JP6456969B2 - 膨張顕微鏡法 - Google Patents
膨張顕微鏡法 Download PDFInfo
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- JP6456969B2 JP6456969B2 JP2016553349A JP2016553349A JP6456969B2 JP 6456969 B2 JP6456969 B2 JP 6456969B2 JP 2016553349 A JP2016553349 A JP 2016553349A JP 2016553349 A JP2016553349 A JP 2016553349A JP 6456969 B2 JP6456969 B2 JP 6456969B2
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Description
本願は、その内容が、その全体において参照により本明細書に援用される2014年2月21日に出願された米国仮特許出願番号61/943,045の利益を主張する。
本発明は、国立衛生研究所により認められた助成金番号1DP1NS087724の下、米国政府の支援によりなされた。政府は、本発明に一定の権利を有する。
本発明は、顕微鏡法、特に生物学的標本の光学的画像化に関する。
顕微鏡法は、固定された細胞および組織中の小さな構造の画像を光学的に拡大することにより、価値のある生物学的情報を提供している。しかしながら、かかる画像化技術の解像度は、発光源の波長の約半分に限定される。スペクトルの可視領域において、これは1ミクロンの1/4〜1/3のオーダーである(250〜330nm)。不幸にも、従来の光学的顕微鏡法ではこの長さの尺度未満の非侵襲性の観察が利用できない非常に多くの範囲の生物学的構造/系がある。
本発明は、生物学的標本自体を物理的に膨張することに基づく、古典的な顕微鏡法回析限界よりも良好な解像度での生物学的標本の光学的画像化のための方法である。この方法において、培養された細胞、固定された組織、または原則として、生物学的材料を含む他の型の目的の試料に、組成物または化学カクテルを注入し、該組成物または化学カクテルで試料材料を包埋し、次いで該組成物を、好ましくはナノスケールの正確さで三次元において全方向に等しく膨張させ得る。
本発明は、顕微鏡法のための拡大された目的の試料、および目的の試料を拡大するための方法、標本中で膨潤性(swellable)ポリマー網目状組織を合成することによる古典的顕微鏡法回析限界よりも良好な解像度での目的の試料の光学的画像化に関し、標本は、物理的に膨張され、物理的な拡大がもたらされ得る。本発明はまた、目的の試料を拡大するための本明細書に記載される組成物の使用を含む。
図1は、本発明の方法の例示的な態様における工程を示すフローチャートである。
Claims (49)
- 顕微鏡法のための目的の試料を拡大するための方法であって、該方法は、
目的の試料を膨潤性(swellable)材料に埋め込む工程、ここで、該試料を膨潤性材料に埋め込む工程が、該試料に、膨潤性材料の前駆体を含む組成物を浸透化させること、およびインサイチュで膨潤性材料を形成することを含む、
該試料を、膨潤前に膨潤性材料に固定する工程、ならびに
該材料を膨潤させることにより該試料を拡大する工程
を含み、該試料を膨潤させる前に、該試料を、機械的、物理的、化学的、生化学的または酵素的な消化に供する、方法。 - 該試料が標識される、請求項1記載の方法。
- 該試料が、免疫蛍光染色、免疫組織化学的染色または免疫細胞化学的染色により標識される、請求項2記載の方法。
- 該組成物が、1つ以上の水溶性モノマー前駆体を含む水溶液である、請求項1記載の方法。
- 該溶液が、アクリレート、アクリルアミドおよびN,N-メチレンビスアクリルアミドを含む、請求項4記載の方法。
- 該膨潤性材料がポリアクリレートである、請求項1〜5いずれか記載の方法。
- 該試料が、水を添加することにより全方向に等しく膨潤され、拡大された試料を生じる、請求項1記載の方法。
- 該試料に膨潤性材料の前駆体を浸透化させる前に、該試料を界面活性剤で処理する、請求項1記載の方法。
- 顕微鏡法のための目的の拡大された試料を調製するための方法であって、該方法は、
目的の試料を多価電解質ヒドロゲルに埋め込む工程、ここで、該試料を多価電解質ヒドロゲルに埋め込む工程が、該試料に、多価電解質ヒドロゲルの前駆体を含む組成物を浸透化させること、およびインサイチュで多価電解質ヒドロゲルを形成することを含む、
該試料を、膨潤前に多価電解質ヒドロゲルに固定する工程、ならびに
該ヒドロゲルを膨潤させることにより該試料を拡大する工程
を含み、該試料を膨潤させる前に、該試料を、機械的、物理的、化学的、生化学的または酵素的な消化に供する、方法。 - 該試料が標識される、請求項9記載の方法。
- 該試料が、免疫蛍光染色、免疫組織化学的染色または免疫細胞化学的染色により標識される、請求項10記載の方法。
- 該多価電解質ヒドロゲルの前駆体が、アクリレート、アクリルアミドおよびN,N-メチレンビスアクリルアミドを含む、請求項9記載の方法。
- 多価電解質ヒドロゲルがポリアクリレートである、請求項9〜12いずれか記載の方法。
- 該試料に多価電解質ヒドロゲルの前駆体を浸透化させる前に、該試料を界面活性剤で処理する、請求項9記載の方法。
- 該試料が、水を添加することにより全方向に等しく膨潤され、拡大された試料を生じる、請求項9記載の方法。
- 該拡大された試料を顕微鏡下で見分することにより試料の高解像度画像を作製する工程をさらに含む、請求項1〜15いずれか記載の方法。
- 顕微鏡下で該拡大された試料を検分することにより該拡大された試料を光学的画像化する工程をさらに含む、請求項1〜15いずれか記載の方法。
- (a)細胞または組織の試料と、結合部分およびアンカーを含むタグを接触させる工程、ここで、該結合部分は、該試料の生体分子成分に結合する;
(b)該試料に、膨潤性材料の前駆体を含む組成物を浸透化させる工程;
(c)重合を開始してインサイチュで膨潤性材料を形成する工程、ここで、該アンカーは、該膨潤性材料に共有結合して、試料−膨潤性材料複合体を形成する;ならびに
(d)水性の溶媒または液体を添加して、試料−膨潤性材料複合体を膨潤させ、それにより該複合体を物理的に拡大させ、全方向に等しく拡大された試料を生じる工程
を含む、細胞または組織の試料を全方向に等しく拡大するための方法。 - 該試料が標識される、請求項18記載の方法。
- 該試料が、免疫蛍光染色、免疫組織化学的染色または免疫細胞化学的染色により標識される、請求項19記載の方法。
- 該液体が水である、請求項18記載の方法。
- 浸透化工程を行う前に、該試料を界面活性剤で処理する、請求項18記載の方法。
- 工程(d)の前に、該試料−膨潤性材料複合体を、機械的、物理的、化学的、生化学的または酵素的な消化に供する、請求項18記載の方法。
- 該膨潤性材料の前駆体を含む組成物が、1つ以上の水溶性モノマー前駆体を含む水溶液である、請求項18記載の方法。
- 該溶液が、アクリル酸ナトリウム、アクリルアミドおよびN,N-メチレンビスアクリルアミドを含む、請求項18〜24いずれか記載の方法。
- 該膨潤性材料がポリアクリレートである、請求項18〜24いずれか記載の方法。
- (a)細胞または組織の試料と、結合部分およびアンカーを含むタグを接触させる工程、ここで、該結合部分は、該試料の生体分子成分に結合する;
(b)該試料に、ヒドロゲルの前駆体を含む組成物を浸透化させる工程;
(c)重合を開始してインサイチュでヒドロゲルを形成する工程、ここで、該アンカーは、該ヒドロゲルに共有結合して、試料−ヒドロゲル複合体を形成する;ならびに
(d)水性の溶媒または液体を添加して、試料−ヒドロゲル複合体を膨潤させ、それにより該複合体を物理的に拡大させ、全方向に等しく拡大された試料を生じる工程
を含む、細胞または組織の試料を全方向に等しく拡大するための方法。 - 該試料が標識される、請求項27記載の方法。
- 該試料が、免疫蛍光染色、免疫組織化学的染色または免疫細胞化学的染色により標識される、請求項28記載の方法。
- 該液体が水である、請求項27記載の方法。
- 浸透化工程を行う前に、該試料を界面活性剤で処理する、請求項27記載の方法。
- 工程(d)の前に、該試料−ヒドロゲル複合体を、機械的、物理的、化学的、生化学的または酵素的な消化に供する、請求項27記載の方法。
- 該ヒドロゲルが多価電解質ヒドロゲルである、請求項27〜32いずれか記載の方法。
- 該ヒドロゲルの前駆体を含む組成物が、少なくとも1つの多価電解質モノマーおよび共有結合架橋剤を含む、請求項33記載の方法。
- 該ヒドロゲルの前駆体が、アクリル酸ナトリウム、アクリルアミドおよびN,N-メチレンビスアクリルアミドを含む、請求項34記載の方法。
- 該多価電解質ヒドロゲルがポリアクリレートである、請求項33記載の方法。
- 該拡大された試料を顕微鏡下で見分することにより試料の高解像度画像を作製する工程をさらに含む、請求項18〜36いずれか記載の方法。
- 顕微鏡下で該拡大された試料を検分することにより該拡大された試料を光学的画像化する工程をさらに含む、請求項18〜36いずれか記載の方法。
- (a)細胞または組織の試料と、結合部分およびアンカーを含むタグを接触させる工程、ここで、該結合部分は、該試料の生体分子成分に結合する;
(b)該試料に、膨潤性材料の前駆体を含む組成物を浸透化させる工程;
(c)重合を開始してインサイチュで膨潤性材料を形成する工程、ここで、該アンカーは、該膨潤性材料に共有結合して、試料−膨潤性材料複合体を形成する;ならびに
(d)水性の溶媒または液体を添加して、試料−膨潤性材料複合体を膨潤させ、それにより該複合体を物理的に拡大させ、全方向に等しく拡大された試料を生じる工程
を含む方法により作製される、全方向に等しく拡大された細胞または組織の試料。 - 該試料が標識される、請求項39記載の試料。
- 該試料が、免疫蛍光染色、免疫組織化学的染色または免疫細胞化学的染色により標識される、請求項40記載の試料。
- 該液体が水である、請求項39記載の試料。
- 浸透化工程を行う前に、該試料を界面活性剤で処理する、請求項39記載の試料。
- 工程(d)の前に、該試料−膨潤性材料複合体を、機械的、物理的、化学的、生化学的または酵素的な消化に供する、請求項39記載の試料。
- 該膨潤性材料がヒドロゲルである、請求項39〜44いずれか記載の試料。
- 該ヒドロゲルが多価電解質ヒドロゲルである、請求項45記載の試料。
- 該膨潤性材料の前駆体を含む組成物が、少なくとも1つの多価電解質モノマーおよび共有結合架橋剤を含む、請求項45記載の試料。
- 該ヒドロゲルの前駆体が、アクリル酸ナトリウム、アクリルアミドおよびN,N-メチレンビスアクリルアミドを含む、請求項47記載の試料。
- 該多価電解質ヒドロゲルがポリアクリレートである、請求項46記載の試料。
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Families Citing this family (53)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CA2850509C (en) | 2011-10-14 | 2023-08-01 | President And Fellows Of Harvard College | Sequencing by structure assembly |
US11021737B2 (en) | 2011-12-22 | 2021-06-01 | President And Fellows Of Harvard College | Compositions and methods for analyte detection |
EP4249605A3 (en) | 2011-12-22 | 2023-11-15 | President And Fellows Of Harvard College | Methods for analyte detection |
WO2013184754A2 (en) | 2012-06-05 | 2013-12-12 | President And Fellows Of Harvard College | Spatial sequencing of nucleic acids using dna origami probes |
US10138509B2 (en) | 2013-03-12 | 2018-11-27 | President And Fellows Of Harvard College | Method for generating a three-dimensional nucleic acid containing matrix |
KR102282990B1 (ko) | 2013-06-04 | 2021-07-28 | 프레지던트 앤드 펠로우즈 오브 하바드 칼리지 | Rna-가이드된 전사 조절 |
CN118240918A (zh) | 2013-06-25 | 2024-06-25 | 普罗格诺西斯生物科学公司 | 采用微流控装置的空间编码生物分析 |
CN106715768B (zh) | 2014-07-30 | 2020-06-16 | 哈佛学院院长及董事 | 用于测定核酸的系统和方法 |
EP4119677B1 (en) | 2015-04-10 | 2023-06-28 | Spatial Transcriptomics AB | Spatially distinguished, multiplex nucleic acid analysis of biological specimens |
US10526649B2 (en) | 2015-04-14 | 2020-01-07 | Massachusetts Institute Of Technology | Augmenting in situ nucleic acid sequencing of expanded biological samples with in vitro sequence information |
US10059990B2 (en) | 2015-04-14 | 2018-08-28 | Massachusetts Institute Of Technology | In situ nucleic acid sequencing of expanded biological samples |
US11408890B2 (en) | 2015-04-14 | 2022-08-09 | Massachusetts Institute Of Technology | Iterative expansion microscopy |
CN108139408B (zh) * | 2015-08-07 | 2020-08-28 | 麻省理工学院 | 蛋白质保持扩展显微法 |
CA2994958C (en) * | 2015-08-07 | 2024-02-13 | Massachusetts Institute Of Technology | Nanoscale imaging of proteins and nucleic acids via expansion microscopy |
GB2559526B (en) | 2015-11-03 | 2021-02-17 | Harvard College | Method and apparatus for volumetric imaging of a three-dimensional nucleic acid containing matrix |
JP6909227B2 (ja) * | 2016-02-25 | 2021-07-28 | マサチューセッツ インスチチュート オブ テクノロジー | 臨床組織標本を膨張させるための方法 |
US20170276578A1 (en) * | 2016-03-22 | 2017-09-28 | University Of Washington | Expansion microscopy methods and kits |
CN116200465A (zh) | 2016-04-25 | 2023-06-02 | 哈佛学院董事及会员团体 | 用于原位分子检测的杂交链反应方法 |
US11397140B2 (en) | 2016-04-29 | 2022-07-26 | Massachusetts Institute Of Technology | Methods for reversible and tunable tissue magnification |
EP3472359B1 (en) | 2016-06-21 | 2022-03-16 | 10X Genomics, Inc. | Nucleic acid sequencing |
GB2570412A (en) | 2016-08-31 | 2019-07-24 | Harvard College | Methods of generating libraries of nucleic acid sequences for detection via fluorescent in situ sequencing |
GB2569252A (en) * | 2016-08-31 | 2019-06-12 | Harvard College | Methods of combining the detection of biomolecules into a single assay using fluorescent in situ sequencing |
ES2969671T3 (es) * | 2016-11-08 | 2024-05-21 | Harvard College | Obtención de imágenes con multiplexación usando MERFISH, microscopía de expansión y tecnologías relacionadas |
WO2018136856A1 (en) | 2017-01-23 | 2018-07-26 | Massachusetts Institute Of Technology | Multiplexed signal amplified fish via splinted ligation amplification and sequencing |
WO2018157048A1 (en) | 2017-02-24 | 2018-08-30 | Massachusetts Institute Of Technology | Methods for examining podocyte foot processes in human renal samples using conventional optical microscopy |
CN106959240B (zh) * | 2017-03-28 | 2019-09-24 | 华中科技大学 | 一种反复膨胀过程实现大块组织体的膨胀透明方法 |
WO2018218150A1 (en) | 2017-05-26 | 2018-11-29 | President And Fellows Of Harvard College | Systems and methods for high-throughput image-based screening |
JP7160350B2 (ja) * | 2017-07-06 | 2022-10-25 | 公立大学法人大阪 | 生体組織透明化法及びその試薬 |
WO2019023214A1 (en) | 2017-07-25 | 2019-01-31 | Massachusetts Institute Of Technology | ATAC IN SITU SEQUENCING |
CN111263819A (zh) | 2017-10-06 | 2020-06-09 | 卡特阿纳公司 | Rna模板化连接 |
US11753676B2 (en) | 2017-10-11 | 2023-09-12 | Expansion Technologies | Multiplexed in situ hybridization of tissue sections for spatially resolved transcriptomics with expansion microscopy |
US11479811B2 (en) | 2017-11-21 | 2022-10-25 | Expansion Technologies | Expansion microscopy compatible and multiplexed in situ hybridization of formalin fixed paraffin embedded tissue sections for spatially resolved transcriptomics |
US11873374B2 (en) | 2018-02-06 | 2024-01-16 | Massachusetts Institute Of Technology | Swellable and structurally homogenous hydrogels and methods of use thereof |
US10915729B2 (en) | 2018-02-20 | 2021-02-09 | The Regents Of The University Of Michigan | Three-dimensional cell and tissue image analysis for cellular and sub-cellular morphological modeling and classification |
JP7082364B2 (ja) * | 2018-03-02 | 2022-06-08 | 日本電信電話株式会社 | フィルム状ソフトマテリアル、蛍光フィルム基材及びフィルム状ソフトマテリアルの製造方法、並びに応力計測方法 |
CN112020638A (zh) * | 2018-04-25 | 2020-12-01 | 加利福尼亚大学董事会 | 使用具有紫外表面激发的显微镜检查对组织的组织学层面三维成像 |
US11414701B2 (en) | 2018-05-24 | 2022-08-16 | The Broad Institute, Inc. | Multimodal readouts for quantifying and sequencing nucleic acids in single cells |
WO2020013833A1 (en) * | 2018-07-13 | 2020-01-16 | Massachusetts Institute Of Technology | Dimethylacrylamide (dmaa) hydrogel for expansion microscopy (exm) |
CN112770776A (zh) | 2018-07-30 | 2021-05-07 | 瑞德库尔有限责任公司 | 用于样品处理或分析的方法和系统 |
US11549135B2 (en) | 2018-09-14 | 2023-01-10 | The Broad Institute, Inc. | Oligonucleotide-coupled antibodies for single cell or single complex protein measurements |
KR102368585B1 (ko) * | 2018-09-28 | 2022-03-03 | 서울대학교산학협력단 | 온도에 따라 가역적 팽창/초기화가 가능한 하이드로젤을 이용한 생체조직 이미징 |
EP3928074A4 (en) * | 2019-02-22 | 2022-11-30 | Massachusetts Institute of Technology | ITERATIVE DIRECT EXPANSION MICROSCOPY |
CN110006862B (zh) * | 2019-03-29 | 2020-09-08 | 华中科技大学 | 膨胀切削显微成像方法及适用于该方法的超吸水水凝胶 |
SG11202111878RA (en) | 2019-05-31 | 2021-11-29 | 10X Genomics Inc | Method of detecting target nucleic acid molecules |
WO2021113505A1 (en) | 2019-12-05 | 2021-06-10 | Massachusetts Institute Of Technology | Method for preparing a specimen for expansion microscopy |
EP4153606A2 (en) | 2020-07-13 | 2023-03-29 | Singular Genomics Systems, Inc. | Methods of sequencing complementary polynucleotides |
EP4121561A4 (en) | 2020-08-06 | 2024-04-17 | Singular Genomics Systems, Inc. | METHODS FOR IN SITU TRANSCRIPTOMICS AND PROTEOMICS |
EP4121557A4 (en) | 2020-08-06 | 2024-05-01 | Singular Genomics Systems, Inc. | SPATIAL SEQUENCING |
US11333588B1 (en) * | 2020-12-07 | 2022-05-17 | Nebulum Technologies Co., Ltd. | Matrix-assisted methods and compositions to prepare biological samples for super-resolution imaging |
CN112881671A (zh) * | 2021-01-11 | 2021-06-01 | 东南大学 | 一种基于高密度荧光标记法的细胞微管膨胀显微成像方法 |
US20240084378A1 (en) | 2022-05-11 | 2024-03-14 | 10X Genomics, Inc. | Compositions and methods for in situ sequencing |
DE102022112065B3 (de) | 2022-05-13 | 2023-08-24 | Abberior Instruments Gmbh | Verfahren, vorrichtung und computerprogramm zur mikroskopischen probenabbildung |
KR20240042887A (ko) * | 2022-09-26 | 2024-04-02 | 연세대학교 산학협력단 | 고배율의 세포 확대 분석을 위한 세포 필름, 이의 제조용 조성물 및 제조 방법 |
Family Cites Families (19)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6107081A (en) | 1999-02-05 | 2000-08-22 | The United States Of America As Represented By The Administrator Of The National Aeronautics And Space Administration | Uni-directional cell stretching device |
AU774336B2 (en) * | 1999-04-27 | 2004-06-24 | Ciphergen Biosystems, Inc. | Probes for a gas phase ion spectrometer |
US6287870B1 (en) * | 1999-08-20 | 2001-09-11 | Robert A. Levine | Method and assembly for separating formed constituents from a liquid constituent in a complex biologic fluid sample |
US6878384B2 (en) * | 2001-03-13 | 2005-04-12 | Microvention, Inc. | Hydrogels that undergo volumetric expansion in response to changes in their environment and their methods of manufacture and use |
JP3812829B2 (ja) * | 2002-08-22 | 2006-08-23 | 独立行政法人科学技術振興機構 | 組織の包埋剤、樹脂包埋体及び樹脂包埋体の製造方法 |
JP2005291759A (ja) * | 2004-03-31 | 2005-10-20 | Michimasa Kishimoto | 二次元画像による病症診断システム |
JP4374435B2 (ja) * | 2004-07-28 | 2009-12-02 | 国立大学法人 大分大学 | 高親水性高分子による組織包埋方法。 |
DK2126586T3 (da) * | 2007-02-01 | 2010-10-11 | Immundiagnostik Ag | Direkte bestemmelse af vitamin D i serum eller plasma |
JP5058676B2 (ja) * | 2007-05-18 | 2012-10-24 | 株式会社アイエスティー | 生体標本の作製方法 |
EP2195413A4 (en) * | 2007-08-30 | 2014-01-01 | Harvard College | MULTI-WELL CULTURE PLATE HAVING A HIGH COMPATIBILITY SURFACE |
US20100041128A1 (en) * | 2008-01-08 | 2010-02-18 | Medtrain Technologies, Llc | Microfluidic Device for Application of Shear Stress and Tensile Strain |
KR20110007600A (ko) * | 2008-01-30 | 2011-01-24 | 제론 코포레이션 | 세포배양 제품 및 스크리닝하는 방법 |
JP4956839B2 (ja) * | 2008-02-13 | 2012-06-20 | 国立大学法人 大分大学 | 高親水性高分子モノマー水溶液による組織包埋方法 |
US20090241681A1 (en) * | 2008-03-27 | 2009-10-01 | Andrew Machauf | Hydrogel-based mems biosensor |
RU2010151919A (ru) | 2008-05-20 | 2012-06-27 | Зе Реджентс Оф Зе Юниверсити Оф Калифорния (Us) | Анализ ex vivo клеток с целью детектирования болезненного состояния и выбора и мониторинга терапевтического агента |
WO2011111876A1 (en) * | 2010-03-12 | 2011-09-15 | Riken | Clearing reagent for biological material, and use thereof |
US20140087412A1 (en) | 2011-04-20 | 2014-03-27 | 4Dx Pty Ltd | Method and Device for Application of Fluid Forces to Cells |
JP5967528B2 (ja) * | 2012-06-22 | 2016-08-10 | 国立研究開発法人理化学研究所 | 生物材料を透明化する方法および生物材料用透明化処理キット |
EP4163617A1 (en) * | 2012-08-09 | 2023-04-12 | The Board of Trustees of the Leland Stanford Junior University | Methods and compositions for preparing biological specimens for microscopic analysis |
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