The polarized AB plot for the frequency-domain analysis and representation of fluorophore rotation and resonance energy homotransfer
The polarized AB plot for the frequency-domain analysis and representation of fluorophore rotation and resonance energy homotransfer
复制标题
DOI：
10.1111/j.1365-2818.2008.02102.x
复制
发表时间：
2008-11-01
影响因子：
2
通讯作者：
Clayton, A. H. A.
中科院分区：
文献类型：
--
作者：
Clayton, A. H. A.
单频相位调制荧光寿命成像数据的图形表示，称为AB图，扩展到考虑到荧光的偏振分量的测量。对于受阻旋转体模型（其特征在于单个激发态寿命、单个旋转相关时间以及限制性的初始和最终各向异性），旋转相关时间和激发寿命可以从以下发射分量中的任何两个的AB图确定：平行、垂直、总发射或其组合。一种解决的战略，从测量的总，平行和垂直分量的排放的两个受阻旋转体的混合物的组件受阻旋转和寿命的开发。该分析不需要初始极限各向异性或稳态各向异性或激发态寿命的先验知识。绘制极化AB空间中的非均质系统，以帮助解释频域动态去极化成像显微镜实验。这些图可以用来区分空间依赖的旋转相关时间异质性的异质性限制各向异性。讨论了噪声和孔径退偏的影响。预计极化AB图将提供一个有用的辅助现有的方法可视化和分析所产生的分子动力学和同质能量转移在单频显微镜应用中的动态极化现象。
The graphical representation of single-frequency phase-modulation fluorescence lifetime imaging data, referred to as the AB plot, is extended to take into account measurements of the polarized components of the fluorescence. For a hindered rotator model (characterized with a single excited-state lifetime, a single rotational correlation time and limiting initial and final anisotropies) the rotational correlation time and the excited lifetime can be determined from the AB plot of any two of the following emission components: parallel, perpendicular, total emission or combinations thereof. A strategy for resolving the component hindered rotations and lifetimes for mixtures of two hindered rotators from measurements of the total, parallel and perpendicular components of the emission is developed. The analysis does not require prior knowledge of the initial limiting anisotropy or of the steady-state anisotropy or of the excited state lifetime. Plots in polarized AB space derived for heterogeneous systems are constructed to aid interpretation of frequency-domain dynamic depolarization imaging microscopy experiments. These plots can be used to distinguish spatially dependent rotational correlation time heterogeneity from heterogeneity in limiting anisotropies. The effects of noise and aperture depolarization are discussed. It is anticipated that the polarized AB plot will provide a useful adjunct to existing methods for visualizing and analysing dynamic polarization phenomena arising from molecular dynamics and homo-energy transfer in single-frequency microscopy applications.