JP5086101B2 - 細胞分類方法及び装置 - Google Patents
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Description
なお、本出願に対応する外国の特許出願においては下記の文献が発見または提出されている。
Claims (40)
- 対向し合う一対の壁面と対向し合う一対の端面とを備えるフローチャンバーであって、前記フローチャンバーの上面に配置された導入口と、フローチャネルを形成する中間検出領域と、選択的な処理領域と、前記フローチャンバーの底面に配置された下流側導出口とを有するフローチャンバーを備え、
前記導入口は、導入シース液体による隔離した第1のシース流れ及び第2のシース流れを受け入れる複数のシース流れ導入口と、分類すべき複数の細胞を含有する試料流体による、前記第1のシース流れと前記第2のシース流れの間の第3の流れを受け取る試料導入口を有し、
前記試料導入口は前記複数のシース流れ導入口の間に配置され、
前記導入シース液体の流量及び圧力のうちの少なくとも1つは、導入された前記試料流体の幅が、前記対向し合う一対の壁面に対して垂直な方向及び前記一対の端面に対して垂直な方向で、直接に収縮されるよう選択され、その結果、導入された前記試料流体が少なくとも前記中間検出領域において、前記導入シース液体により収縮された試料流れとなり、
前記収縮は、前記対向し合う一対の壁面に対して垂直な方向にせん断を発生させ、発生したせん断流により前記複数の細胞が、その扁平面が前記一対の壁面に平行になるように向きが定まるべくアライメントされ、
前記中間検出領域に配置され、アライメントされた前記複数の細胞から複数の所望の細胞を検出してそれを示す出力を生成する検出器と、
前記選択的な処理領域に設置され、前記試料流れから所望の細胞を分類するために、前記細胞の流量を変化させる分類器と、
少なくとも1つが、前記所望の細胞を受け取る、隔離して配置された複数の領域又はチャネルと、
を備えた、複数の細胞を分類及び配向するフロー装置。 - 前記分類器は、前記検出器の下流に存在し、前記試料流れから前記複数の所望の細胞を分類する、請求項1に記載のフロー装置。
- 前記分類器は、前記検出器の下流において、少なくとも一方が前記複数の所望の細胞を受け取る第1、第2の複数の隔離した領域が形成された、前記フローチャンバーの1つの部分を含む、請求項2に記載のフロー装置。
- 前記分類器は、前記複数の所望の細胞を第1領域へと案内し、複数の所望でない細胞を第2領域へと案内する目的で、前記検出器に応答して、1つの領域又はそれより多くの複数の領域へと選択的に圧力を印加する圧力変換器を含む、請求項3に記載のフロー装置。
- 前記分類器は、複数の所望の細胞を複数の所望でない細胞から分離する、請求項1から4のいずれか1項に記載のフロー装置。
- 前記一対の壁面は、一対の平面部材である、請求項1から5のいずれか1項に記載のフロー装置。
- 前記複数の細胞は前記試料流体内に懸濁されている、請求項1から6のいずれか1項に記載のフロー装置。
- 前記第1のシース流れ及び第2のシース流れの、前記第3の流れに相対的な前記流量又は圧力は、少なくとも前記中間検出領域において、前記一対の壁面に対して垂直な方向の前記第3の流れの幅が、前記試料導入口での前記一対の壁面に対して垂直な方向の前記第3の流れの幅の10パーセント以下となるように前記第3の流れを収縮させ、それにより前記中間検出領域に進入する複数の細胞を互いに対して、また前記一対の壁面に対して略平行な1つの方向に配向させる、請求項7に記載のフロー装置。
- 前記中間検出領域は、入力光信号を集中させる領域を含む、請求項8に記載のフロー装置。
- 前記下流側導出口は、第1と第2の複数の隔離した領域により形成され、第1の領域は所望の細胞を受け取り、第2の領域は、所望でない種の複数の細胞を受け取る、請求項1に記載のフロー装置。
- 前記検出器は、前記中間検出領域において、前記複数の細胞に入力光を光学的に照射する光源を備える、請求項1から10のいずれか1項に記載のフロー装置。
- 前記複数の細胞は前記入力光に反応して出力光を生成する反応性を有しており、前記検出器は前記複数の細胞が生成する光に反応する光検出器を備える、請求項11に記載のフロー装置。
- 前記出力光は、前記複数の細胞から散乱する光と、前記複数の細胞が生成する蛍光光の少なくとも一方を含む、請求項12に記載のフロー装置。
- 前記検出器は、前記複数の細胞からの出力光を受光するための、また、当該出力光を集光してそれによる複数の集光光ビームレットを生成する第1光学素子を含む、光学系配列を備え、前記複数の集光光ビームレットは、前記細胞の前記入力光との相互作用に従って生成される、前記細胞の多様な角度における複数の画像を示す、請求項12に記載のフロー装置。
- 前記検出器は、前記光学系配列の中央光軸に沿っており、前記中央光軸に沿った複数の選択された軸上ビームレットを複数の対応する光検出器へと方向付ける光検出器を更に備える、請求項14に記載のフロー装置。
- 前記検出器は、前記中央光軸からずれて方向付けられる複数の光路を有する複数の軸ずれビームレットを検出するための、1つの対応する軸ずれ光路を有する少なくとも1つの軸ずれ光検出器を更に備える、請求項15に記載のフロー装置。
- 前記少なくとも1つの軸ずれ光検出器の前記光路内の光スプリッタであって、前記複数の集光光ビームレットから選択される複数のビームレットを前記対応する軸ずれ検出器へと方向づける光スプリッタを更に備える、請求項16に記載のフロー装置。
- 前記複数の集光光ビームレットは、散乱光と蛍光光を含み、前記検出器は、前記複数の集光光ビームレットから前記散乱光と前記蛍光光の少なくとも一方をフィルタリングする複数の光フィルターを更に備える、請求項15に記載のフロー装置。
- 前記分類器は、細胞種ごとに1つのチャネルと、前記フローチャネルに接続され、前記試料流れの複数の選択位置において前記流量を変更して前記複数の細胞を複数のチャネルうちの選択した複数のチャネルへと案内するための駆動部材とを備える、請求項2に記載のフロー装置。
- 前記駆動部材は、前記フローチャネルに圧力を印加し、前記圧力を変化させるための圧力変換器を備える、請求項19に記載のフロー装置。
- 前記駆動部材は、圧電変換器、加熱器、容量膜、及び、光加熱器の少なくとも1つを備える、請求項19に記載のフロー装置。
- 細胞寿命を延長するためには、前記フロー装置を、摂氏2度から10度にて運転することができる、請求項1から21のいずれか1項に記載のフロー装置。
- 前記所望の細胞は、X染色体細胞及びY染色体細胞のうちの一方である請求項5に記載のフロー装置。
- 前記所望でない細胞は、前記X染色体細胞及び前記Y染色体細胞のうちの他方である請求項23に記載のフロー装置。
- 前記所望でない細胞は、レーザーの使用により死滅させられる請求項24に記載のフロー装置。
- 前記細胞は***細胞であり、前記分類器は、前記***細胞の性別識別を実行する請求項5に記載のフロー装置。
- 前記細胞に対して実行される分類及び配向は、選択的な特徴識別のために実行される請求項5に記載のフロー装置。
- 前記試料流体及び前記シース流体はマルチチャネル化されており、マルチチャネル構成に配置されている請求項5に記載のフロー装置。
- 前記マルチチャネル構成は、
導入された複数の並列な試料流体及びシース液体と、
前記導入された複数の並列な試料流体及びシース液体のそれぞれに対する、隔離して配置された複数の領域又は導出チャネルとを備える請求項28に記載のフロー装置。 - 前記導入された複数の並列な試料流体及びシース液体のそれぞれに対して提供されるレンズシステムであって、複数の検出器を備え、前記複数の検出器のそれぞれは、前記導入された流れとシース液体流のそれぞれに対して専用であるレンズシステムを更に備える請求項29に記載のフロー装置。
- レーザービームを生成するレーザーデバイスと、
前記レーザービームを複数のビームレットに分割するビームスプリッタであって、前記複数のビームレットは、前記導入された複数の並列な試料流体及びシース流体のうちの1つに対応する前記中間検出領域に対して方向付けられるビームスプリッタと、
前記複数のビームレットを複数の平行なビームにして出力する平行化レンズと、
前記平行化レンズから出力された前記複数の平行なビームを方向付けする円筒状レンズであって、対物面において、前記複数の平行なビームのそれぞれを導入された試料流体に集中させる円筒状レンズとを更に備える請求項30に記載のフロー装置。 - 集光レンズと
前記複数のビームレットのそれぞれから光を検出する検出器アレイデバイスとを更に備え、
前記複数のビームレットの1つ又は全ての光と前記導入された流れは前記集光レンズへと進入して、散乱光と蛍光光の両方が集光される請求項31に記載のフロー装置。 - 前記ビームスプリッタは、回折格子又はホログラムである請求項31に記載のフロー装置。
- 前記検出器アレイデバイスは、光電子増倍管のアレイである請求項32に記載のフロー装置。
- 前記選択的な処理領域内において、前記所望でない細胞を死滅させることによって、前記所望でない細胞から前記所望の細胞が分割される請求項5に記載のフロー装置。
- 前記所望でない細胞を死滅させることは、レーザー死滅、電気パルス活性化電気穿孔、又は細胞活性化の少なくとも1つを通して実行される請求項35に記載のフロー装置。
- 前記検出器の下流に配置された、所望でない細胞を死滅させる殺細胞装置を更に備える請求項1に記載のフロー装置。
- 前記殺細胞装置は、前記所望でない細胞に、致死的エネルギーを衝突させる電極及びレーザーの少なくとも1つを備える、請求項37に記載のフロー装置。
- 前記殺細胞装置は、前記細胞内の、前記細胞を死滅させる対象を活性化するための光源を備える、請求項37に記載のフロー装置。
- 前記検出器の下流に配置された、所望でない細胞を処理又は死滅させる、細胞電気穿孔を更に備える請求項1に記載のフロー装置。
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Also Published As
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BRPI0607095B1 (pt) | 2017-11-21 |
ES2595047T3 (es) | 2016-12-27 |
CN102212457B (zh) | 2014-09-10 |
HK1203626A1 (en) | 2015-10-30 |
BRPI0607095A2 (pt) | 2009-08-04 |
ES2639306T3 (es) | 2017-10-26 |
EP3109616A1 (en) | 2016-12-28 |
HK1112288A1 (en) | 2008-08-29 |
EP1846748A2 (en) | 2007-10-24 |
EP1846748B1 (en) | 2016-09-14 |
CN102212457A (zh) | 2011-10-12 |
DK2166340T3 (en) | 2017-09-18 |
CN101189504A (zh) | 2008-05-28 |
JP2012013714A (ja) | 2012-01-19 |
EP1846748A4 (en) | 2009-05-20 |
HK1160222A1 (en) | 2012-08-10 |
CN102128778A (zh) | 2011-07-20 |
EP2166340A1 (en) | 2010-03-24 |
EP2166340B1 (en) | 2017-08-02 |
US7355696B2 (en) | 2008-04-08 |
WO2006083907A3 (en) | 2006-12-21 |
USRE46992E1 (en) | 2018-08-14 |
EP2801813A1 (en) | 2014-11-12 |
US20080144037A1 (en) | 2008-06-19 |
EP2410314A2 (en) | 2012-01-25 |
US20060170912A1 (en) | 2006-08-03 |
US7545491B2 (en) | 2009-06-09 |
EP2410314A3 (en) | 2012-07-11 |
JP2008530985A (ja) | 2008-08-14 |
WO2006083907A2 (en) | 2006-08-10 |
CN102128778B (zh) | 2013-12-18 |
BR122017014567B1 (pt) | 2018-04-17 |
CN101189504B (zh) | 2012-05-30 |
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