Biochemical Core

生化核心

• 批准号: 7312498
• 负责人: Richard J. Roman
• 金额: $ 30.55万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7312498
• 关键词: analytical chemistry; biomedical facility; cytochrome P450; eicosanoids; enzyme activity; high performance liquid chromatography; immunocytochemistry; in situ hybridization; liquid chromatography mass spectrometry; polymerase chain reaction; radioimmunoassay; western blottings

The Biochemical Core Laboratory of this Program Project Grant will provide a consolidated highly specialized well-equipped and professional unit capable of performing a variety of radioimmunoassay, biochemical and molecular procedures as required by project investigators. No single investigator laboratory supported by R01 would be capable of equipping and staffing this resource. The Core will carry out all the analytical techniques: 1) Assay of CYP and NOS activities by radiochemical HPLC. 2) Measurement of 20- HETE and EETs in tissue samples, CSF and microdialysates by LC/MS. 3) Identification of novel metabolites generated by CYP4F and CYP4X enzymes by LC?MS/MS. This Core is also equipped to measure IP3, DAG, microglial and astroglial products in the brain and cerebral vasculature. In addition we will measure CYP and NOS protein using western blots and immunohistochemistry. Molecular measurements with real time RT-PCR and in situ hybridizations will be carried out as outlined in the projects. Access to the analytical expertise and equipment in the central facility is essential to each of the proposed projects and justified by the large number of biochemical and molecular determinations required for successful completion of the proposed studies.

本项目的生化核心实验室项目资助将提供综合的高度 设备精良的专业单位，能够按照项目研究人员的要求进行各种放射免疫分析、生化和分子程序。 R01 支持的任何一个研究实验室都没有能力为这一资源配备设备和人员。核心将执行所有分析技术：1) 通过放射化学 HPLC 测定 CYP 和 NOS 活性。 2) 通过 LC/MS 测量组织样本、CSF 和微透析液中的 20-HETE 和 EET。 3) 通过 LC?MS/MS 鉴定 CYP4F 和 CYP4X 酶产生的新代谢物。该核心还可以测量大脑和脑血管系统中的 IP3、DAG、小胶质细胞和星形胶质细胞产物。此外，我们将使用蛋白质印迹和免疫组织化学来测量 CYP 和 NOS 蛋白。将按照项目中概述的方式进行实时 RT-PCR 和原位杂交的分子测量。 获得中央设施的分析专业知识和设备对于每个拟议项目至关重要，并通过成功完成拟议研究所需的大量生化和分子测定来证明。