Development of Laser Microbeam Cell Micromanipulation and Its Bioengineering Application

激光生物学正在开拓新的前沿领域，激光微束细胞微操作技术正在开创新的生物技术。该系统本质上是一个倒置的显微镜，其恒温器连接到He-Ne激光器和<Ar^+>激光器，用于微束细胞照射。每一束激光都由柔性光纤引导，光纤可以连接多个激光装置、激光束选择器和显微镜。通过激光束选择器，选择所需的激光照射到样品上。在TV监视器屏幕上目视观察样品的相位对比图像，并在激光照射后的一段时间内记录在录像机和帧存储器中。激光的焦点，通过一个双色镜，叠加在试样图像和聚焦照射在任何所需的地点的试样是通过移动机械阶段获得。激光束被引导到快门控制器，快门控制器在给定的时间段内释放激光器的快门。用<micro>10倍物镜可获得直径为10 μ m的聚焦光斑，并利用该系统进行了活细胞的显微穿孔和显微切割等细胞显微手术。激光照射核仁导致巨细胞形成。激光微束细胞显微操作系统在研究激光对不同类型细胞的影响方面可能比活体系统更有用。该系统在研究激光对细胞器亚细胞水平的影响方面具有许多优势。

The new frontier of laser biology is being developed and a novel biotechnique of cell micromanipulation by laser microbeam is being pioneered. The system is essentially an inverted microscope with a thermostat interfaced to a He-Ne laser and a <Ar^+> laser, employed for microbeam cell irradiation. Each laser beam is guided by flexible optical fiber which can connect between several laser apparatus, a laser beam selector and then a microscope. By a laser beam selector, the desired laser is selected to be irradiated to the specimen. The phase-contrasted image of the specimen is visually observed on a TV monitor screen and registered in a video tape recorder and a frame memory for a period of time after the laser irradiation. The focal spot of the laser, through a dichromatic mirror, is superimposed on the specimen image and the focused irradiation at any desired site of the specimen is acquired by moving the mechanical stage. The laser beam is guided to the shutter controller which releases the shutter of the laser for a given period of time. The focused laser spot size of 10 <micro> m in diameter is obtainable with a 10-fold objective.Cell microsurgery such as microperforation and microdissection of living cells was performed by laser microbeam using this system. Laser irradiation on the nucleolus resulted in giant cell formation. Also the possibility of cell fusion by laser irradiation was shown.The present system of laser microbeam cell micromanipulation is probably more useful for studying the effects of laser irradiation on various type of cells than the in vivo systems. And our system has many advantages for detailed investigation of the effects of laser irradiation on the subcellular level of organella.