CORE--MOLECULAR ANALYSIS

Core C的目的是为本项目的项目研究者提供分子分析服务。Western免疫印迹、northern和原位杂交，以及[定量的竞争性]聚合酶链反应（RT-PCR）分析；[DNA碎片;电动势位移试验；乙酰胆碱（ACh）相关酶和ICE酶测定；细胞因子生物活性分析（细胞因子周期蛋白）现在是可用的。此外，其他蛋白质及其编码mrna的分析可能感兴趣（例如，其他细胞因子，生长因子或结构蛋白）将根据要求提供。目前已有S100beta和GFAP酶联免疫吸附测定试剂盒，其他酶联免疫吸附测定试剂盒可用于快速筛选大量样品。作为细胞培养中特定mrna的快速筛选，微原位杂交也是可行的。[待分析的组织将来自人类和啮齿动物的大脑（项目1-4），以及细胞培养（项目3）。]本核心中进行的分析数据将作为原始数据和初步分析数据提供给项目调查员。例如，来自北部和西部免疫印迹分析和RT- PCR的数据将作为原始数据（凝胶图片和转移片）和提取物的ng/ ml返回给研究者，相对于总蛋白、总RNA、G3PDH或杂交核糖体RNA。elisa的数据将以每个样品光密度单位的形式作为原始数据返回，同时生成标准曲线，并计算ng/mug蛋白和ng/mul提取物。原位杂交后，切片或细胞培养物将返回给研究者，或者，如果需要，发送给[项目4]进行图像分析。培养物微原位杂交的数据将以cpm/well（相对于总聚腺苷化mRNA）的形式返回给研究者。[在本核心中执行的服务的验证和价值将存档在本核心的独立计算机数据库中。]

The purpose of Core C is to provide molecular analysis services to the Project investigators in this Program Project. Western immunoblot, northern and in situ hybridization, and [quantitative, competitive] polymerase chain reaction (RT-PCR) analyses; [DNA fragmentation; electromotive shift assay; enzyme assays for acetylcholine (ACh)-related enzymes and for ICE; and cytokine bioactivity analysis for cytokine cycle proteins] are now available. In addition, analysis of other proteins and their encoding mRNAs that may be of interest (e.g., other cytokines, growth factors, or structural proteins) will be available upon request. S100beta and GFAP ELISAs are available, and other ELISAs can be constructed for rapid screening of large numbers of samples. As a rapid screen for specific mRNAs in cell cultures, micro-in situ hybridization is also available. [The tissue to be analyzed will from human and rodent brain (Projects 1-4), and from cell cultures (Project 3).] Data from analyses performed in this Core will be provided to the Project Investigators as both raw data and preliminary analyzed data. For example, data from northern and western immunoblot analyses and RT- PCR will be returned to the investigator as raw data (pictures of gels and films of transfers) and as ng/mul of extract, relative to total protein, total RNA, G3PDH, or hybridizable ribosomal RNA. Data from ELISAs will be returned as raw data in the form of optical density units per sample, together with a standard curve generated at the same time, and calculation of ng/mug protein and ng/mul extract. Following in situ hybridization, sections or cell cultures will be returned to the investigator or, if requested, sent direction to [project 4] for image analysis. Data from micro-in situ hybridization of cultures will be returned to the investigator as cpm/well relative to total polyadenylated mRNA. [Verification of and values from services performed in this Core will be archived in the free standing computer database in this core.]