Protein Translational Mobility and Concentration Near Membrane Surfaces

全内反射与荧光相关光谱学将被用来确定控制在透明平面底物上沉积的单磷脂双层附近的荧光蛋白的平移迁移率和浓度的因素。氩离子激光束将在衬底/溶液界面处进行内部反射，产生一个仅延伸到邻近溶液一小段距离的倏逝场。随着消失的深度，在光学显微镜的中间成像平面上放置的孔径将在膜表面附近定义一个小体积。这个小体积内的分子产生的荧光，用高孔径物镜收集，用快速硅雪崩光电二极管检测，随着单个荧光蛋白进出观察体积，荧光会随时间波动。标准化的荧光波动自相关函数将提供有关溶质平移迁移率和浓度的信息。多种多样的生物过程涉及溶液中的分子（如蛋白质）与生物细胞表面的受体分子的相互作用。这些相互作用的细节导致了特定的生物行为，据预测，这些相互作用的细节在很大程度上取决于直接靠近细胞表面的溶液中蛋白质的浓度和运动。这两种性质，浓度和平移迁移率，将通过使用激光激发和光学显微镜新开发的方法，在透明平面上构建的细胞表面模型旁边测量荧光蛋白。这些结果将用于扩大目前关于细胞如何检测和响应其环境的一般知识水平，从而产生生物功能。

9728116 Thompson Total internal reflection with fluorescence correlation spectroscopy will be used to determine the factors which govern the translational mobilities and concentrations of fluorescent proteins adjacent to single phospholipid bilayers deposited on transparent planar substrates. An argon ion laser beam will be internally reflected at the substrate/solution interface, creating an evanescent field that extends only a small distance into the adjacent solution. Along with the evanescent depth, an aperture placed at an intermediate image plane of an optical microscope will define a small volume next to the membrane surface. The fluorescence arising from molecules within this small volume, collected with a high aperture objective and detected with a fast silicon avalanche photodiode, will fluctuate with time as individual fluorescent proteins move in and out of the observed volume. The normalized fluorescence fluctuation autocorrelation function will provide information about solute translational mobility and concentration. A large variety of biological processes involve the interactions of molecules in solution, such as proteins, with receptor molecules present on the surfaces of biological cells. The details of these interactions, which result in specific biological behavior, are predicted to depend strongly on the concentrations and movements of the proteins in solution directly adjacent to the cell surfaces. These two properties, concentration and translational mobility, will be measured for fluorescent proteins next to models of cell surfaces constructed on transparent planar surfaces, by using newly developed methods that employ laser excitation and optical microscopy. The results will be used to expand the current level of general knowledge about how cells detect and respond to their environments, giving rise to biological function.