ShEEP Request for Nikon TIRF STORM microscope

超分辨率显微镜为成像单个分子或多组分子提供了前所未有的机会。 分辨率低于50 nm的分子。然而，具有这些功能的显微镜，如果从 商业供应商提供的服务，如准备组装，超出了大多数调查员的预算。和切割 建造自己的建筑需要对光学的理解，这超出了大多数人的知识范围。 生物学家与爱荷华州市退伍军人医疗中心有关的多名调查人员已经达到了一个临界点， 将超分辨率显微镜应用于活细胞的研究需要将他们的研究 性新这项研究针对的是与美国退伍军人相关的疾病：肺 肺气肿，慢性感染如丙型肝炎、巨细胞病毒或 利什曼病，青光眼和肥胖症。目前在弗吉尼亚州或爱荷华州大学还没有显微镜 校园，这将使这些类型的研究。因此，我们要求尼康Eclipse Ti-e2 配备有全内反射（TIRF）和随机光学重建的倒置显微镜 显微镜（STORM）模块。TIRF光学将有利于那些研究受体的信号通过组装 衔接蛋白在质膜（T细胞受体，博士主教和斯台普顿）或保留活性 进入内体后的信号传导（McGowan博士）。结合TIRF和STORM将使麦高恩博士， 艾伦和安德森研究的组装podosomes或重塑皮层下的细胞骨架。 STORM将使Wilson、Meier和斯台普顿博士能够观察病原体的蛋白质组装， 这些来自宿主细胞的信息可以用来了解病毒或寄生虫如何逃避宿主的防御。Meier博士将使用STORM来 研究病毒或哺乳动物蛋白质的靶向降解或病毒蛋白质与染色质的相互作用。所有 这些项目中的一个需要高分辨率的荧光成像，以检测和定位光子， 帧速率快，方向和时间精度高。我们选择了尼康H-TIRF/N STORM系统，因为它在一个包中满足了所有这些要求， 这对于多用户仪器是非常期望的。为了实现活细胞成像， 显微镜将配备一个阶段顶部培养箱，以控制环境，并允许试剂 在不改变视野的情况下引入。我们还要求强大的PC工作站和尼康 获取和分析图像所需的软件。更完整的仪器说明可 在具体目标部分可以找到。因为这个包涉及新兴的最先进的技术，我们 我希望它也将加强新的研究人员的研究计划，因为他们加入退伍军人管理局的研究计划 并追求针对美国退伍军人未来健康需求的新项目。

Super-resolution microscopy has opened unprecedented opportunities to image single molecules or groups of molecules with resolution below 50 nm. However microscopes with these capabilities, if purchased from a commercial vendor as ready to assemble, are beyond the budgets of most individual investigators. And cutting expenses by building one's own requires an understanding of optics which is beyond the knowledge of most biologists. Multiple investigators associated with the Iowa City VA Medical Center have reached a point in their investigations where applying super-resolution microscopy to living cells is required to move their research forward. This research is targeted towards diseases which are relevant to US military veterans: pulmonary emphysema, alterations in host defense observed in chronic infections such as hepatitis C, cytomegalovirus, or Leishmaniasis, glaucoma and obesity. There is currently no microscope on the VA or University of Iowa campuses which will enable these types of studies. Therefore, we are requesting a Nikon Eclipse Ti-e2 inverted microscope equipped with total internal reflection (TIRF) and stochastic optical reconstruction microscopy (STORM) modules. TIRF optics will benefit those studying receptors which signal by assembling adapter proteins at the plasma membrane (T-cell receptor, Drs. Bishop and Stapleton) or retain active signaling after entering endosomes (Dr. McGowan). Combining TIRF and STORM will enable Drs. McGowan, Allen and Anderson to study the assembly of podosomes or remodeling of the sub-cortical cytoskeleton. STORM will enable Drs. Wilson, Meier, and Stapleton to observe the assembly of proteins from pathogens with those from host cells to learn how viruses or parasites evade host defenses. Dr. Meier will use STORM to study targeted degradation of viral or mammalian proteins or the interaction of viral proteins with chromatin. All of these projects require multicolor fluorescent imaging, at high resolution, to detect and localize photons, at rapid frame rates and with high directional and temporal precision. We have chosen the Nikon H-TIRF/N- STORM system because it meets all of these requirements in a package which provides these features in a largely automated format, which is highly desirable for a multi-user instrument. To enable live cell imaging the microscope will be equipped with a stage-top incubator to control the environment and allow reagents to be introduce without altering the field of view. We are also requesting powerful PC workstations and Nikon software which is required to acquire and analyze images. A more complete description of the instrument may be found in the Specific Aims section. Because this package involves emerging state-of the art technology, we hope that it will also enhance the research programs of new investigators as they join the VA research program and pursue novel projects tailored to the future health needs of US veterans.