Acquisition of an Advanced Flow Cytometer

有了这份提案，我们申请资金购买一台多用途的最先进的流式细胞仪/细胞分选机(Becton Dickinson FACS VantageTM细胞分选机系统或Coulter EPICS Elite ESP系统)，以更新和扩展我们目前的内布拉斯加州大学林肯生物技术中心的多用户流式细胞仪核心研究设施。新设备将取代我们已有14年历史的Coulter EPICS V系统，该系统现在需要频繁维修。这种仪器只剩下很短的寿命了。目前的流式细胞仪也不能满足UNL校园内几名研究人员的需要，他们希望在研究工作中进行双变量染色体分析、多色荧光分析和最先进的染色体和细胞分类。作为一项多功能设施，我们需要一种新的流式细胞仪的配件，它将允许：用户友好的操作系统；多色荧光分析和至少两个激光功能；在更短的时间内进行细胞分选；计算机化的液滴延迟设置；大颗粒分析和分选；将1-100,000个细胞分选到微型托盘中，放到显微镜载玻片或过滤器上。重要的是，我们选择的新流式细胞仪系统是为适应未来的升级和应用而设计的。即使有目前仪器的局限性，25个以上研究实验室的项目仍受益于流式细胞术核心研究设施提供的服务。购买最先进的仪器将更好地为现有用户提供服务，而且肯定会吸引更多的用户，这些用户的需求无法用目前使用的仪器来满足。新仪器的可用性将导致具体研究活动的例子包括：1)为了构建染色体特异DNA文库，对重要植物物种的染色体进行单变量和双变量流核型分析和分类。对番茄和玉米染色体的研究已经在进行中。我们将积极寻求世界各地的基因组研究人员的合作，他们希望从其他重要植物中分离出染色体。我们的目标是使我们的流式细胞术核心研究设施成为国际公认的植物基因组分析和染色体分选研究中心。2)用流式细胞术分析动物和植物(包括农作物和天然植物)的细胞核，以及动物和植物的细胞系，以确定倍性水平和/或估计核DNA含量。3)免疫荧光分析(1-4色)，并对血细胞亚群和细胞系进行分类，以确定表面抗原、受体或内部物质的表达。

With this proposal, we request funding for the purchase of a multi-purpose state-of-the-art flow cytometer / cell sorter (Becton Dickinson FACS VantageTM Cell Sorter system or Coulter EPICS Elite ESP system) to update and expand our current multi-user Flow Cytometry Core Research Facility of the University of Nebraska-Lincoln (UNL) Center for Biotechnology. The new equipment will replace our fourteen year-old Coulter EPICS V system which now requires frequent repairs. This instrument has only a short life span remaining. The present flow cytometer is also inadequate for the needs of several investigators on the UNL campus who wish to perform bivariate chromosome analysis, multicolor fluorescence analysis, and state-of-the-art sorting of chromosomes and cells in their research endeavors. As a multipurpose facility, we are requesting accessories for a new flow cytometer which will permit: a user friendly operating system; multicolor fluorescence analysis and at least two laser capability; cell sorting in less time; computerized drop-delay setup; large-particle analysis and sorting; sorting of 1-100,000 cells into microtitier trays, onto microscope slides or filters. Importantly, the new flow cytometer system we have chosen is designed to accommodate future upgrades and applications. Even with the limitations of the present instrumentation, projects in over 25 research laboratories have benefited from the services provided by the Flow Cytometry Core Research Facility. Acquisition of a state-of-the-art instrument will better service current users and most certainly will attract additional users whose needs cannot be met with the instrument presently in use. Examples of specific research activities that would result from the availability the new instrument include: 1) Univariate and bivariate flow karyotyping and sorting of chromosomes from important plant species for the purpose of construction of chromosome-specific DNA libraries. Work is already in progress with tomato and corn chromosomes. We will actively seek collaboration from genome researchers throughout the world who want sorted chromosomes from other important plants. Our goal is to make our Flow Cytometry Core Research Facility an internationally recognized center for plant genome analysis and chromosome sorting research. 2) Flow cytometric analysis of nuclei of animals and plants (both crop plants and native plants), and cell lines from animals and plants for the determination of ploidy level and/or estimation of nuclear DNA content. 3) Immunofluorescence analysis (1-4 color) and sorting of blood cell subsets and cell lines for the expression of surface antigens, receptors or internal substances.