NEW FLUORESCENT THIOL PROBES FOR FLOW CYTOMETRY AND PCR

我们建议合成新的探针用于细胞内的测定。 谷胱甘肽和其他细胞硫醇。在第一阶段，我们合成了五个新的 优于现有硫醇染料CPM的硫醇活性荧光团 关于量子产额、斯托克斯位移和灵敏度的辅助DACM 用于硫醇定量。至少有一种染料比现有的要好 胞内硫醇染料的选择性和敏感度 谷胱甘肽在其他细胞内硫醇存在的情况下。在第二阶段，我们 计划制备更多具有相同基本分子结构的染料 但带有取代基会提供更长的波长吸收和 发射，在水介质中有更好的溶解性。合成的探针 将在体外和细胞培养中测试吸收和排放 与谷胱甘肽和其他蛋白质硫醇结合时的性质。细胞 这些染料的毒性和是否适合用于流式细胞术也将是 下定决心。新的染料也将与现有的染料进行比较，如 荧光标记在其他应用中的应用，如高效液相色谱检测 硫醇、酶分析和聚合酶链式反应技术。

We propose to synthesize new probes for the determination of intracellular glutathione and other cellular thiols. In Phase I we synthesized five new thiol reactive fluorophores which are superior to existing thiol dyes CPM aid DACM with respect to quantum yield and Stokes shifts and sensitivity for thiol quantitation. At least one of the dyes was better than existing intracellular thiol dyes with respect to selectivity and sensitivity for glutathione in the presence of other intracellular thiols. In Phase II we plan to prepare additional dyes with the same basic molecular structure but with substituents which will impart longer wavelength absorption and emission and better solubility in aqueous media. The synthesized probes will be tested in vitro and in cell cultures for absorption and emission properties when bound to glutathione and other protein thiols. Cell toxicity and suitability of the dyes for flow cytometry will also be determined. The new dyes will also be evaluated against existing dyes as fluorescent markers in other applications such as HPLC detection of thiols, enzyme assays, and in PCR technology.