150 mm and a length of 1,300 mm. Furthermore, non-woven fabric (50 g / m) made of polyester fiber with an average fiber diameter of 33 μm 2 The basis weight was a second pre-filter, and it was wound with a width of 150 mm and a length of 1.300 mm. A polyethylene mesh was wound on the outside with a width of 150 mm and a length of 150 mm. The cylinder diameter was 39 mm. Both ends of this cylinder are closed with urethane, a cylindrical polycarbonate container with an inner diameter of 41 mm having a blood inlet and outlet at the ceiling and bottom, respectively, the outer peripheral surface of the cylinder is the blood inlet of the container, and the inner peripheral surface is the blood outlet The leukocyte removal filter was used.
[0050]
Bovine fresh blood with heparin added as an anticoagulant (white blood cell count: 5.300 / μL, platelet count: 170.000 / μL) (heparin concentration: 1000 IU / L) 3000 mL of heparin at a constant flow rate of 50 mL / min at room temperature Rinse and remove white blood cells. The blood inlet was opened and air was allowed to flow to the column outlet at 50 ml / min. After detecting the pressure increase due to air mixing with the pressure detector, 200 ml of physiological saline is flowed from the circuit immediately after the leukocyte removal filter outlet at a flow rate of 50 ml / min, and 200 ml of blood from which leukocytes remaining in the circuit after the leukocyte removal filter have been removed are removed. It was collected. The histamine concentration and serotonin concentration of 380 ml of the collected blood and 200 ml of the collected gas and 180 ml of the physiological saline solution were measured with an EIA kit (manufactured by Immunotech). Using formulas (1) and (2) described in Example 1, the histamine concentration increase rate and the serotonin concentration increase rate were determined, respectively. Further, in the same manner as in Example 1, the leukocyte count of the blood after the leukocyte removal filter treatment and the leukocyte count of the blood before the treatment were obtained at 1000 ml treatment, 2000 ml treatment, and 3000 ml treatment, and these values were averaged. Was the white blood cell count of the blood after the leukocyte removal filter treatment and the white blood cell count of the blood before the treatment, and the leukocyte removal rate was used using the formula (3) described in Example 1.
[0051]
[Comparative Example 2]
Using the same leukocyte removal filter as in Example 2, the blood from which leukocytes had been removed was collected using 380 ml of 5% glucose solution instead of using air before the leukocyte removal filter, and used as a collected solution. The histamine and serotonin concentrations and the white blood cell count were sampled in exactly the same manner as in Example 2. Using the formulas (1), (2), and (3) described in Example 1, the histamine concentration increase rate and serotonin concentration, respectively. The increase rate and leukocyte removal rate were determined. The results are shown in Table 2.
[0052]
[Table 2]
【The invention's effect】
According to the present invention, blood (red blood cells, plasma, etc.) from which leukocytes remaining in the leukocyte removal filter unit have been removed can be recovered while suppressing the release of cytokines and mediators.
Claims (7)
白血球除去フィルター内に残留する白血球が除去された血液を気体により白血球除去フィルターの出口を過ぎるまで押し出す工程及び白血球除去フィルターの出口と下流側回路の接続部位から無菌的に生理的溶液を流し込み下流側回路に残留する白血球が除去された血液を下流側回路外に押し出す工程とを有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液回収方法。The leukocyte removal filter unit of the leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit connected fluid-tightly to the outlet side of the leukocyte removal filter flows blood to capture leukocytes and then remains in the leukocyte removal filter In a method of collecting blood from which leukocytes to be removed are collected,
The process of extruding the blood from which leukocytes remaining in the leukocyte removal filter have been removed by gas until the outlet of the leukocyte removal filter is passed by the gas, and the physiological solution is poured aseptically from the connection site between the leukocyte removal filter outlet and the downstream circuit downstream possess a step of extruding a blood leukocytes remaining in the circuit is removed from the downstream circuit, when recovering the blood remaining in the leukocyte-removing filter, suppressing the increase in the concentration of captured the leukocyte-derived mediators A method for recovering residual blood in a leukocyte removal filter unit.
白血球除去フィルター内に残留する白血球が除去された血液を気体により白血球除去フィルターの出口を過ぎるまで押し出す工程及び白血球除去フィルターの出口と下流側回路の接続部位から無菌的に生理的溶液を流し込み下流側回路に残留する白血球が除去された血液を下流側回路外に押し出す工程とを有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球中の顆粒球由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液回収方法。The process of extruding the blood from which leukocytes remaining in the leukocyte removal filter have been removed by gas until the outlet of the leukocyte removal filter is passed by the gas, and the physiological solution is poured aseptically from the connection site between the leukocyte removal filter outlet and the downstream circuit downstream A step of extruding the blood from which the leukocytes remaining in the circuit have been removed to the outside of the downstream circuit, and when collecting the blood remaining in the leukocyte removal filter, the mediator derived from granulocytes in the captured leukocytes A method for recovering residual blood in a leukocyte removal filter unit, characterized by suppressing an increase in concentration.
白血球除去フィルター内に残留する白血球が除去された血液を気体により白血球除去フィルターの出口を過ぎるまで押し出す工程及び白血球除去フィルターの出口と下流側回路の接続部位から無菌的に生理的溶液を流し込み下流側回路に残留する白血球が除去された血液を下流側回路外に押し出す工程とを有し、前記白血球除去フィルターは、血小板も捕捉し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された血小板由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液回収方法。The process of extruding the blood from which leukocytes remaining in the leukocyte removal filter have been removed by gas until the outlet of the leukocyte removal filter is passed by the gas, and the physiological solution is poured aseptically from the connection site between the leukocyte removal filter outlet and the downstream circuit downstream And extruding the blood from which the white blood cells remaining in the circuit are removed to the outside of the downstream circuit, and the white blood cell removal filter captures platelets and is captured when the blood remaining in the white blood cell removal filter is collected. A method for collecting residual blood in a leukocyte removal filter unit, characterized by suppressing an increase in the concentration of a platelet-derived mediator.
少なくとも白血球を捕捉した白血球除去フィルター入口に気体を送気する手段、
気体が白血球除去フィルター出口に達したとき気体を検出する手段、
白血球除去フィルター出口と下流側回路の接続部位に生理的溶液を注入する手段を有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液を回収する残留血液回収装置。The leukocyte removal filter unit of the leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit connected fluid-tightly to the outlet side of the leukocyte removal filter flows blood to capture leukocytes and then remains in the leukocyte removal filter A device for collecting blood from which white blood cells have been removed,
Means for supplying gas to the inlet of the leukocyte removal filter that captures at least leukocytes,
Means for detecting the gas when the gas reaches the leukocyte removal filter outlet;
Have a means for injecting a physiological solution in connection site of leukocyte-removing filter outlet and the downstream circuit, when recovering the blood remaining in the leukocyte-removing filter, suppressing the increase in the concentration of captured the leukocyte-derived mediators A residual blood recovery device for recovering residual blood in a leukocyte removal filter unit.
少なくとも白血球を捕捉した白血球除去フィルターの上流側に気体を送気する回路、
白血球除去フィルターの出口と下流側回路の接続部位に生理的溶液を注入する回路を有し、前記白血球除去フィルターに残留する血液を回収する際に、捕捉された前記白血球由来のメディエーターの濃度上昇を抑えることを特徴とする白血球除去フィルターユニット内の残留血液を回収する血液回収システム。The leukocyte removal filter unit of the leukocyte removal filter unit comprising at least a leukocyte removal filter and a downstream circuit connected fluid-tightly to the outlet side of the leukocyte removal filter flows blood to capture leukocytes and then remains in the leukocyte removal filter A circuit system that collects blood from which white blood cells have been removed,
A circuit that sends gas to the upstream side of the leukocyte removal filter that captures at least leukocytes,
Have a circuit for injecting a physiological solution in connection site of the outlet and downstream circuit leukocyte removal filter, when recovering the blood remaining in the leukocyte-removing filter, the increase in concentration of captured the leukocyte-derived mediators A blood collection system for collecting residual blood in a leukocyte removal filter unit characterized by suppressing the blood.
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