JP2019511913A - 生体試料のための自動化された分析ツール - Google Patents
生体試料のための自動化された分析ツール Download PDFInfo
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- JP2019511913A JP2019511913A JP2018544262A JP2018544262A JP2019511913A JP 2019511913 A JP2019511913 A JP 2019511913A JP 2018544262 A JP2018544262 A JP 2018544262A JP 2018544262 A JP2018544262 A JP 2018544262A JP 2019511913 A JP2019511913 A JP 2019511913A
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Abstract
Description
蛍光染料から得られる蛍光シグナルのための励起ユニットおよび検出ユニットを有する蛍光システムと、
少なくとも1つの生体標本を蛍光染料と共に含む少なくとも1つのコンテナを保持するための開口部と、
蛍光シグナルを消光する消光ユニットと、
液体をコンテナに供給する、かつ/または液体をコンテナから取り出す、液体処理システムと、
開口部、蛍光システム、消光ユニットおよび液体処理システムのうちの少なくとも1つを、1つの作業平面を規定する少なくとも2つの直交する次元で移動させるメカニズムと、
さらに蛍光染料の励起、蛍光シグナルの検出および収集ならびに蛍光シグナルの消光を含む1つのルーチンを、自動化された手法で実行する制御ユニットと、
を含む。
少なくとも1つの生体標本を蛍光染料と共に、ステージ上の開口部におけるコンテナ内に保持すること、
蛍光システムによって、蛍光染料を励起し、蛍光染料から得られる蛍光シグナルを検出すること、
開口部、ステージ、蛍光システムおよび消光ユニットのうちの少なくとも1つを、1つの作業平面を規定する少なくとも2つの直交する次元において、生体標本が消光ユニットと隣り合うまで移動させること、
さらに消光ユニットによって蛍光シグナルを消光すること、
を含む。
本発明において使用される蛍光試薬は、蛍光部分と抗原認識部分とを含んでおり、オプションとしてこれらはスペーサ分子により結合される。
用語「抗原認識部分」とは、細胞標本の細胞において発現したマーカに対して向けられた、任意の種類の抗体、または断片化抗体、または断片化抗体派生物のことを指す。この用語は、完全に無損傷の抗体、断片化抗体または断片化抗体派生物に関連するものであり、たとえばFab,
生体標本は、蛍光試薬の抗原認識部分により検出/認識可能な標的部分を有する。生体標本は、動物全体、無脊椎動物(たとえばシノラブディス・エレガンス、キイロショウジョウバエ)、脊椎動物(たとえばゼブラフィッシュ、アフリカツメガエル)、および哺乳動物(たとえばハツカネズミ、人類)の器官、組織切片、細胞集合体、または単一細胞のような任意の試料に由来するものとすることができる。生体標本は、組織切片、細胞集合体、懸濁細胞または粘着細胞の形態を有することができる。細胞を、生体または死体とすることができる。これらの試薬各々を別個の液体容器に収容することができ、この場合、液体容器各々が試薬、検出部分を有する抗原認識部分、蛍光染料を含む抗体、抗生物質、生体栄養素、毒素、染料および酸化剤のうちの少なくとも1つを含むようにする。
R−フィコエリトリン(R−PE)の励起のためには520〜560nmの緑色光というように、蛍光試薬に対して調節された適切な波長の光の照射を介して、蛍光励起が実施される。したがって励起ユニットによって、特定の蛍光を吸収するスペクトル範囲内の電磁放射が供給される。次いで蛍光体は、赤色方向にシフトした波長(シフトは一般に約20nm)の蛍光を放出し、励起放射からこれを別個に検出することができる。蛍光顕微鏡励起ユニットの一般的な具現化形態は、アークランプ、キセノンランプまたはメタルハライドランプなどのような白色光源と、励起に必要とされるスペクトル帯域を生成するための後続のフィルタとを含んでいる。レーザまたは発光ダイオードも使用される。
用語「消光」および「退色」は、本明細書では入れ替え可能に用いられ、蛍光体または標識された生体標本への蛍光体の付着の化学的または放射による変質の結果として、標識された生体標本からの蛍光の減少を意味する、と理解されたい。したがって消光システムはさらに、試薬の蛍光性能または生体標本への試薬の付着を変質させる化学薬品または放射のうち少なくとも一方のソースを有することができる。
本発明による自動化されたシステムのコンテナは、各々が別個の生体標本を含む複数の液体ウェルを有することができる。好ましくは、このコンテナはタイタプレートである。
本発明によるシステムは、蛍光シグナルを測定する蛍光システムと、蛍光システムの上方で少なくとも1つの生体標本を含むディスポーザブル品を保持する開口部と、生体標本からの蛍光シグナルを消光するためにクエンチング光を供給する消光システムと、コンテナに液体を供給可能および/またはコンテナから液体を除去可能な液体処理システムと、開口部、蛍光システム、消光ユニットおよび液体処理システムのうちの少なくとも1つを、1つの作業平面を規定する少なくとも2つの直交する次元で移動させるメカニズムと、さらに蛍光染料の励起、蛍光シグナルの検出および収集ならびに蛍光シグナルの消光を含む1つのルーチンを、自動化された手法で実行する制御ユニットと、を含むことができる。
これまで、例示的な蛍光イメージングシステム30および例示的な蛍光消光システム40の構成要素について説明してきたが、次にこの装置を用いる方法について説明する。なお、この方法によって、既述の実施形態における構成要素のあらゆるすべてのものを使用できる、ということを理解されたい。以下では、この方法全般について概略的に説明し、続いて図10に示した固有のアルゴリズムについて述べる。
Claims (21)
- 少なくとも1つの蛍光染料により染色された生体標本を分析するための自動化されたシステムであって、前記システムは、
前記蛍光染料から得られる蛍光シグナルのための励起ユニットおよび検出ユニットを有する蛍光システムと、
少なくとも1つの生体標本を前記蛍光染料と共に含む少なくとも1つのコンテナを保持するための開口部と、
前記蛍光シグナルを消光する消光ユニットと、
液体を前記コンテナに供給する、かつ/または、液体を前記コンテナから取り出す、液体処理システムと、
前記開口部、前記蛍光システム、前記消光ユニットおよび前記液体処理システムのうちの少なくとも1つを、1つの作業平面を規定する少なくとも2つの直交する次元において移動させるメカニズムと、
さらに前記蛍光染料の励起、前記蛍光シグナルの検出および収集ならびに前記蛍光シグナルの消光を含む1つのルーチンを、自動化された手法で実行する制御ユニットと、
を含む自動化されたシステム。 - 前記液体処理システムは、蛍光染料、前記蛍光シグナルを消光する配合物、洗浄液および緩衝剤のうちの少なくとも1つを、前記生体標本へ供給する、
請求項1記載の自動化されたシステム。 - 前記液体処理システムは、ステージ上に配置されたロボット制御型のピペットシステムを有し、前記ピペットシステムは、前記作業平面に対し直交するz軸に沿って移動可能である、
請求項1または2記載の自動化されたシステム。 - 前記液体処理システムは、前記ピペットシステムの外側を洗浄して付着した液体を除去する手段を有する、
請求項1から3までのいずれか1項記載の自動化されたシステム。 - 前記制御ユニットは、蛍光染料で染色された前記生体標本のイメージとして、前記蛍光シグナルを収集する、
請求項1から3までのいずれか1項記載の自動化されたシステム。 - 前記自動化されたシステムはさらに、少なくとも1つの付加的な液体容器を有し、前記付加的な液体容器は、試薬、検出部分を有する抗原認識部分、蛍光染料を含む抗体、抗生物質、生体栄養素、毒素、染料および酸化剤のうちの少なくとも1つを含む、
請求項1から4までのいずれか1項記載の自動化されたシステム。 - 前記コンテナは、各々別個の生体標本を含む複数の液体ウェルを有する、
請求項1から5までのいずれか1項記載の自動化されたシステム。 - 前記コンテナは、タイタプレートである、
請求項6記載の自動化されたシステム。 - 前記コンテナは、透明部分と不透明部分とを有し、前記透明部分に前記生体標本が配置され、前記不透明部分は、前記蛍光シグナルを反射するかまたは吸収する、
請求項1から7までのいずれか1項記載の自動化されたシステム。 - 前記消光ユニットは、前記蛍光システムに横方向で隣り合って配置された光源を有し、前記光源は、消光放射を放出する、
請求項1から9までのいずれか1項記載の自動化されたシステム。 - 前記消光ユニットは、それぞれ異なる波長の消光放射を放出する2つ以上の光源を有する、
請求項1から9までのいずれか1項記載の自動化されたシステム。 - 前記消光ユニットはさらに、少なくとも1つのミラーを有し、前記ミラーは、前記生体標本または前記蛍光染料によっても吸収されない消光放射の一部分を反射して、前記生体標本に戻す、
請求項9記載の自動化されたシステム。 - 前記消光ユニットは、前記生体標本の表面領域あたりの放射および/または放射強度を均質化する手段を有する、
請求項1から9までのいずれか1項記載の自動化されたシステム。 - 前記消光ユニットは、400nmよりも短くかつ900nmよりも長い波長の放射を前記消光放射から実質的に取り除くフィルタを有する、
請求項1から9までのいずれか1項記載の自動化されたシステム。 - 前記消光ユニットは、さらに蛍光検出器を含み、前記蛍光検出器は、前記蛍光シグナルの減衰を消光データとして監視する、
請求項9記載の自動化されたシステム。 - 前記制御ユニットは、前記消光データに基づきフィードバックループにより前記消光ユニットを制御する、
請求項1から12までのいずれか1項記載の自動化されたシステム。 - 前記制御ユニットは、x方向またはy方向において±約1〜200μmの精度を伴って、前記開口部を前記作業平面内で移動させる、
請求項1から13までのいずれか1項記載の自動化されたシステム。 - 前記自動化されたシステムは、さらに温度制御ユニットを有し、前記温度制御ユニットは、前記生体標本の温度を10〜40℃に制御する、
請求項1から13までのいずれか1項記載の自動化されたシステム。 - 生体標本を分析するための自動化された方法であって、前記方法は、
少なくとも1つの生体標本を蛍光染料と共にステージ上の開口部におけるコンテナ内に保持するステップと、
蛍光システムによって、前記蛍光染料を励起し、前記蛍光染料から得られる蛍光シグナルを検出するステップと、
前記開口部、前記ステージ、前記蛍光システムおよび消光ユニットのうちの少なくとも1つを、1つの作業平面を規定する少なくとも2つの直交する次元において、前記生体標本が前記消光ユニットと隣り合うまで移動させるステップと、
さらに前記消光ユニットによって前記蛍光シグナルを消光するステップと、
を含む自動化された方法。 - 前記自動化された方法はさらに、
前記消光後に前記生体標本をイメージングするステップを含む、
請求項19記載の自動化された方法。 - 前記自動化された方法はさらに、
液体処理システムにより一連の液体を、前記生体標本を保持する前記コンテナに搬送するステップと、
さらに前記蛍光染料の励起、前記蛍光シグナルの検出および収集ならびに前記蛍光シグナルの消光を含む1つのルーチンを、一連の液体を用いて自動化された手法で実行するステップと、
を含む、
請求項19または20記載の自動化された方法。
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EP3420339A1 (en) | 2019-01-02 |
US20170241911A1 (en) | 2017-08-24 |
ES2955671T3 (es) | 2023-12-05 |
JP6914265B2 (ja) | 2021-08-04 |
WO2017144338A1 (en) | 2017-08-31 |
CN108700518A (zh) | 2018-10-23 |
CN108700518B (zh) | 2022-09-02 |
EP3420339B1 (en) | 2023-06-21 |
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