CRYOELECTRON MICROSCOPE AND CRYO-ULTRAMICROTOME

请拨资金购买一台高分辨率低温电子设备 显微镜。用户的研究建议需要具备以下能力 观察冰冻水合状态下的颗粒和切片材料。 该显微镜还将用于低剂量和常规 显微镜。 拥有经过同行审查的NIH资金的用户一直在调查结构 在分子水平上使用负染和阴影技术。 这些方法现在已经在很大程度上被观察到的 冷冻水合状态下的未染色标本，保存它们 以一种本质上是本地的形式。这种方法是必要的，以便 回答问题时不沾染、固定或干燥人工制品，并 因为它能够显著提高分辨率 很多情况下。它也是一种观察时间分辨的强大技术 可以通过结合快速冷冻和 例如，来自笼中前体的代谢物的释放。项目 需要使用这种技术包括对分子的研究 肌丝、血红蛋白晶体、DNA-蛋白质的结构 相互作用和包被的囊泡。 冷冻电子显微镜将是加州大学的第一台 马萨诸塞州医疗中心。收购这台仪器的目的是 对于满足用户当前的需求和满足 在项目中启动新的方向，这些项目现在不可能 认识到。这项技术将提供一个天然而强大的补充 对于校园里已经在使用的结构技术， 包括X射线结晶学、细胞的3D光学显微镜研究 结构、高分辨核磁共振与常规和冷冻取代 亚细胞冷冻断口的电子显微镜研究 结构。

Funds are requested for the purchase of a high resolution cryo-electron microscope. Research proposals of the users require the ability to observe particulate and sectioned material in the frozen-hydrated state. The microscope will also be available for low dose and conventional microscopy. Users with peer-reviewed NIH funding have been investigating structures at the molecular level using negative staining and shadowing techniques. These approaches have now been largely superseded by observation of unstained specimens in the frozen-hydrated state, which preserves them in an essentially native form. This approach is necessary in order to answer questions without staining, fixation or drying artifacts and because is capable of providing significantly improved resolution in many cases. It is also a powerful technique for observing time resolved structural changes that can be captured by combining rapid freezing with, for example, the release of metabolites from caged precursors. Projects requiring the use of this technique include studies of the molecular structure of muscle filaments, hemoglobin crystals, DNA-protein interactions and coated vesicles. The cryo-electron microscope will be the first at the University of Massachusetts Medical Center. The acquisition of this instrument is essential to provide for the current needs of users and for the initiation of new directions in projects that are now impossible to realize. This technology will provide a natural and powerful complement to the structural techniques that are already in use on this campus, including X-ray crystallography, 3D light microscopic studies of cellular structure, high resolution NMR and conventional and freeze-substitution and freeze fracture electron microscopic studies of subcellular structure.