Production of novel proteins using the micro system that permits sorting and functional evaluation at single molecular level

使用允许在单分子水平上进行分类和功能评估的微系统生产新型蛋白质

• 批准号: 13558088
• 负责人: FUNATSU Takashi
• 金额: $ 9.02万
• 依托单位: Waseda University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13558088/
• 关键词: Micro-system; Molecule Sorter; Single-molecule Probe

We have developed a micro fabricated fluorescence-activated bio-molecular sorter using thermosensitive hydrogel as a microvalve. The sorter was composed of a microfluidic chip with Y-shaped channels with one inlet and two outlets. Biological specimen containing fluorescently labeled cells and thermosensitive polymer was introduced into a Y-shaped microchannel. Thermosensitive polymer changed the state from sol to gel when it was heated. The sol-gel transition was locally induced by site directed infrared laser irradiation in order to plug the one of the microchannel and collect the cells. As an application of this sorter, GFP-expressing E. coll. CD8-expressing lymphocytes labeled with fluorescence antibody, and DNA molecules labeled with TOTO-1 were separated. The sorting time of 120 ms and recovery ratio of 90 % was achieved by this sorter.

我们已经开发了使用热敏水凝胶作为微脱毛的微型荧光激活的生物分子分散蛋白。该分散器由带有Y形通道的微流体芯片组成，带有一个入口和两个插座。将含有荧光标记的细胞和热敏聚合物的生物标本引入Y形微通道中。热敏聚合物加热时，热敏聚合物将状态从SOL变为凝胶。 Sol-Gel转变是通过位置的定向红外激光照射局部诱导的，以插入微通道之一并收集细胞。作为该分散者的应用，表达GFP的E. Coll。分离用荧光抗体标记的CD8表达淋巴细胞，并分离用Toto-1标记的DNA分子。该分类器实现了120毫秒的排序时间和90％的回收率。

项目成果

Fujiwara, I., S.Takahashi, H.Tadakuma, T.Funatsu, S.Ishiwata: "Microscopicanalysis of polymerization dynamics with individual actin filaments"Nature Cell Biol.. 4. 666-673 (2002)

Fujiwara, I., S.Takahashi, H.Tadakuma, T.Funatsu, S.Ishiwata：“用单个肌动蛋白丝进行聚合动力学的微观分析”Nature Cell Biol.. 4. 666-673 (2002)

座古保, 船津高志: "生体機能を1分子レベルで探る"分光研究. 51. 3-14 (2002)

Zakoyasu，Takashi Funatsu：“探索单分子水平的生物功能”光谱研究。51. 3-14 (2002)。

多田隈尚史, 船津高志: "生きた細胞の核内mRNAの1分子蛍光イメージング"細胞工学. 20. 672-677 (2001)

Takashi Tadakuma、Takashi Funatsu：“活细胞中核 mRNA 的单分子荧光成像”《细胞工程》20. 672-677 (2001)。

Funatsu, T. et al.: "Rapid and sensitive detection method of a bacterium using GFP reporter phage"Microbiol.Immuno.. 46. 365-369 (2002)

Funatsu, T. 等人：“使用 GFP 报告噬菌体的细菌快速灵敏检测方法”Microbiol.Immuno.. 46. 365-369 (2002)

Shirasaki, Y. et al.: "A novel biomolecule sorter using thermosensitive hydrogel in micro flow system"Micro Total Analysis System 2002. 925-927 (2002)

Shirasaki, Y. 等人：“在微流系统中使用热敏水凝胶的新型生物分子分选机”微全分析系统 2002. 925-927 (2002)