PROBES FOR FLOW CYTOMETRY

流式细胞术探针

基本信息

批准号：
3168288
负责人：
ZBIGNIEW DARZYNKIEWICZ
金额：
$ 33.34万
依托单位：
NEW YORK MEDICAL COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
1990
资助国家：
美国
起止时间：
1990-10-01 至 1995-02-28
项目状态：
已结题

项目摘要

The primary long term objective of this program is to continue development of new cytochemical, biophysical and molecular probes and techniques to measure content and/or conformation of various constituents of individual cells. These probes (techniques) will be designed for cell analysis by flow cytometry and cell sorting and are expected to have clinical applications in cancer screening, diagnosis, classification and prognosis, in evaluation of drug sensitivity of tumors and in monitoring treatment effects. Specific current aims (projects) of this study are: 1) evaluation of the possible effect of differences in chromatin structure on DNA content estimates in colon and breast tumors; 2) RNA content estimates: a) development of sensitive method(s) of RNA measurement applicable to nuclei isolated from solid tumors; b) studies of mechanisms responsible for differential staining of rRNA in ribosomes vs polyribosomes in situ with pyronin Y, and application of this phenomenon to a new lymphocyte stimulation assay; (3) studies and application of the resonance energy transfer between pyronin Y and rhodamine 123 as a new mitochondrial probe; (4) extension of the method for assay of DNA denaturation in situ to nuclei isolated from colon, breast, lung and bladder tumors; (5) analysis of nuclear proteins selectively released by antitumor drugs having affinity to nucleic acids, and testing antibodies developed against these proteins as a possible probe of drug effects on target cells. The second objective (specific aim or project 6) is to apply the developed probes to studies of cell growth, progression through the mitotic cycle, differentiation and response of cells to different antitumor drugs. The data will be correlated with kinetics of those processes and will provide a more complete description of metabolic changes and cell heterogeneity during the cell cycle. The third objective (specific aim or project 7) is, by using biophysical and biochemical methods, to study molecular interactions between the probes and various cell constituents, predominantly nucleic acids. Because many probes are either antitumor drugs themselves or drug analogs, these studies in addition to being helpful in developing new diagnostic techniques, are expected to reveal information on mechanisms of drug action on cells at the molecular level. Correlating these data with drug studies on whole cells will provide a comprehensive view on intracellular drug targets, mechanisms involved in drug binding, and cytotoxicity as related to cell kinetics or metabolic state. This in turn will help assess the cellular features predicting susceptibility to particular drugs and aid in new drug design.

该计划的主要长期目标是继续开发新的细胞化学，生物物理和分子探针和技术测量个人各个成分的内容和/或构象细胞。这些探针（技术）将设计用于细胞分析流式细胞仪和细胞分类，预计将具有临床在癌症筛查，诊断，分类和预后中的应用，在评估肿瘤的药物敏感性和监测治疗方面效果。本研究的具体目前目的（项目）是：1）评估染色质结构差异的可能影响结肠和乳腺肿瘤中的DNA含量估计； 2）RNA含量估计： a）开发适用于RNA测量的敏感方法从实体瘤分离的细胞核； b）研究负责的机制核糖体中rRNA的差异染色与原位的多核糖体吡隆蛋白Y，并将这种现象应用于新的淋巴细胞刺激测定；（3）共振能量的研究和应用作为新的线粒体探针，吡咯蛋白Y和Rhodamine 123之间的转移；（4）扩展DNA变性的方法原位对核的延伸从结肠，乳房，肺和膀胱肿瘤中分离出来；（5）分析核蛋白质由具有亲和力的抗肿瘤药物选择性释放核酸和针对这些蛋白的抗体作为一种药物对靶细胞的影响可能探测。第二个目标（特定目标或项目6）是应用开发的研究细胞生长的研究，通过有丝分裂循环进展的探针，细胞对不同抗肿瘤药物的分化和反应。这数据将与这些过程的动力学相关，并将提供代谢变化和细胞异质性的更完整描述在细胞周期中。第三个目标（特定目标或项目7）是使用生物物理和生化方法，研究探针之间的分子相互作用以及各种细胞成分，主要是核酸。因为很多探针本身就是抗肿瘤药物，或者是药物类似物，这些研究除了有助于开发新的诊断技术之外预计将揭示有关药物作用机制对细胞的机制的信息分子水平。将这些数据与全细胞的药物研究相关联将提供有关细胞内药物目标的全面观点参与药物结合和与细胞动力学有关的细胞毒性或代谢状态。反过来，这将有助于评估细胞特征预测对特定药物的敏感性并有助于新药设计。