Building of a Confocal Scanning Differential Polarization Microscope

本申请是申请延长为期两年的生物仪器赠款的资金，用于建造和测试共焦差分偏振扫描显微镜。该仪器的基本光学和电子学设计和建造已经在我们的实验室完成。该显微镜已在复杂生物系统的原位研究中显示出巨大的潜力。使用这台仪器，我们第一次能够在空间上解析长程手性结构域，其中光捕获复合体II组织在叶绿体基粒中。我们请求再提供三年的经费，以扩大本仪器的能力，以开展：a)显微光谱偏振测量物体中给定位置的测量值。目前，该显微镜可以在固定的照明波长下将物体的光学(线性和圆形)各向异性空间分解到光的衍射极限内。所提出的仪器的建立将使我们能够在研究对象中定位感兴趣的微观区域，并获得该局部区域的圆二色谱和线二色谱：b)发光去偏振成像。利用共焦外照射光学的优点和偏振调制和同步解调的扩展灵敏度，我们提出了一种用于空间分辨发光标记物种的分子动力学的仪器。该仪器将首先应用于肥大细胞中IgE受体的旋转动力学成像。

The present application is a request for renewal of funding on a two-year Biological Instrumentation Grant for the construction and testing of a Confocal Differential Polarization Scanning Microscope. The basic optical and electronic design and building of this instrument has been completed in our laboratory. This microscope has already shown its enormous potential as an analytical tool in the study of complex Biological systems in-situ. Using this instrument, we have been able, for the first time, to resolve spatially the long-range chiral domains in which the Light Harvesting Complex II is organized in the chloroplast grana. We request funding for an additional period of three years to extend the capabilities of the present instrument to carry out: a) Microspectro-polarimetric measurements at a given location in the object. Presently, the microscope can spatially resolve the optical (linear and circular) anisotropy of the object to within the diffraction limits of the light, at a fixed wavelength of illumination. The building of the proposed instrumentation will allow us to locate a microscopic domain of interest in the studies object and to obtain the Circular Dichroism and linear Dichroism spectra of that localized domain: b) Luminescence Depolarization Imaging. Using the advantages of confocal epi-dillumination optics and the extended sensitivity of polarization modulation and synchronous demodulation, we propose to develop the instrumentation to spatially resolve the molecular dynamics of luminescence-labelled species. This instrumentation will be applied first to imaging the rotational dynamics of IgE receptors in mast cells.