Development of real-time analysis system of molecular interactions using molecular imaging technique

2008 Fiscal Year Final Research Report

• Project/Area Number: 18370066
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Biophysics
• Research Institution: Kyoto University (2008); Tokyo Metropolitan Organization for Medical Research (2006-2007)
• Principal Investigator: HARADA Yoshie Kyoto University, 物質-細胞統合システム拠点, 教授 (10202269)
• Co-Investigator(Kenkyū-buntansha): 横田 浩章 京都大学, 物質-細胞統合システム拠点, 特定拠点講師 (90415547)
• Project Period (FY): 2006 – 2008
• Keywords: マイクロアレイ / 全反射顕微鏡 / 生体分子 / マイクロビーズ

Research Abstract

生体分子を結合させたマイクロビーズをガラス基板上にアレイ化する技術と全反射蛍光顕微鏡を組み合わせ、生体分子間相互作用を高感度に解析できる顕微鏡システムの開発を行った。このシステムは、微量な試料で複数の生体分子間の結合・解離過程を同時に実時間解析することができる。このシステムを使って、抗原-抗体反応、タンパク質-DNA 間相互作用、低分子化合物-タンパク質間の相互作用解析だけでなく、抗体を利用し、生物試料抽出液中の特定タンパク質の濃度を結合速度から定量できることを示した。

Research Products

• [Journal Article] Single-molecule visualization of binding modes of helicase to DNA on PEGylated surface (2009)
  • Author(s): H.Yokota, Y.-W. Han, J.-F. Allemand, X.-G. Xi, D. Bensimon,V. Croquette,Y. Ito, and Y. Harada
  • Journal Title: Chemistry Letters 38 Pages: 308-309
• [Journal Article] Single-cell chemical lysis method for analyses of intracellular molecules using an array of picoliter-scale microwells Anal (2008)
  • Author(s): Y. Sasuga, T. Iwasawa, K. Terada, Y. Oe, H. Sorimachi, O. Ohara, and Y. Harada
  • Journal Title: Chem 80 Pages: 9141-9149
• [Journal Article] Direct observation of the reversible unwinding of a single DNA molecule caused by the intercalation of ethidium bromide (2007)
  • Author(s): Masahito Hayashi, and Yoshie Harada
  • Journal Title: Nucleic Acids Research 35(19) Pages: e125
• [Journal Article] Direct observation of DNA rotation during branch migration of Holliday junction DNA by Escherichia coli RuvA-RuvB protein complex (2006)
  • Author(s): Yong-Woon Han, Tomomi Tani, Masahito Hayashi, Takashi Hishida, Hiroshi Iwasaki, Hideo Shinagawa and Yoshie Harada
  • Journal Title: Proc. Natl. Acad Sci. USA 103(31) Pages: 11544-11548
• [Journal Article] Development of a microscopic platform for real-time monitoring of biomolecular interactions (2006)
  • Author(s): Yasuhiro Sasuga, Tomomi Tani, Masahito Hayashi, Hisashi Yamakawa, Osamu Ohara, and Yoshie Harada
  • Journal Title: Genome Res 16 Pages: 132-139
• [Presentation] Direct observation of DNA rotation during branch migration of Holliday junction DNA by Escherichia coli RuvA-RuvB protein complex (2009)
  • Author(s): Yong-Woon Han, Tomomi Tani, Masahito Hayashi, Takashi Hishida, Hiroshi Iwasaki, Hideo Shinagawa, and Yoshie Harada
  • Organizer: International symposium on Bio-nanosystems
  • Place of Presentation: 仙台
  • Year and Date: 20090901-03
• [Presentation] A novel protein array using microbeads aligned in a microfluidic chip (2009)
  • Author(s): Yoshihiro Ooe, Yasuhiro Sasuga Osamu Ohara and Yoshie Harada
  • Organizer: Biophysical Society 53^<rd> Annual Meeting
  • Place of Presentation: Boston USA
  • Year and Date: 20090228-0304
• [Presentation] A simple single-cell lysis method for analyses of intracellular molecules using a picoliter-scale microwell array (2009)
  • Author(s): Yasuhiro Sasuga, Yoshihiro Ooe, Osamu Ohara, Yoshie Harada
  • Organizer: The 6^<th> Asian Biophysics Association (ABA) Synposium 2009
  • Place of Presentation: Hong Kong
  • Year and Date: 20090111-15
• [Presentation] Harada A novel protein array using microbeads aligned in a microfluidic chip (2008)
  • Author(s): Yoshihiro Ooe, Ysuhiro Sasuga, Osamu Ohara and Yoshie Harada
  • Organizer: 日本生物物理学会 第46 回年会
  • Place of Presentation: 福岡
  • Year and Date: 20081203-05
• [Presentation] Single-molecule Observation of DNA/helicase Interaction by Novel Microscopy.. The first iCeMS Symposium (2008)
  • Author(s): Yokota, H., Han, Y-W., Allemand, J-F., Xie, X., Croquette, V., Bensimon, D. and Harada, Y
  • Organizer: featuring mesoscopic interactions in cells and cellular membranes and the 11th International Membrane Research Forum
  • Place of Presentation: Kyoto City
  • Year and Date: 20080220-22
• [Presentation] Single-molecule Observation of DNA/helicase Interaction by Novel Microscopy (2008)
  • Author(s): Yokota, H., Han, Y-W., Allemand, J-F., Xie, X., Croquette, V., Bensimon, D., Harada, Y
  • Organizer: Joint Meeting of the Biophysical Society 52^<nd> Annual Meeting &amp; 16^<th> International Biophysics Congress
  • Place of Presentation: Long Beach
  • Year and Date: 20080202-06
• [Presentation] Real Time Quantitation of an Endogenous mRNA in Single Living Cells (2008)
  • Author(s): Okabe, K., Harada, Y. and Funatsu, T
  • Organizer: Joint Meeting of the Biophysical Society 52^<nd> Annual Meeting &amp; 16^<th> International Biophysics Congress
  • Place of Presentation: Long Beach
  • Year and Date: 20080202-06
• [Presentation] Effect of Zipcode Sequence and Molecular Weight of mRNA on Its Subcellular Localization Revealed by Single Molecule Imaging (2008)
  • Author(s): Yamagishi, M., Ishihama, Y., Shirasaki, Y., Kurama, H. Sasuga, Y., Terada, K., Harada, Y., Funatsu, T
  • Organizer: Joint Meeting of the Biophysical Society 52^<nd> Annual Meeting &amp; 16^<th> International Biophysics Congress
  • Place of Presentation: Long Beach
  • Year and Date: 20080202-06
• [Presentation] Studies on biomolecules using single-molecule imaging and manipulation techniques (2008)
  • Author(s): Yoshie Harada
  • Organizer: 8^<th> International Conference on Excitonic Processes in Condensed Matter (EXCON'08)
  • Place of Presentation: Kyoto (Invited)
  • Year and Date: 2008-06-26
• [Presentation] マイクロ流路とマイクロビーズを組み合わせた新規生体分子間相互作用検出法 (2008)
  • Author(s): 大江良洋、貴家康尋、原田慶恵
  • Organizer: 第8 回日本蛋白質科学会年会
  • Place of Presentation: 東京
  • Year and Date: 2008-06-11
• [Presentation] マイクロビーズアレイとマイクロウェルを使った1細胞タンパク質解析法の開発 (2008)
  • Author(s): 原田慶恵
  • Organizer: バイオナノ研究会
  • Place of Presentation: 宮城 青根温泉
  • Year and Date: 2008-05-09
• [Presentation] Single-molecule experiments on DNA motors The First iCeMS Symposium and the 11th Membrane Research Forum (2008)
  • Author(s): Yoshie Harada
  • Place of Presentation: Kyoto
  • Year and Date: 2008-02-21
• [Presentation] マイクロウェルアレイを用いた単一細胞分泌タンパク質解析法の開発 (2007)
  • Author(s): 貴家康尋, 寺田佳代子, 大江良洋, 小原收, 原田慶恵
  • Organizer: 日本生物物理学会第45 回年会
  • Place of Presentation: 横浜
  • Year and Date: 20071221-23
• [Presentation] 一細胞中の酵素活性測定技術の開発 (2007)
  • Author(s): 岩澤朋之、貴家康尋、原田慶恵
  • Organizer: 2007 年生体運動研究研究合同班会議
  • Place of Presentation: 金沢
  • Year and Date: 20070107-09
• [Presentation] マイクロビーズを利用したin vitro生体分子間相互作用解析法と単一細胞解析技術の開発 (2007)
  • Author(s): 貴家康尋, 小原收, 原田慶恵
  • Organizer: BMB2007 ワークショップ
  • Place of Presentation: 横浜(招待講演)
  • Year and Date: 2007-12-11
• [Presentation] マイクロ流路を利用した迅速なタンパク質相互作用検出システムの開発 (2007)
  • Author(s): 薄井(青木)一恵, 貴家康尋, 岡崎規理子,小原玲子, 原田慶恵, 小原收
  • Organizer: BMB2007
  • Place of Presentation: 横浜
  • Year and Date: 2007-12-11
• [Presentation] Labeling of iron-sulfur protein with a fluorescent dye to detect its conformation change in the cytochrome bc1complex (2006)
  • Author(s): Miki, T., Sakamoto, S., Yokota, H., and Harada, Y
  • Organizer: Fifth East Asian Biophysics Symposium &amp; Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Okinawa, Japan
  • Year and Date: 20061112-16
• [Presentation] Direct observation of T7 RNA polymerase rotation during transcription (2006)
  • Author(s): Kato, Y., Yokota, H., Hayashi, M., Terada, K., Sasuga, Y. and Harada, Y
  • Organizer: Fifth East Asian Biophysics Symposium &amp; Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Ginowan City,Japan
  • Year and Date: 20061112-16
• [Presentation] Direct observation of the twisting motion of a single DNA molecule caused by the intercalation of ethidium bromide (2006)
  • Author(s): Hayashi, M. and Harada, Y
  • Organizer: Fifth East Asian Biophysics Symposium & Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Ginowan City, Japan
  • Year and Date: 20061112-16
• [Presentation] Novel microscopy for simultaneous single molecule measurement of DNA/protein interaction (2006)
  • Author(s): Yokota, H., Han, Y-W., Allemand, J-F., Xi, X., Croquette V. , Bensimon, and D., Harada Y
  • Organizer: Fifth East Asian Biophysics Symposium &amp; Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Ginowan City, Japan
  • Year and Date: 20061112-16
• [Presentation] Live imaging of growth cone regeneration after axotomy of dorsal root ganglion neurons (2006)
  • Author(s): Nomura, M., Tani, T., and Harada, Y
  • Organizer: Fifth East Asian Biophysics Symposium &amp; Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Ginowan City, Japan
  • Year and Date: 20061112-16
• [Presentation] Real Time Imaging of Specific Messenger RNA in a Living Cell Using Artificial Nucleic Acids (2006)
  • Author(s): Okabe, K., Harada, Y. and Funatsu, T
  • Organizer: Fifth East Asian Biophysics Symposium &amp; Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Ginowan City, Japan
  • Year and Date: 20061112-16
• [Presentation] Development of a microscopic platform for single-cell analyses using picoliter microwell array (2006)
  • Author(s): Sasuga, Y., Iwasawa, T., Terada, K., Sorimachi, H., Ohara, O. and Harada, Y
  • Organizer: Fifth East Asian Biophysics Symposium &amp; Forty-Fourth Annual Meeting of the Biophysical Society of Japan
  • Place of Presentation: Ginowan City, Japan
  • Year and Date: 20061112-16
• [Presentation] Development of a microscopic platform for real-time monitoring of biomolecular interactions (2006)
  • Author(s): Sasuga, Y., Tani, T., Hayashi, M., Yamakawa, H., Ohara, O. and Harada, Y
  • Organizer: Gordon Research Conference 2006 Single Molecule Approaches To Biology
  • Place of Presentation: New London, USA
  • Year and Date: 20060617-22
• [Presentation] A simple single-cell chemical lysis method for analyses of intracellular molecules using a picoliter-scale microwell array Gordon Research Conference
  • Author(s): Sasuga, Y., Iwasawa, T., Ooe, Y., Ohara, O., Harada, Y
• [Book] バイオマシン・生物分子モーター (2008)
  • Author(s): 原田慶恵、横田浩章
  • Total Pages: 878-883
  • Publisher: 細胞工学秀潤社(Vol.25 No.8)
• [Book] 1分子が見えた!.1枚の写真館 (2008)
  • Author(s): 原田慶恵
  • Total Pages: 425
  • Publisher: 細胞工学(27(5))
• [Book] ナノバイオ辞典, テクノシステム (2007)
  • Author(s): 原田慶恵
  • Publisher: 一分子観察, 蛍光顕微鏡, 蛍光色素, 光ピンセット
• [Book] 一分子蛍光法化学と生物 (2007)
  • Author(s): 原田慶恵
  • Total Pages: 340-346
  • Publisher: 日本農芸化学会 会誌(vol.45 No.5)
• [Book] DNA モーター生物物理学ハンドブック (2007)
  • Author(s): 原田慶恵
  • Total Pages: 353-356
  • Publisher: 朝倉書店
• [Book] エヌ・ティー・エス (2007)
  • Author(s): 原田慶恵
  • Total Pages: 738-747
  • Publisher: DNA モーター非侵襲・可視化技術ハンドブック
• [Book] 光ピンセット.生命科学のための機器分析実験ハンドブック (実験医学別冊) (2007)
  • Author(s): 横田浩章, 原田慶恵
  • Total Pages: 99-104
  • Publisher: 第3章顕微解析4.(羊土社)
• [Book] 1分子イメージング. 生命科学のための機器分析実験ハンドブック(実験医学別冊) (2007)
  • Author(s): 原田慶恵
  • Total Pages: 237-242
  • Publisher: 第7章イメージング解析 3.(羊土社)
• [Book] 光トラップ 分子細胞生物学辞典
  • Author(s): 原田慶恵
  • Publisher: 光トラップ 分子細胞生物学辞典
• [Patent(Industrial Property Rights)] マイクロ流路形成体を利用したマイクロビーズアレイ用チップ、マイクロビーズアレイ及びこれらを用いた被検物質を検出する方法
  • Industrial Property Number: 特許出願2007-195476