Spectromicroscopy for biochemical analysis of sperm

描述(由申请人提供)：我们建议发展以下能力 X射线光谱显微镜，使其能够应用于生物化学成像 正常和不正常的精子。这将通过聚焦“软”X射线来实现(在此 例如，光子能量为200-800 eV的X射线)到最小的远场 聚焦任何波长的电磁辐射，并扫描干燥或 冷冻水化的标本穿过焦点形成图像。图像在一个 然后一系列的光子能量将被组合在一起，以提供 整个精子的化学态敏感的X射线吸收光谱 空间分辨率优于50 nm。从这个数据来看，主要的生物化学 单个精子的成分将通过与参照物的拟合来确定 分离化合物的光谱；此外，预期的和可能的意外的 组件之间的空间相关性将使用原理进行研究 成分分析方法。这项工作将使用x射线进行。 我们小组在石溪开发的显微镜，它在软X射线下工作 布鲁克海文国家同步加速器光源上的波荡器光束线 国家实验室。增加更好的显微镜将使显微镜得到改进。 用于定量光谱分析的有序分类光学，并通过配备低温冷凝器 具有更精确的压电台和更高效率的探测器的显微镜 用于研究冷冻的水化样品，而不会造成过度的辐射损害。 这些仪器的开发将以我们研究 精子形态变异与生化变异的相关性 以更好地了解男性不育的原因。通过获得x射线 不育男性不同精子形态的光镜数据 希望对体外受精(IVP)临床医生选择精子有一定的指导作用 卵胞浆内单精子注射(ICSI)以提高成功率 程序的速率。将制作光谱显微数据分析软件 在开发过程中，可供其他研究人员免费下载。

Description (provided by applicant): We propose to develop the capabilities of x-ray spectromicroscopy to allow it to be applied to biochemical imaging of normal and abnormal sperm. This will be done by focusing "soft" X rays (in this case, x rays with a photon energy of 200-800 eV) to the smallest far-field focus of electromagnetic radiation of any wavelength, and scanning a dry or frozen hydrated specimen through that focal spot to form an image. Images at a series of photon energies will then be combined to deliver chemical-state-sensitive x-ray absorption spectra from an entire sperm at better than 50 nm spatial resolution. From this data, major biochemical constituents in individual sperm will be determined by fitting to reference spectra of isolated compounds; furthermore, expected and possibly unexpected spatial correlations between components will be studied using principal component analysis methods. This work will be carried out using x-ray microscopes developed by our group at Stony Brook which operate at a soft x-ray undulator beamline at the National Synchrotron Light Source at Brookhaven National Laboratory. The microscopes will be improved by the addition of better order sorting optics for quatitative spectroscopy, and by equipping a cryo microscope with a more accurate piezo stage and a higher efficiency detector for studies of frozen hydrated specimens without excessive radiation damage. These instrumentation developments will be guided by our goal of studying the correlation between morphological and biochemical variations in sperm, in order to better understand the causes of male infertility. By obtaining x-ray spectromicroscopic data on different sperm morphologies in infertile males, we hope to guide the in vitro fertilization (IVP) clinician in the choice of sperm use for intracytoplasmic sperm injection (ICSI) so as to improve the success rate of the procedure. Spectromicroscopic data anaysis software will be made available for free downloading by other researchers as it is developed.