Building of a Confocal Scanning Differential Polarization Microscope

目前申请的是一项为期两年的生物仪器拨款的续期申请，用于共聚焦差偏振扫描显微镜的建造和测试。本仪器的基本光学和电子设计与制造已在实验室完成。这种显微镜已经显示出其巨大的潜力，作为一种分析工具，在复杂的生物系统的原位研究。利用该仪器，我们首次能够在空间上分辨出叶绿体颗粒中光收获复合体II组织的远程手性结构域。我们要求额外拨款三年，以扩大现有仪器的能力，以便进行：a)在物体的给定位置进行显微光谱偏振测量。目前，显微镜可以在空间上分辨物体的光学各向异性（线性和圆形），在光的衍射极限内，在固定波长的照明。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域，并获得该局部域的圆二色性和线性二色性光谱：b)发光去偏振成像。利用共聚焦外延照光学的优势以及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解析发光标记物种分子动力学的仪器。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.