A Field Emission, Intermediate Voltage Electron Microscope for Structural Biology

本申请建议购买飞利浦CM 300-FEG电子显微镜，最大加速电压为300 keV，场发射照明。CM 300-FEG是完全计算机化的，具有革命性的测角仪和功能，特别适合电子断层扫描和冷冻电子显微镜（cryoEM）。 将购买一个专用的保持器和抗污染器，以便于生物标本的冷冻电镜，并从Gatan购买一个配有Macintosh计算机系统的CCD摄像机，以便于开发计算机化的在线数据收集，并获得关于标本制备质量的即时反馈。该仪器的功能将增强6名全职教师的研究，他们将电子显微镜作为其研究的一个组成部分。研究项目包括昆虫飞行肌的电子断层扫描（Taylor）、辅肌动蛋白（Taylor）和Ca 2 +-ATP酶（Taylor）的电子晶体学、烟草花叶病毒（Caspar）定向凝胶的电子衍射、猪蛔虫（Roberts Stewart）主要精子蛋白（MSP）螺旋丝的三维重建&、丝状细菌病毒和皮利的三维重建（Makowski），免疫复合物的三维几何形状的研究（Roux）和细胞骨架结构的研究（Keller）。该仪器将加强这些项目（1）通过对较厚样品的高分辨率成像，（2）通过在线图像记录在数据收集之前对样品进行数字测量，（3）通过其电动和计算机化的样品台，能够开发用于断层扫描的自动倾斜系列数据收集协议，（4）通过促进定向的电子衍射，细丝的多层阵列，（5）通过促进蛋白质和多蛋白质复合物的2-D晶体和准晶体阵列的高分辨率成像，（7）通过利用场发射电子源的高相干性和由300 keV电子提供的高焦深来实现单分子和小复合物的高分辨率成像。

This application proposes the purchase of Philips CM300-FEG electron microscope with a maximum 300 kev accelerating voltage and field emission illumination. The CM300-FEG is fully computerized with a revolutionary goniometer and capabilities that are particularly well designed for electron tomography and cryoelectron microscopy (cryoEM). A specialized holder and anticontaminator will be purchased to facilitate cryoEM of biological specimens and a CCD camera from Gatan with Macintosh computer system to facilitate development of computerized on-line data collection and to obtain instant feedback on the quality of specimen preparation. The capabilities of this instrument will enhance the research of 6 full time faculty who use electron microscopy as an integral part of their research. Research projects include electron tomography of insect flight muscle (Taylor), electron crystallography of actinin (Taylor) and the Ca2+-ATPase (Taylor), electron diffraction of oriented gels of tobacco mosaic virus (Caspar), 3-D reconstruction of helical filaments of the major sperm protein (MSP) from Ascaris suum (Roberts & Stewart), 3-D reconstruction of filamentous bacteria viruses and pili (Makowski), studies of the 3-D geometry of immune complexes (Roux) and studies on the structure of the cytoskeleton (Keller). This instrument will enhance these projects (1) through high resolution imaging of thicker specimens, (2) through digital survey of specimens prior to data collection by means of on-line image recording, (3) by enabling development of protocols for automated tilt series data collection for tomography by virtue of its motorized and computerized specimen stage (4) by facilitating electron diffraction of oriented, multilayered arrays of filaments, (5) by facilitating high resolution imaging of 2-D crystalline and paracrystalline arrays of proteins and multiprotein complexes, (7) by enabling high resolution imaging of single molecules and small complexes by utilizing the high c oherence of the field emission electron source and the high depth of focus provided by 300 kev electrons.