Shared Resource: Cell Sorting Facility

细胞/分子分析共享资源：细胞分选因子（CSF） 项目总结/摘要 细胞分选设施（CSF）为癌症中心成员提供访问和技术支持， 流式细胞术技术，一种用于分析和分离（分选）复杂混合物中的细胞的方法，基于 包括光散射和荧光。在理查德的指导下， 哈代博士（BCDF）和1.3名全职员工的目标是提供最高质量的分析和排序 癌症中心成员的能力。随着流式细胞术技术的成熟， 研究的组成部分，在不同的实验室使用。31名癌症中心成员使用CSF， 2014年日历。2014年CSF中记录的94%的服务时间用于支持同行评审 资助的研究。来自所有5个研究项目的48名癌症中心成员在2011年至2016年期间使用了CSF。 2014.由于仪器的复杂性，成本和口径，服务最好是方便，虽然 中央CSF。在每个实验室都有单独的流式细胞仪是成本高昂的。 流式细胞术分析和/或分选的性质并不真正适合外部（商业） 的服务.也就是说，样品通常是活细胞，在分析前不久制备（因此非常容易腐烂）， 许多实验涉及使用新试剂（荧光染料标记的 单克隆抗体）。哈代28年的经验和专业知识是实施国家的关键， 先进的技术和创新的方法。哈代的领导力加上机构的承诺导致了 购买高度定制的FACSAria（509 500美元）。这种新仪器最佳地激发了PE和PE 串联缀合物与绿色激光，相对于蓝色488 nm激发提供更大的信噪比。的 绿色激光还能够检测使用先前的荧光检测器不能检测到的某些荧光蛋白。 Vantage分拣机。Aria还运行当前版本的仪器控制软件，并具有编程的自动 每天早上在设置时进行校准。LSR-II也进行了升级，通过增加一个绿色 激光器（能够最有效地激发PE和PE串联），取代红色激光器以增加激光 功率，并增加荧光检测器的数量。此仪器还运行当前仪器 控制软件每天早上自动校准。这个自动校准软件的咏叹调和LSR轨道 每个通道的灵敏度，因此提供了额外的质量保证，在 他们会走向失败内部开发的数据管理系统Flow-LIMS允许所有癌症患者 中心成员可在其本地计算机上注释、访问和管理流式细胞术数据 实验室，旨在促进实验数据的共享，并有助于促进合作 实验室之间。CSF由一个专门的设施咨询委员会（FAC）提供咨询， 每年。FAC的建议直接提交给管理机构母公司监督委员会， 以确保CSF继续拥有必要的资源，以提供最高质量的服务。

CELLULAR/MOLECULAR ANALYSES SHARED RESOURCE: CELL SORTING FACILITY (CSF) PROJECT SUMMARY/ABSTRACT The Cell Sorting Facility (CSF) provides Cancer Center members access to and technical support for the technology of flow cytometry, a method for analyzing and isolating (sorting) cells in a complex mixture, based on a variety of parameters, including light scatter and fluorescence. The CSF under the direction of Richard Hardy, PhD (BCDF) and a staff of 1.3 FTEs has a goal to provide the highest quality analytic and sorting capabilities to Cancer Center members. As flow cytometry technology has matured, it has become an essential component of research, utilized in a diverse array of laboratories. 31 Cancer Center members used the CSF in calendar year 2014. 94% of service hours logged in the CSF during 2014 were in support of peer-review funded research. 48 Cancer Center members from all 5 Research Programs used the CSF in the period 2011- 2014. Due to the complexity, cost and caliber of the instrumentation, the services are best facilitated though the centralized CSF. It would be cost prohibitive to have individual flow cytometers available in each laboratory. The nature of flow cytometry analysis and/or sorting is not really appropriate for an external (commercial) service. That is, samples are often live cells, prepared shortly before analysis (therefore quite perishable) and many experiments involve the use of diverse combinations of novel reagents (fluorochrome-labeled monoclonal antibodies). The 28 years of experience and expertise of Hardy is key in implementing state-of- the-art technology and innovative approaches. Hardy's leadership coupled with institutional commitment led to the purchase of a highly customized FACSAria ($509,500). This new instrument optimally excites PE and PE tandem conjugates with a green laser, providing greater signal-to-noise relative to blue 488nm excitation. The green laser also enables detection of certain fluorescent proteins that could not be detected using the previous Vantage sorter. The Aria also runs the current version of instrument control software with a programmed auto- calibration at setup each morning. The LSR-II was also upgraded to enhance its capabilities, by adding a green laser (enabling most efficient excitation of PE and PE tandems), replacing the red laser to increase laser power, and increasing the number of fluorescence detectors. This instrument also runs the current instrument control software with auto-calibration each morning. This auto-calibration software on the Aria and LSR tracks the sensitivity in each channel and so provides added quality assurance, flagging potential problems before they progress to failure. The in-house developed data management system, Flow-LIMS, that allows all Cancer Center members to annotate, access, and manage flow cytometry data on computers in their local laboratories, has been designed to facilitate sharing of experimental data, and it helps to foster collaboration between laboratories. The CSF is advised by a dedicated Facility Advisory Committee (FAC) that meets annually. FAC recommendations are directed to a governing Facility Parent Oversight Committee, that serves to ensure that the CSF continues to have the necesssary resources to provide the highest quality services.