Cell Sorting

项目总结(见说明)： 细胞分选设备(CSF)为福克斯大通癌症中心(Fccc)的研究人员提供了通过多达14种独立测量来分析单细胞悬液的能力，其中12种基于免疫荧光，两种基于光散射。在2009日历年，CCSG研究小组五个项目中的四个项目的31名同行评审、资助的调查人员使用了该机制。大的 大多数设施的使用(98.5%)是由同行审查、资助的调查人员使用的。流式细胞术的主要应用包括识别和分离(分选)细胞亚群(高达20,000个/秒)，分离罕见的变异(1/10‘)细胞亚群，肿瘤或细胞系的特征，以及通过直接沉积到96孔板中进行细胞亚群的单细胞克隆。目前的分拣机提供高速4流 分类和数字检测。具有紫外线功能的激光可以测量钙离子通量和活细胞DNA含量。除了3激光12色细胞分选机(FACS-VantageSE/DIVA)外，该设施还包括一台仅供分析的10色LSR-II和一台三色FACScan。2009年，该中心为海湾生物系统公司JSAN台式细胞分选机提供了资金(240,000美元)。用户操作的JSAN能够进行高达8色的2流分选，并利用3光束激励和流动池检测，包括UV激光。该设施向用户提供关于实验设计(包括试剂和适当的荧光试剂的可用性)、样品制备和数据解释的建议。FLOW-LIMS是利用fccc生物统计和生物信息学设施(BBF)开发的软件，用于数据注释、组织和 存档。分析是在研究人员的实验室计算机上使用高度交互的图形软件程序FlowJo进行的。使用时数，每年2,778小时(目前估计超过理论容量的)，自上次审查以来增加了34%，并预计在未来五年内随着所有五个项目的计划招募人员而增加，哈代(免疫细胞发展和宿主防御) 计划)，自1987年以来一直指导这一资源，并负责设施运营。通过设施政策、设施咨询委员会(FAC)和管理设施母公司监督委员会(FPOC)的监督，确保公平使用设施。

PROJECT SUMMARY (See instructions): The Cell Sorting Facility (CSF) offers Fox Chase Cancer Center (FCCC) investigators the capability of analyzing single cell suspensions by up to 14 independent measurements, 12 of which are based on immunofluorescence and two on light scattering. In calendar year 2009 the Facility was used by 31 peer-reviewed, funded investigators across four of the five CCSG Research Programs. The large majority of Facility usage (98.5%) was by peer-reviewed, funded investigators. Major applications of flow cytometry include discrimination and isolation (sorting) of cell subpopulations (at up to 20,000 cells/sec), isolation of rare variant (1/10'') cell subsets, characterization of tumors or cell lines, and single cell cloning of cell subpopulations by direct deposition into 96-well plates. The current sorter provides high-speed 4-stream sorting and digital detection. A UV-capable laser enables measurement of calcium flux and viable cell DNA content. In addition to the 3-laser 12-color cell sorter (FACS-VantageSE/DiVa), the Facility also includes an analysis-only 10-color LSR-II and a 3-color FACScan. In 2009 the Center provided funds ($240,000) for a new Bay Biosystems JSAN desktop cell sorter. The user-operated JSAN is capable of up to 8-color 2-stream sorting and utilizes 3-beam excitation with flow-cell detection, including a UV laser. The Facility aids users with advice on experimental design (including availability of reagents and appropriate fluorochromes), sample preparation, and data interpretation. Software developed with the FCCC Biostatistics and Bioinformatics Facility (BBF), the Flow-LIMS, is used for data annotation, organization, and archiving. Analysis is done on investigators' laboratory computers using a highly interactive, graphical software program, FlowJo. Hours of usage, 2,778/year (currently estimated at over 64% of theoretical capacity), has increased 34% since the last review and is projected to increase over the next five years with planned recruits across all five Programs, Hardy (Immune Cell Development and Host Defense (ICDHD) Program), has directed this resource since 1987 and has responsibility for Facility operations. Equitable access to the Facility is assured by Facility policy, oversight from a Facility Advisory Committee (FAC) and a governing Facilities Parent Oversight Committee (FPOC).