Photo-activatable fluorophores and techniques for biological imaging applications

用于生物成像应用的光激活荧光团和技术

• 批准号: 7237988
• 负责人: Wen-hong Li
• 金额: $ 27.44万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-01 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7237988
• 关键词: Abnormal Cell; Acinus organ component; Amines; Animal Model; Arrhythmia; Biological; Biological Assay; Biological Models; Caenorhabditis elegans; Cataract; Cell Communication; Cell Lineage; Cells; Cellular biology; Class; Color; Communication; Coumarins; Coupling; Cultured Cells; Data Set; Demyelinations; Development; Developmental Biology; Dextrans; Dimensions; Disease; Dreams; Dyes; Embryo; Exocytosis; Fluorescence Microscopy; Four-dimensional; Gap Junctions; Goals; Homeostasis; Image; Imaging Techniques; Invasive; Laser Scanning Microscopy; Life; Light; Measures; Methods; Molecular; Molecular Probe Techniques; Monitor; Movement; Mus; Nematoda; Neurons; Numbers; Optics; Organic Synthesis; Organism; Pancreas; Parents; Photochemistry; Physiological; Population; Preparation; Property; Protocols documentation; Reporting; Research; Research Personnel; Role; Shapes; Spectrum Analysis; System; Techniques; Viola; Water; alpha benzopyrone; biomaterial compatibility; cellular imaging; concept; deafness; design; dextran; fluorophore; gap junction channel; image processing; improved; intercellular communication; molecular dynamics; novel; optical imaging; photolysis; research study; size; spatiotemporal; tool; two-photon

DESCRIPTION (provided by applicant): Photo-activatable fluorophores (caged dyes) have wide applications in tracking the spatiotemporal dynamics of molecular movements in biological systems. Previously developed caged fluorophores were designed for photolysis using ultra-violet (UV) light. To exploit the excellent three-dimensional (3D) selectivity of two photon (2P) excitation, new caged fluorophores and techniques suitable for 2P uncaging and imaging need to be developed. The long-term objective of this project is to develop caged dyes and techniques for biological imaging applications. The proposed project will focus on developing new methods to track molecular movements between cells through gap junction (GJ) channels. Using photo-activatable fluorophores to image cell-cell communication represents a new application of caged fluorophores. We aim to achieve three goals: first, to develop a 2P uncaging and imaging technique using cell permeable and caged fluorophores to study the dynamics of cell coupling in physiological preparations consisting of intact cell populations in three dimension; second, to develop 2P activatable, water soluble, and amine reactive fluorophores for bioconjugations. We plan to prepare two classes of bioconjugates (Type 1 and Type 2) that have distinct biological applications; third, to apply these bioconjugates of caged fluorophores to develop 2P uncaging and imaging techniques to trace cell lineage and to monitor GJ intercellular communication (GJIC) in living model organisms such as the nematode C.elegans. This organism is ideal for optical imaging because of its transparency and small size. The proposed research will combine organic synthesis, photochemistry, cell biology, and two photon laser scanning microscopy (2PLSM) to achieve these goals. Although the biological focus of this developmental project is on GJIC, concepts, techniques, and tools developed here should be of general utilities to study both intra- and intercellular molecular dynamics in a variety of biological systems. Abnormal cell-cell communication has been implicated in a number of diseases including cardiac arrhythmia, deafness, neuronal demyelination, and cataracts, developing new techniques to study cell communication is important for devising new strategies for treating these diseases.

描述（由申请人提供）：光激活荧光团（笼状染料）在跟踪生物系统中分子运动的时空动态方面具有广泛的应用。先前开发的笼式荧光团设计用于使用紫外线 (UV) 光解。为了利用双光子 (2P) 激发的优异三维 (3D) 选择性，需要开发新的笼式荧光团和适用于 2P 解笼和成像的技术。该项目的长期目标是开发用于生物成像应用的笼式染料和技术。拟议的项目将重点开发新方法来通过间隙连接（GJ）通道追踪细胞之间的分子运动。使用光激活荧光团对细胞间通讯进行成像代表了笼式荧光团的新应用。我们的目标是实现三个目标：首先，开发一种使用细胞可渗透和笼状荧光团的2P解笼和成像技术，以研究由三维完整细胞群组成的生理制剂中细胞耦合的动态；其次，开发用于生物共轭的 2P 可激活、水溶性和胺反应性荧光团。我们计划制备两类具有不同生物学应用的生物结合物（1 型和 2 型）；第三，应用这些笼状荧光团的生物共轭物来开发 2P 解笼和成像技术，以追踪细胞谱系并监测线虫等活体模型生物中的 GJ 细胞间通讯 (GJIC)。这种生物体因其透明且尺寸小而非常适合光学成像。拟议的研究将结合有机合成、光化学、细胞生物学和双光子激光扫描显微镜（2PLSM）来实现这些目标。尽管该开发项目的生物学重点是 GJIC，但这里开发的概念、技术和工具应该具有通用性，可用于研究各种生物系统中的细胞内和细胞间分子动力学。异常的细胞间通讯与许多疾病有关，包括心律失常、耳聋、神经元脱髓鞘和白内障，开发研究细胞通讯的新技术对于设计治疗这些疾病的新策略非常重要。