Development of Nano-space Video Microscopy and its Application to the Analysis of New Pharmacological Dyanamics

1)新型纳米空间分辨率视频显微镜的研制采用视频增强对比度的方法，利用绝缘体获得60 nm的分辨率，提高了视频显微镜的分辨率。用奥林巴斯公司研制的聚光透镜的暗场显微镜成功地观察到了10 nm尺度的金胶体。2)荧光退偏振成像视频显微镜的研制为了观察分子的迁移率，在荧光视频显微镜的光路中，通过旋转滤光盒交替地改变了一组偏振器(平行和垂直)。用图像处理技术分析细胞膜中分子的运动。3)激光定标视频显微镜的研制：视频显微镜可以检测到两束交叉激光交叉点的干涉条纹。新的药理动力学分析方法的发展：4)利用高分辨率视频显微镜，测量了细胞骨架分子在非中性粒细胞吞噬和胞吐过程中的作用。分析了不同药物分子对吞噬和胞外过程的影响，表明细胞骨架分子在吞噬和胞外过程中起着关键作用。用荧光去偏振视频显微镜观察到，当吸收促进分子攻击小肠上皮膜时，其膜分子发生蠕动。采用脉冲流动注射与显微镜观察相结合的方法，对有无祛痰药时的纤毛搏动进行了监测。结果表明，氨溴索对纤毛搏动有促进作用，为药物的药理动力学分析提供了新的工具。

1) Development of New Video-microscopes for Nano-space ResolutionThe resolution of video microscope was improved by video enhanced contrast method and using insulators to get 60nm. Agold colloid in10nm scale was successfully observed by the dark field microscopy whose condenser lens was developed by Olympus Co.,2) Development of a Video Microscopy for Imaging of Fluorescence DepolarizationIn order to visualize the molecular mobility, one set of polarizers(parallel and perpendicular)was alternatively changed by rotating filter cassette in optical path of afluorescence video microscope. The molecular movement in the cell membrane was analyzed by image processing3) Development of Laser Scaling Video-microscopy : The interference fringe pattern at the crossing point of two crossed laser beams was found to be detected by video-microscopy. This fringe can be used as the optical scale for the evaluation of 2-&3-dimensional shapes of cells.Development of Analytical Methods for New Pharmacological Dynamics :4) Using high resolution video microscope, the role of cytoskeleton molecules in the sequence of phagocytosis and exocytosis of aneutrophil was measured. The effect of various medicinal molecules on the time course of phago-and exo-cytosis were analyzed to show the cytoskeleton molecules have akey role for it. Using fluorescence depolarization video-microscopy, it was shown that the membrane molecules of small intestinal epitheliums were squirming when the absorption promoting molecules have attacked on it. The combination of pulse flow injection method and microscopic observation, the beating of bronchial cilium in the presence and absence of expectorants was monitored.It was shown that the beating was promoted by Na-ambroxol.The developed microscopies are found to be new tools for the analysis of pharmacological dynamics of drugs.