Acquisition of a DeltaVision OMX Super-Resolution Imaging System

描述（由申请人提供）：生物学家的一个主要目标是“看到”细胞内部，发现每个蛋白质在何时何地起作用。传统的光学显微镜方法将我们的空间分辨率限制在~250纳米，在一些情况下，这只比生物体本身略小。这个经典的物理限制已经被超分辨率显微镜的发展所突破。我们的建议是获得DeltaVision OMX超分辨率显微镜系统，这将允许我们的用户实现亚衍射亚细胞分辨率，以解决一些重要的生物学问题。该系统的强大之处在于超分辨率是通过传统的激光和计算机技术实现的，显微镜系统是标准的高端宽视场显微镜，可以使用传统的荧光团，无需开发新的试剂，通过使用多条激光线可以实现多达4种颜色的标记，并且系统可以由经验丰富的细胞生物学家维护和操作。实际上，该系统在横向和轴向上都将分辨率屏障扩展到约150nm。该系统还设计为能够获得表达荧光蛋白的活细胞的极快同时多色成像。该系统的速度和分辨率在任何其他仪器中都是无与伦比的。该仪器将允许我们的用户接近广泛的问题跨越三个重要的生物医学领域（微生物生物学，染色质生物学和有丝分裂），是由现有的显微镜方法的限制。微生物细胞生物学的一个主要障碍是，医学上重要的微生物，如肺炎链球菌，只有~0.5 m，仅比光学显微镜的分辨率略大，这使得对蛋白质亚细胞分布的成像几乎不可能。不对称在细菌分化和生物膜形成中的交流中非常重要，强调了能够以高分辨率动态成像这些生物体中成分分布的重要性。同样，染色质的表观遗传修饰对基因表达至关重要，越来越多的证据表明，特定的DNA位点在染色质结构中具有空间定义。确定染色质标记何时以及如何分布提供了超分辨率成像如何对重要研究领域产生影响的额外示例。最后，细胞骨架的动态组织对正确的细胞形态和有丝分裂过程至关重要。超分辨率成像能够识别重要的动态调节蛋白在何时何地起作用，超高速成像将使我们对微管细胞骨架的动态调节有新的认识。总的来说，拟议的仪器将对我校生命科学研究人员的研究方法和进展产生重大影响。

DESCRIPTION (provided by applicant): A major goal of biologists is to "see" inside cells to discover where and when each individual protein acts. Conventional light microscopy approaches limit our spatial resolution to ~250 nm, which in several instances is only slightly smaller than the organism itself. This classic physical limitation has been broken through the development of Super-Resolution microscopy. Our proposal is to acquire the DeltaVision OMX SuperResolution microscope system that will allow our users to achieve sub-diffraction subcellular resolution to address a number of important biological problems. The power of this system is that the super resolution is achieved with conventional lasers and computer technology, the microscope system is a standard high-end widefield microscope, conventional fluorophores can be used obviating the need to develop new reagents, up to 4 color labeling can be achieved by the use of multiple laser lines, and the system can be maintained and operated by an experienced cell biologist. Practically speaking, the system has extended the resolution barrier to approximately 150 nm in both the lateral and axial direction. The system is also engineered to be able to acquire extremely rapid simultaneous multi-color imaging of living cells expressing fluorescent proteins. The speed and resolution of this system are unsurpassed in any other instrument. This instrumentation will allow our users to approach a wide array of questions spanning three important biomedical areas (microbial biology, chromatin biology, and mitosis) that are limited by existing microscopy methods. A major obstacle in microbial cell biology is that medically important microbes, such as Streptococcus pneumoniae are only ~0.5 5m, which is only slightly larger than the resolution of the light microscope, making imaging of the subcellular distribution of proteins virtually impossible. Asymmetry is very important in bacterial differentiation and communication in biofilm formation, highlighting the importance of being able to dynamically image the distribution of components in these organisms at high resolution. Likewise the epigenetic modifications of chromatin are critical to gene expression, and increasing evidence shows that specific DNA sites are spatially defined in chromatin structure. Identifying when and how chromatin marks are distributed provide additional examples of how super resolution imaging will make an impact on an important area of research. Finally, the dynamic organization of the cytoskeleton is critical for proper cell morphology and for mitotic progression. SuperResolution imaging has the capacity to identify when and where important dynamics regulatory proteins act, and the super speed imaging will allow us to gain novel insights into the dynamic regulation of the microtubule cytoskeleton. Overall the proposed instrumentation would have a significant impact on the research approaches and progress made by life sciences researchers at our university.