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Development of multiphoton laser fabrication system for tissue engineering

用于组织工程的多光子激光制造系统的开发

基本信息

批准号：
13355004
负责人：
NAKAMURA Osamu
金额：
$ 22.96万
依托单位：
Osaka University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

项目摘要

We developed a three-dimensional laser fabrication system which can be applied for modeling living tissue structure in cultural condition. A mode-locked Ti : Sapphire laser (pulse width : 80-100fs, wavelength : 750-900nm, repetition rate : 82MHz) was used as a light source and was focused into a photopolymerizable material. At the focusing point, the material has multiphoton absorption and is polymerized. Since the polymerization area is localized within the focal volume, scanning of the focus enables us to create any shape of structures in three dimensions. Scanning laser in x-y directions was made by a pair of galvanometer mirrors, and scanning for z direction was done by scanning sample with using a piezoelectric stage. The size of structures which can be fabricated by the developed system is about 500μm x 500μm x 100μm with the spatial resolution of about 200nm when NA1.0 objective lens is used. The fabricated three-dimensional structure was used as a scaffold which control cell growth in three-dimensional space. For culturing cells on a structure, we used photocurable gelatin as a photopolymerizable material. We optimized experimental conditions for fabrication, such as laser power, exposure time, pulse width, wavelength of the laser, and concentration of gelatin. We also investigated the maximum size of structures that can be fabricated in order to confirm the performance of the system. By culturing and observing rat cardiomyocytes and human cancer cells on the structure, we confirmed that growth of both type of cells can be controlled with the fabricated microstructure.

我们开发了一套三维激光加工系统，可用于在培养条件下模拟活体组织结构。锁模Ti：使用蓝宝石激光器（脉冲宽度：80- 100 fs，波长：750- 900 nm，重复频率：82 MHz）作为光源并聚焦到可光聚合的材料中。在聚焦点处，材料具有多光子吸收并聚合。由于聚合区域位于焦点体积内，因此焦点的扫描使我们能够在三维中创建任何形状的结构。利用振镜实现了激光在x-y方向的扫描，利用压电平台实现了样品在z方向的扫描。当使用NA 1.0物镜透镜时，该系统可加工出的结构尺寸约为500 μ m × 500μm × 100μm，空间分辨率约为200 nm。所制造的三维结构被用作在三维空间中控制细胞生长的支架。为了在结构上培养细胞，我们使用光固化明胶作为光聚合材料。我们优化了制备的实验条件，如激光功率、曝光时间、脉冲宽度、激光波长和明胶浓度。我们还研究了可以制造的结构的最大尺寸，以确认系统的性能。通过在该结构上培养和观察大鼠心肌细胞和人类癌细胞，我们证实了这两种类型的细胞的生长都可以通过制造的微结构来控制。