PROBES FOR FLOW CYTOMETRY

这一计划的主要长期目标是继续发展 新的细胞化学、生物物理和分子探针和技术 测量个体的各种成分的含量和/或构象 细胞。这些探针(技术)将被设计用于细胞分析，通过 流式细胞术和细胞分选，有望在临床上 在癌症筛查、诊断、分类和预后方面的应用 在评估肿瘤药物敏感性和监测治疗中的作用 效果。本研究目前的具体目标(项目)为：1) 对染色质结构差异可能影响的评估 结肠和乳腺肿瘤的DNA含量估计；2)RNA含量估计： A)发展灵敏的核糖核酸测量方法(S)，适用于 从实体肿瘤中分离出的细胞核；b)对导致 核糖体与多聚核糖体rRNA原位差异染色 派罗宁Y，以及这一现象在一种新淋巴细胞中的应用 (3)共振能量的研究与应用 派罗宁Y与罗丹明123之间作为新的线粒体探针的转移； (4)核内DNA原位变性检测方法的推广 分离自结肠、乳腺、肺和膀胱肿瘤；(5)分析 抗肿瘤药物选择性释放具有亲和力的核蛋白 核酸，以及针对这些蛋白质开发的测试抗体 可能探索药物对靶细胞的影响。 第二个目标(具体目标或项目6)是应用开发的 探索细胞生长，有丝分裂周期的进展， 细胞对不同抗肿瘤药物的分化和反应。这个 数据将与这些过程的动力学相关联，并将提供 更完整地描述代谢变化和细胞异质性 在细胞周期中。 第三个目标(具体目标或项目7)是通过使用生物物理学 和生化方法，以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。因为很多人 探针要么是抗肿瘤药物本身，要么是药物类似物，这些研究 除了有助于开发新的诊断技术外，还有 有望揭示药物对细胞作用机制的信息 分子水平。将这些数据与对整个细胞的药物研究相关联 将提供关于细胞内药物靶点、机制的全面观点 参与药物结合，以及与细胞动力学或细胞动力学有关的细胞毒性 新陈代谢状态。这反过来将有助于评估蜂窝功能 预测对特定药物的敏感性，并帮助新药设计。

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.