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TWO DIMENSIONAL ANALYTICAL PREPARATIVE ULTRACENTRIFUGE

二维分析制备型超速离心机

基本信息

批准号：
2332125
负责人：
NORMAN G ANDERSON
金额：
$ 10万
依托单位：
LARGE SCALE BIOLOGY CORPORATION
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-05-01 至 1998-06-15
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2332125
关键词：
analytical ultracentrifugation biomedical equipment development charge coupled device camera computer program /software density gradient ultracentrifugation fiber optics laboratory rat method development sedimentation spectrometry

项目摘要

DESCRIPTION: (Adapted from the applicant's abstract) This proposal is to develop a combined preparative-analytical two-dimensional zonal ultracentrifuge system, based on early prototypes developed at Oak Ridge, to both measure accurately the sedimentation rates (s) and isopycnic banding densities (p) of cells, subcellular organelles and infectious agents, and to perform high resolution preparative separations based on these parameters. The purified fractions will be anaylyzed by high resolution two-dimensional electrophoresis with the objective of assigning to each protein that can be detected a specific subcellular location. Initial demonstration and feasibility studies will be concerned with separating and analyzing different classes of liver and brain nuclei. LSB is currently preparing standard reference maps of human and rodent proteins by organ, is identifying proteins by composition or sequence, and is mapping the effects of drugs, hormones, and toxic agents of gene expression by 2DE. We now require methods for producing subcellular fractions (nuclei, mitochondria, peroxisomes, lysosomes, etc.) reproducibly, and for further subfractionating these into nuclear membranes, nucleoli, nuclear pore complexes, inner and outer mitochondrial membranes, etc., so that the location of each protein gene product can be determined. The 2D centrifuge systems proposes will also be useful for the detection and isolation of infectious agents from tissues and body fluids.

描述：(改编自申请人的摘要)本提案旨在开发一种制备-分析相结合的二维分区基于橡树岭开发的早期原型的超速离心机系统，既要准确测量沉积速率(S)又要准确测量等压细胞、亚细胞器和感染性细胞器的条带密度(P) 试剂，并执行基于以下的高分辨率制备分离这些参数。对提纯后的馏分进行高效液相色谱分析。分辨率双向电泳法的目标是为每个可以检测到的蛋白质分配一个特定的亚细胞地点。初步论证和可行性研究将是涉及分离和分析不同类别的肝脏和大脑核团。LSB目前正在编制标准参考图人类和啮齿动物的蛋白质通过器官识别蛋白质组成或序列，并正在绘制药物、激素、用2DE检测毒物对基因表达的影响。我们现在需要用于产生亚细胞组分(核、线粒体、过氧化物体、溶酶体等)可重复的，并进一步细分这些核膜、核仁、核孔复合体，内、外线粒体膜等，使每个蛋白质基因的位置可以确定产品。所提出的2D离心机系统还将可用于检测和分离来自中国的组织和体液。