ANALYTICAL PROTEIN PROFILING

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 中心，不一定是研究者的机构。 I.描述 蛋白质组学实验室  迈克尔·鲍威尔，领导者 蛋白质组学核心实验室拥有表面增强激光解吸电离（SELDI）质谱仪和Thermo-Finnigan LTQ LC/MS/MS系统。还可获得BioRad自动点选器（Proteome Works）和BioRad凝胶成像仪（Molecular imager FX）。与此软件包捆绑的是最新版本的PDQuest 8.0，用于分析凝胶的表达数据。在2007年，我们还增加了一个生物荧光珠分选仪，它允许分析多重测定，如ELISA从一个单一的，小体积的样品。这些设备使我们能够提供蛋白质鉴定和蛋白质修饰分析等服务。我们还可以通过凝胶和LC分析提供差异蛋白表达分析。 分析化学实验室  丹尼尔冯多伊奇，领导人 MSM ACL配备了两个Agilent（HP）1100系列高效液相色谱系统。超过 在过去的一年里，我们从药理学和毒理学系的Ward Kirlin博士那里获得了第二个安捷伦系统。ACL中的HPLC系统包括：1）配备二极管阵列检测器（DAD）、可变波长检测器（VWD）和1046 A荧光检测器（FD）的HP（Agilent）1100系列HPLC; 2）配备1200系列荧光检测器（FD）和高灵敏度Gilson荧光检测器的HP（Agilent）1100系列HPLC; 3）配备有125 S溶剂模块、508自动进样器和168二极管阵列检测器的Beckman Gold HPLC系统。此外，该系统还配备了一个SS 420 x模拟/数字盒，其上连接有荧光和电化学检测器。这些检测器包括高灵敏度Gilson荧光检测器、Agilent 1049 A电化学检测器（ECD）和配备多个5011型高灵敏度双通道分析池的ESA Coulochem II ECD系统。除HPLC系统外，ACL还配备了一台Spectromax Plus（UV/维斯可变波长微量滴定板读数器，配有Softmax Pro软件，用于进行比色分光光度法和免疫测定）和两台Agilent气相色谱仪 (GC)用于检测药物及其代谢物的系统。GC包括具有火焰离子化检测器（FID）和氮磷检测器（NPD）的Agilent（HP）6890系列毛细管GC和与5972系列质谱仪（MS-EI）耦合的Agilent（HP）5890系列毛细管GC。可以在扫描或选择离子监测（SIM）模式下采集数据。 服务费用： 蛋白质组学实验室/2D凝胶实验室 我们目前有一个收费表的所有服务。例如，我们按 LC/MS/MS运行用于LTQ分析。该费用包括胰蛋白酶消化样品、LC/MS运行和后续数据库检索。包括翻译后修饰在内的数据库搜索会对典型的修饰（如磷酸化、硝化、泛素化等）收取额外费用。其他更复杂的分析则根据所需时间的估计收费。目前，我们通过让用户购买自己的试剂盒，然后为他们提供一个工作站，其中包含分析所需的所有材料，来提供生物分析。我们正在制定用户费用，以支付材料成本和平台校准和验证所需的时间。随着今年核心经理的加入，我们应该能够提高我们的效率 电荷结构 分析化学实验室 分析化学实验室有一个标准的服务收费表。费用结构基于处理样品（提取、衍生化等）所需的时间以及用品和试剂的成本。对于非NIH赞助的项目（MSM和外部机构和公司），评估劳动力成本。此外，由于ACL没有从NIH或MSM获得运营预算，因此向所有项目征收额外费用，以帮助支付设备维护和其他实验室管理费用。成本定期调整，以反映供应商价格的变化。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. I. DESCRIPTION Proteomics Lab  Michael Powell, Leader The Proteomics core lab houses a Surface Enhanced Laser Desorption-Ionization (SELDI) mass spectrometer, and a Thermo-Finnigan LTQ LC/MS/MS system. Also available are a BioRad automated spot picker (Proteome Works) and BioRad gel imager (Molecular imager FX). Bundled with this package is the latest version of PDQuest 8.0 for analysis of expression data from gels. In 2007 we have also added a BioPlex bead sorter, which allows the analysis of multiplex assays such as ELISAs from a single, small volume sample. Together this equipment allows us to offer services such as protein identification and analysis of protein modifications. We can also offer differential protein expression analysis by both gel and LC analysis. Analytical Chemistry Lab  Daniel von Deutsch, Leader The MSM ACL is equipped with two Agilent (HP) 1100 series high HPLC systems. Over that past year, we obtained a second Agilent system from Dr. Ward Kirlin in the Department of Pharmacology and Toxicology. The HPLC systems in the ACL include: 1) an HP (Agilent) 1100 series HPLC equipped with a Diode Array Detector (DAD), a Variable Wavelength Detector (VWD) and a 1046A Fluorescence Detector (FD); 2) an HP (Agilent) 1100 series HPLC equipped with a 1200 series Fluorescence Detector (FD) and a high sensitivity Gilson fluorescence detector; 3) a Beckman Gold HPLC system equipped with a 125S Solvent Module, a 508 autosampler and a 168 diode array detector. In addition, the system is equipped with an SS420x analog/digital box to which fluorescence and electrochemical detectors are attached. These detectors include a high sensitivity Gilson fluorescence detector, an Agilent 1049A Electrochemical Detector (ECD) and an ESA Coulochem II ECD system outfitted with multiple Model 5011 high sensitivity two channel analytical cells. In addition to the HPLC systems, the ACL has a Spectromax Plus is a UV/VIS variable wavelength microtiter plate reader with Softmax Pro software used for performing colorimetric spectrophotometry and immunoassays and two Agilent Gas Chromatographs (GC) systems for the detection of drugs and their metabolites. The GCs include an Agilent (HP) 6890 Series capillary GC with Flame Ionization Detector (FID) and Nitrogen Phosphorus Detector (NPD) and an Agilent (HP) 5890 Series capillary GC coupled with 5972 Series Mass Spectrometer (MS - EI). Data can be acquired in the SCAN or Selected Ion Monitoring (SIM) modes. Service Charges: Proteomics Lab/2D Gel Lab We currently have a fee schedule for all services. For example, we charge by the LC/MS/MS run for analysis on the LTQ. This charge includes a digestion of the sample by trypsin, an LC/MS run, and the subsequent database searches. Database searches including post translational modifications incur an additional charge for typical modifications such as phosphorylation, nitration, ubiquitination, etc. Other, more complicated analysis, are charged from an estimate of the time required. Currently, we are offering BioPlex analysis by having users purchase their own kits and then providing them with a workstation with all of the materials needed for analysis. We are working on a user fee to cover the cost of materials and the time necessary for calibrations and validations for the platform. With the addition of our core manager this year we should be able to increase the efficiency of our charge structure. Analytical Chemistry Lab The Analytical Chemistry Laboratory has a standard fee schedule for services. The fee structure is based on the amount of time required to process the samples (extraction, derivatization, etc) and the cost of supplies and reagents. For non-NIH sponsored projects (MSM and outside institutions and corporations), labor costs are assessed. In addition, since the ACL receives no operating budget from either the NIH or MSM, an additional fee is assessed to all projects to help cover equipment maintenance and other laboratory overhead. Costs are periodically adjusted to reflect vendor price changes.