A Frequency Domain Fluorescence Lifetime Instrument for the Aladdin Synchrotron Light Source

这项建议描述了在威斯康星大学麦迪逊分校阿拉丁同步加速器的研究资源的建设，致力于生物系统的时间分辨荧光光谱学。其具体目标是(1)建立一个利用阿拉丁同步加速器光源的时间结构，特别是其皮秒分辨率和宽激发波长能力的频域荧光计；(2)与荧光设施一起建立一个制备生物样品实验室；(3)培训学生使用同步辐射的频域方法的原理，并操作该设施；以及(4)对生物系统动力学进行核心研究。为了实现这些目标，我们将借鉴我们以前的经验(1)在意大利的Frascati建立一个同步辐射设施，(2)开发利用脉冲源的谐波含量的频域荧光仪器，(3)设计和建造一个微波超外差荧光仪，以及(4)建立和运营一个国际发光中心(荧光动力学实验室)，为生物科学家提供一个最先进的荧光实验室和一个功能性生物化学实验室。荧光时间分辨实验在探索蛋白质和核酸生物聚合物或细胞细胞器等生物系统的动力学方面具有不可估量的价值。阿拉丁同步加速器源在时间分辨率、亮度、连续激发波长(紫外线和可见光)、相位稳定性和脉冲间幅度稳定性方面的独特性，使其成为美国同步辐射实验室的最佳选择。经过前期建设，荧光同步辐射实验室将成为生命科学界的研究资源。阿拉丁的初步实验充分证明了多束团模式下频域测量的可行性，并证实了我们在信号稳定性和信号强度方面的预测，这是研究复杂生物系统的关键特征。

This proposal describes the construction of a research resource at the Aladdin Synchrotron, University of Wisconsin- Madison, dedicated to time-resolved fluorescence spectroscopy of biological systems. The specific aims are to (1) build a frequency domain fluorometer that exploits the time structure of the Aladdin Synchrotron light source especially its picosecond resolution and broad excitation wavelength capabilities; (2) establish a preparative biological sample laboratory in conjunction with the fluorescence facility; (3) train students in the principles of frequency domain methods using synchrotron radiation, and to operate the facility; and (4) conduct core research on the dynamics of biological systems. To accomplish these goals, we will draw upon our prior experiences (1) establishment of a synchrotron radiation facility at Frascati (Italy), (2) development of frequency domain fluorescence instrumentation exploiting the harmonic content of pulsed sources, (3) design and construction of a microwave superheterodyning fluorometer and (4) establishment and operation of an international luminescence center (Laboratory for Fluorescence Dynamics), which provides biological scientists with a state-of-the-art fluorescence laboratory coupled with a functional biochemistry laboratory. Fluorescence time-resolved experiments are invaluable in exploring the dynamics of biological systems such as protein and nucleic acid biopolymers, or cellular organelles.. The uniqueness of the Aladdin Synchrotron source in terms of time resolution, brightness, continuous excitation wavelengths (ultraviolet and visible), phase stability, and pulse-to-pulse amplitude stability, makes it the best choice in the United State for a synchrotron radiation laboratory. After the initial construction period, the Fluorescence Synchrotron Radiation Laboratory will become a research resource for the life sciences community. Preliminary experiments at Aladdin have fully demonstrated the feas ibility of frequency domain measurements in the multibunch mode and have confirmed our predictions in terms of signal stability and signal intensity which are crucial characteristics for the study of complex biological systems.