JP7454734B2 - サンプルの収集、安定化、および保存のためのシステム、方法およびデバイス - Google Patents
サンプルの収集、安定化、および保存のためのシステム、方法およびデバイス Download PDFInfo
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Description
[0001] 本出願は、2015年9月9日出願の米国仮特許出願第62/216,312号の優先権を主張するものとし、2015年11月25日出願の米国仮特許出願第62/260,172号の優先権を主張するものとし、2016年7月26日出願の米国仮特許出願第62/367,056号の優先権を主張するものとし、2016年7月29日出願の米国仮特許出願第62/368,817号の優先権を主張するものとし、その全体の開示は参照により本明細書に組み込まれる。
参照による援用
ョン可能なDNA配列を生成するステップと、1本鎖DNA環を生成するために、リガーゼを用いて、修復されたライゲーション可能なDNA配列の分子内ライゲーションを実施するステップと、を含むことができる。これらのステップは、介在する単離ステップまたは精製ステップを一切行わずに、単一の反応容器内で行うことができる。リン酸供与体は、グアノシン三リン酸(GTP)、シチジン三リン酸(CTP)、ウリジン三リン酸(UTP)、デオキシチミジン三リン酸(dTTP)またはそれらの組み合わせであり得る。線状DNAは、二本鎖または一本鎖DNAのいずれかであり得る。DNAは、循環DNAなどの断片化されたDNAの区画であり得る。ライゲーション可能なDNAは、二本鎖形態であれば、分子内ライゲーション反応の前に変性させる必要があり得る。ライゲーション反応には、鋳型に依存しない、一本鎖DNA配列の分子内ライゲーションが可能なプレアデニル化リガーゼを用いることができる。他の実施形態では、線状DNAから一本鎖DNA環を生成するための方法は、分子内ライゲーションステップの前にDNAプレアデニル化ステップを用いることができる。線状DNAを、任意により、アデノシン三リン酸(ATP)の存在下でポリヌクレオチドキナーゼと一緒にインキュベートして、5’末端に
リン酸基を、3’末端にヒドロキシル基を含むライゲーション可能なDNA配列を生成す
ることができる。線状DNAが高度に断片化された形態である場合、線状DNAからライゲーション可能なDNA配列を生成することが好ましい。次いで、線状DNAまたはライゲーション可能なDNA配列を、ATPの存在下でアデニル化酵素とインキュベートして、5’アデニル化DNA配列を生成することができる。5’-アデニル化DNA配列は、
非アデニル化リガーゼとインキュベートすることができ、これは、鋳型に依存しない5’
-アデニル化DNA配列の分子内ライゲーションが可能であり、一本鎖DNA環を生成することができる。この方法のすべてのステップは、介在する単離ステップまたは精製ステップを一切行わずに、単一の反応容器内で行うことができる。非アデニル化リガーゼがATP依存性リガーゼである場合、分子内ライゲーション反応の前に、ATPは、(例えば、反応混合物をホスファターゼで処置することによって)反応混合物から取り出され得る。5’-アデニル化DNAが二本鎖形態である場合、分子内ライゲーション反応の前に変
性させる必要があり得る。DNA環状化のこれらの方法および他の方法、ならびに増幅方法は、例えば、米国特許出願公開第2015/0031086号に見出され得る。
列を含む二本鎖DNAオリゴヌクレオチドにライゲートされるステップを含むことができる。この二本鎖DNAオリゴヌクレオチドは、二本鎖領域内のRNAポリメラーゼのためのプロモーターを含むことができ、その後に3′オーバーハングを形成する一本鎖DNAのセグメントが続く。3′オーバーハングがチミジン残基のストリングを含む場合、二本鎖DNAの一本鎖部分はメッセンジャーRNA(mRNA)ポリ(A)尾部の3′末端にハイブリダイズすることができる。リガーゼの添加後、mRNAは3’末端にライゲートされた二本鎖DNA配列のうちの1つの鎖を有し得る。RNAポリメラーゼを添加すると、RNA-DNAハイブリッド分子が効率よく転写され、cRNAを合成することができる。RNAポリメラーゼを用いた転写反応は各鋳型を複数回転写することができるため、この方法では効果的なRNA増幅が可能になる。上記と同様の別の方法は、上記のようにDNAオリゴヌクレオチドをRNAにライゲートすることを含み得る。しかし、DNAオリゴヌクレオチドは、固体支持体に付着すること、または親和性タグを含有することのいずれか一方が可能である。これは、RNA分子の非常に効率的な共有結合および/または捕捉を可能にし、様々なあらゆる目的に使用され得る。さらなる方法では、ライゲーションおよびその後の転写を利用して、cRNAの5’末端にユーザ定義の配列を含む相補的
RNAを作製することができる。この配列「タグ」は、RNAポリメラーゼプロモーターとライゲートされたRNA分子の3’末端との間に配置することができる。ユーザ定義の
配列は、cRNAの精製もしくは同定または他の配列特異的操作に使用することができる。続いて、記載された方法に従ってcRNA産物が実質的にライゲートされ、再増幅された場合、その結果得られた二本鎖増幅産物は、元のセンス鋳型に関して、センスであり得て、この新規産物は、5’末端に位置する2つの別個のユーザ定義の配列を有し得る。こ
れらの配列は、cDNAの合成に使用することができ、全長合成および方向性クローニングを可能にする。当業者であれば、ユーザ定義配列の有無にかかわらず、この二重増幅法がRNA量の有意な増加をもたらすことができ、これまで小さすぎて考慮に入れることができなかったサンプルの分析が可能になることを理解するであろう。DNA断片の増幅のためのこれらの方法および他の方法は、例えば、米国特許出願公開第2008/0003602号に見出され得る。
核酸の溶出
パンスルホン酸(MOPS)、2-(N-モルホリノ)エタンスルホン酸(MES)、2-[[1,3-ジヒドロキシ-2-(ヒドロキシメチル)プロパン-2-イル]アミノ]エタンスルホン酸(TES)、カコジル酸塩、グリシン、炭酸塩、またはそれらの任意の組み合わせであり得る。1種以上の緩衝剤は、約0.1mM~約500mM、約0.1mM~約400mM、約0.1mM~約300mM、約0.1mM~約200mM、約0.1mM~約100mM、約0.1mM~約50mM、約0.1mM~約25mM、約0.1mM~約20mM、約0.1mM~約15mM、約0.1mM~約10mM、約0.1mM~約5mM、約0.1mM~約4mM、約0.1mM~約3mM、約0.1mM~約2mM、約0.1mM~約1mM、約0.1mM~約0.9mM、約0.1mM~約0.8mM、約0.1mM~約0.7mM、約0.1mM~約0.6mM、約0.1mM~約0.5mM、約0.1mM~約0.4mM、約0.1mM~約0.3mM、または約0.1mM~約0.2mMの濃度で存在してもよい。緩衝剤は、500mM未満、400mM未満、300mM未満、200mM未満、100mM未満、50mM未満、25mM未満、20mM未満、15mM未満、10mM未満、5mM未満、4mM未満、3mM未満、2mM未満、1mM未満、0.9mM未満、0.8mM未満、0.7mM未満、0.6mM未満、0.5mM未満、0.4mM未満、0.3mM未満、0.2mM未満、または0.1mM未満の濃度で存在してもよい。緩衝剤は、500mM超、400mM超、300mM超、200mM超、100mM超、50mM超、25mM超、20mM超、15mM超、10mM超、5mM超、4mM超、3mM超、2mM超、1mM超、0.9mM超、0.8mM超、0.7mM超、0.6mM超、0.5mM超、0.4mM超、0.3mM超、0.2mM超、または0.1mM超の濃度で存在し得る。
ル、ジメルカプトプロパノール、エチレンジアミン四酢酸(EDTA)、エチレンジオキシ-ジエチレン-ジニトリロ-四酢酸、エチレングリコール-ビス-(2-アミノエチル)-N,N,N’,N’-テトラ酢酸(EGTA)、金属ニトリロトリ酢酸(NTA)、
サリチル酸、もしくはトリエタノールアミン(TEA)であり得る。緩衝液中の1種以上のキレート剤の濃度は、約0.1mM、1mM、5mM、10mM、20mMまたは25mMであり得る。緩衝液中の1種以上のキレート剤の濃度は、0.1mM、1mM、5mM、10mM、20mMまたは25mM未満であり得る。緩衝液中の1種以上のキレート剤の濃度は、0.1mM、1mM、5mM、10mM、20mMまたは25mM超であり得る。
タンパク質の溶出
リノ)プロパンスルホン酸(MOPS)、2-(N-モルホリノ)エタンスルホン酸(MES)、2-[[1,3-ジヒドロキシ-2-(ヒドロキシメチル)プロパン-2-イル]アミノ]エタンスルホン酸(TES)、カコジル酸塩、グリシン、炭酸塩、またはそれらの任意の組み合わせであり得る。緩衝剤は、0.1mM~約500、0.1mM~約400mM、0.1mM~約300mM、0.1mM~約200mM、0.1mM~約100mM、0.1mM~約50mM、0.1mM~約25mM、0.1mM~約20mM、0.1mM~約15mM、0.1mM~約10mM、0.1mM~約5mM、0.1mM~約4mM、0.1mM~約3mM、0.1mM~約2mM、0.1mM~約1mM、0.1mM~約0.9mM、0.1mM~約0.8mM、0.1mM~約0.7mM、0.1mM~約0.6mM、0.1mM~約0.5mM、0.1mM~約0.4mM、0.1mM~約0.3mM、または0.1mM~約0.2mMの濃度で存在し得る。緩衝液は、500mM未満、400mM未満、300mM未満、200mM未満、100mM未満、50mM未満、25mM未満、20mM未満、15mM未満、10mM未満、5mM未満、4mM未満、3mM未満、2mM未満、1mM未満、0.9mM未満、0.8mM未満、0.7mM未満、0.6mM未満、0.5mM未満、0.4mM未満、0.3mM未満、0.2mM未満、または0.1mMの濃度で存在し得る。緩衝剤は、約0.1mM、1mM、10mM、25mMまたは50mMで存在し得る。緩衝剤は、少なくとも0.1mM、1mM、10mM、25mM、または50mMで存在し得る。
Claims (15)
- 一体型のサンプル獲得およびサンプル安定化デバイスであって、
(i)1つ以上の穿刺要素を含むサンプル獲得構成要素と、
(ii)2つ以上の膜を含むサンプル安定化ユニットであって、前記2つ以上の膜が積み重ねられており、前記2つ以上の膜が互いに接触せず、前記2つ以上の膜の少なくとも1つが、サンプル分離段階およびサンプル収集段階を含む、サンプル安定化ユニットと、
(iii)チャネルであって、前記チャネルが、前記サンプル獲得構成要素の開口部から前記チャネルを通じて前記2つ以上の膜の上に生物学的サンプルを移動させるように構成される、チャネルと、
を有し、
前記サンプル安定化ユニットの前記2つ以上の膜が、前記生物学的サンプル内の1つ以上の生物学的構成要素を分離および安定化するように構成されている、
デバイス。 - 前記デバイスがアクチュエータを含み、前記アクチュエータが、前記サンプル獲得構成要素を作動させ、前記1つ以上の穿刺要素に対象の皮膚を穿刺させて前記生物学的サンプルを放出させるように構成されている、請求項1に記載のデバイス。
- 前記デバイスが、前記対象の前記皮膚から前記1つ以上の穿刺要素を引き抜くように構成されている、請求項2に記載のデバイス。
- 前記2つ以上の膜が、セルロース、酢酸セルロース、ガラス繊維、またはそれらの組み合わせを含む多孔質マトリックスである、請求項1に記載のデバイス。
- 前記2つ以上の膜が、実質的に乾燥しており、含水量が2%未満である、請求項1に記載のデバイス。
- 前記チャネルがマイクロチャネルを含むか、または前記チャネルが1つのチャネルである、請求項1に記載のデバイス。
- 前記生物学的サンプルが、500μL未満の容積を含む、請求項1に記載のデバイス。
- 前記生物学的サンプルが、血液である、請求項1に記載のデバイス。
- 前記2つ以上の膜が、最上部膜および前記最上部膜の下にある膜を含み、前記生物学的サンプルの少なくとも一部分が、前記最上部膜および前記最上部膜の下にある前記膜に接触する、請求項1に記載のデバイス。
- 前記2つ以上の膜が、前記デバイスから取り出されるように構成されている、請求項1に記載のデバイス。
- 前記デバイスが、真空によって、前記サンプル獲得構成要素の前記開口部から前記チャネルを通じて前記2つ以上の膜の上に前記生物学的サンプルを移動させるように構成されている、請求項1に記載のデバイス。
- 前記デバイスが、自然発生の毛細管力、吸収材料を介するウィッキング、またはそれらの組み合わせによって、前記サンプル獲得構成要素の前記開口部から前記2つ以上の膜の上に前記生物学的サンプルを移動させるように構成されている、請求項1に記載のデバイス。
- 前記1つ以上の穿刺要素が、ばねに接続されている、請求項1に記載のデバイス。
- 前記2つ以上の膜が、2つの膜である、請求項1に記載のデバイス。
- 前記2つ以上の膜の各々が、同じ組成を有する、請求項1に記載のデバイス。
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JP2024037159A JP2024083344A (ja) | 2015-09-09 | 2024-03-11 | サンプルの収集、安定化、および保存のためのシステム、方法およびデバイス |
Applications Claiming Priority (11)
Application Number | Priority Date | Filing Date | Title |
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US201562216312P | 2015-09-09 | 2015-09-09 | |
US62/216,312 | 2015-09-09 | ||
US201562260172P | 2015-11-25 | 2015-11-25 | |
US62/260,172 | 2015-11-25 | ||
US201662367056P | 2016-07-26 | 2016-07-26 | |
US62/367,056 | 2016-07-26 | ||
US201662368817P | 2016-07-29 | 2016-07-29 | |
US62/368,817 | 2016-07-29 | ||
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