Development of molecular imaging microscope for biological specimen and its application for tissue examination

生物标本分子成像显微镜的研制及其在组织检查中的应用

• 批准号: 15300161
• 负责人: ARAKI Tsutomu
• 金额: $ 10.69万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15300161/
• 关键词: bio-SHG; nonlinear optical effect; ultra-short duration pulse laser; collagen molecule; tissue diagnosis; microscopic image; skin

Aim of this research project is to develop an effective method that can sense objective biomolecles without staining and to apply this method to identification of the situation of the molecles. For ths purpose, we have focused on non-linear optical effect in the tissue that is induced by the irradiation with high-intensity very short duration pulsed laser light such as a femtosecond laser. In the actual research, we have developed three microscopes ; fluorescence lifetime imaging microscope in nanosecond range, a collagen sensitive microscope using second harmonic generation light (SCG light) and a coherent anti Stokes coherent Roman scattering (CARS) microscope.The most successful one is the SHG microscope. Based on the reflection-type microscope with a probe light spot of 10 micrometers in diameter and the polarization measurement of SHG light induced by collagen molecules, we have determined distribution of collagen fiber orientation in various human tissues. The resulting SHG data in human skin imply that the reticular dermis posses a tangled structure of collagen fibers which is consistent with the result of the anatomical examination of the skin. We have improved the SHG microscope to obtain 3D dissection image using the con-focal optics and the 3D sample scanning stage. With this microscope, tomograph of the collage fiber orientation in the human skin and bone have been measured.Performance of the fluorescence lifetime imaging microscope was demonstrated by the measurement of the autofluorescence in liposome. Also we have measured the distribution of hyaluronic acid around an erythrocyte with CARS microscope.

本研究的目的是开发一种不需要染色就能检测目标生物分子的有效方法，并将其应用于分子状态的识别。为此目的，我们已经集中在组织中的非线性光学效应，这是由高强度极短持续时间的脉冲激光，如飞秒激光照射引起的。在实际研究中，我们研制了三种显微镜：纳秒级荧光寿命成像显微镜、二次谐波发生光（SCG光）胶原敏感显微镜和相干反斯托克斯相干罗马散射（汽车）显微镜，其中最成功的是二次谐波显微镜。利用直径为10 μ m的反射式显微镜和胶原分子诱导的二次谐波光的偏振测量，我们测定了人体各种组织中胶原纤维的取向分布。在人皮肤中得到的SHG数据暗示网状真皮是胶原纤维的缠结结构，这与皮肤的解剖学检查的结果一致。我们改进了SHG显微镜，利用共焦光学系统和三维样品扫描平台获得三维解剖图像。用该显微镜对人皮肤和骨中胶原纤维的取向进行了断层扫描，并通过对脂质体中自发荧光的测量验证了荧光寿命成像显微镜的性能。我们还用汽车显微镜测量了透明质酸在红细胞周围的分布。

项目成果

Measurement of 3D orientation of collagen fiber in tissue using a femtosecond laser light

使用飞秒激光测量组织中胶原纤维的 3D 取向

• 发表时间: 2004
• 期刊: Proc.18th Japan Soc. ME & BME
• 作者: Kudo;Y;Tsutomu Araki
• 通讯作者: Tsutomu Araki

フェムト秒パルス光を用いた多重散乱光 除去型グルコース濃度測定法の開発

开发利用飞秒脉冲光消除多重散射光的葡萄糖浓度测定方法

• 发表时间: 2004
• 期刊: 日本エム・イー学会 生体医工学シンポジウム (CD-ROM)
• 作者: J.Nagayama;H.Tsuji;T.Iida;R.Nakagawa;T.Matsueda;H.Hirakawa;J.Fukushige;T.Watanabe;池昌子;堀 泰明 他
• 通讯作者: 堀 泰明 他

生体の非線形光学効果を用いた組織コラーゲン線維のin situ配向分布測定

利用生物非线性光学效应原位测量组织胶原纤维的取向分布

• 发表时间: 2003
• 期刊: Optics Japan 2003講演予稿集
• 作者: Ohsawa;S;安井武史
• 通讯作者: 安井武史

Determination of collagen orientation in skin dermis based on optical nonlinear effect in biological tissue.

基于生物组织光学非线性效应测定皮肤真皮中胶原蛋白的取向。

• 发表时间: 2003
• 期刊: Proc.2003 JSME Mechanical Engin.Congr. Vol.5
• 作者: Yoshisuke Nonaka;Sadanori Konishi;Masato Ohmi;Takeshi Yasui
• 通讯作者: Takeshi Yasui

Characterization of collagen orientation in human dermis by two-dimensional second-harmonic-generation polarimetry.

• DOI: 10.1117/1.1644116
• 发表时间: 2004-03
• 期刊: Journal of biomedical optics
• 影响因子: 3.5
• 作者: T. Yasui;Y. Tohno;T. Araki