Laser Scanning Cytometer

描述(由申请人提供)：我们正在申请一台计算机ICYS激光扫描细胞仪(LSC)的资金。LSC是一种相对较新开发的仪器，它结合了传统显微镜和流式细胞术的许多优点，显著扩展了单细胞分析的范围。它不像显微镜那样随机地对整个视野成像，而是选择、成像和分析满足某些可调标准(如大小或荧光)的细胞。与传统的流式细胞术不同，LSC通常用于分析固定在水平表面的细胞，如载玻片上的培养细胞或各种培养容器中的培养细胞。活细胞可以成像和细胞计量学分析荧光信号，各种类型的固定标本也可以用这种方法进行研究。该仪器使用自动化的纳米级X-Y工作台来移动样品通过物镜，施加多达三种不同的激光来激发荧光发射，并使用四个光电倍增管(PMT)探测器阵列对这些荧光进行定量。因此，LSC本质上是一种强大的混合技术，它在单一平台上结合了成像和细胞分析。重要的是，随着LSC图像成为参数，详细的显微图像被存储并直接参考细胞学数据。我们计划广泛使用的该仪器的一个非常吸引人的功能是，它能够从荧光标记的组织切片的微观区域收集定量荧光数据，并以在许多情况下可与传统流式细胞术获得的真实细胞测量数据相媲美的方式显示和分析此类数据。我们计划将ICYS LSC安置在我们完善的中央流式细胞术核心实验室并进行操作，那里将有充足的专业知识和监督，以确保该仪器得到充分利用。核心实验室有一位经验丰富的资深教员(史蒂文·波塞利博士)作为其主管，并雇用了三名全职教员级别的助理来指导操作和提供技术监督。还有一批内部顾问，他们在与开发将使用LSC的应用程序密切相关的领域具有专门知识。核心实验室拥有完善的财务计划和严密的监督，并从该机构和我们的NCI资助的癌症中心获得大量的机构支持，以确保其高效和稳定的运行。通过将ICYS LSC纳入这个核心实验室，从事许多由NIH资助的各种基础和翻译研究项目的大量研究人员将获得这项重要新技术的便捷和负担得起的机会。公共卫生相关性：这项申请申请的资金将允许我们的流式细胞仪核心实验室获得一种新的分析仪器，称为Compucyte ICYS激光扫描细胞仪。这台仪器将为我们机构的许多研究人员提供一种强大的新方法来分析细胞和组织。使用该仪器进行的研究将促进我们对与生物医学科学高度相关的广泛领域的了解，包括癌症生物学、免疫学、自身免疫性疾病和传染病。

DESCRIPTION (provided by applicant): We are requesting funds for a Compucyte iCys Laser Scanning Cytometer (LSC). The LSC is a relatively recently developed instrument that joins many of the advantages of traditional microscopy and flow cytometry, and significantly extends the limits of single cell analyses. Rather than randomly imaging an entire field like a microscope does, it selects, images and analyzes cells that meet certain adjustable criteria (such as size or fluorescence). Unlike conventional flow cytometry, the LSC is usually used to analyze cells fixed to horizontal surface, such as cultured cells on slides or in a variety of culture vessels. Live cells can be imaged and cytometrically analyzed for fluorescent signals, and a variety of types of fixed specimens can also be studied in this way. The instrument uses an automated nanostep X-Y stage to move samples through the objective lens, applying up to three different lasers to excite fluorescent emissions and quantitating these using an array of four photomultiplier tube (PMT) detectors. Thus, the LSC is essentially a powerful hybrid technology that combines imaging and cytometric analysis in a single platform. Importantly, with the LSC image becomes a parameter, and detailed microscopic pictures are stored and referenced directly to cytometric data. A highly appealing feature of the instrument that we plan to use extensively is its ability to gather quantitative fluorescence data from microscopic areas of fluorescently labeled tissue sections, and to display and analyze such data in a manner that is comparable in many cases to true cytometric data such as one obtains with traditional flow cytometry. We plan to house and operate the iCys LSC in our well established central Flow Cytometry Core Laboratory, where there will be ample expertise and oversight to ensure that the instrument is used to its full capacity. The Core Laboratory has an experienced senior faculty member (Dr. Steven Porcelli) as its supervisor, and employs three full time faculty level associates to direct operations and provide technical supervision. There is also a range of internal consultants with expertise in areas of major relevance to the development of applications that will use the LSC. The Core Laboratory has a well developed and carefully monitored financial plan, and receives substantial institutional support from the institution and from our NCI-funded Cancer Center to ensure its efficient and stable operation. By incorporating the iCys LSC into this core laboratory, a large body of investigators pursuing many diverse NIH-funded basic and translational research projects will gain convenient and affordable access to this important new technology. Public Health Relevance: This application requests funding that will allow our Flow Cytometry Core Laboratory to acquire a new analytical instrument called the Compucyte iCys Laser Scanning Cytometer. This instrument will provide many researchers at our institution with a powerful new way to analyze cells and tissues. The studies that will be done with this instrument will advance our understanding of a wide spectrum of areas with high relevance to biomedical science, including cancer biology, immunology, autoimmune diseases and infectious diseases.