GLOBAL ANALYSIS OF UNMATCHED POLARIZED FLUORESCENCE DECAY CURVES
GLOBAL ANALYSIS OF UNMATCHED POLARIZED FLUORESCENCE DECAY CURVES
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DOI：
10.1021/j100133a005
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发表时间：
1993-08-05
影响因子：
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通讯作者：
DESCHRYVER, FC
中科院分区：
文献类型：
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作者：
CRUTZEN, M;AMELOOT, M;DESCHRYVER, FC
系统地研究了在长时间零极限各向异性的非结合（同质）和结合（异质）模型情况下，在不同实验条件下（包括参考卷积）收集的非匹配偏振荧光衰减曲线的同时分析性能。基于观测曲线和拟合曲线下面积等约束，采用一种改进的快速收敛算法进行了研究。讨论了改变实验条件的效率，如发射偏振器的方向、发射/激发波长和定时校准，以及参数恢复和区分荧光各向异性的可选模型的能力。当总荧光和各向异性的衰减为单指数衰减时，当旋转弛豫时间与荧光弛豫时间相差不大且大于通道的定时校准时，在发射偏振器的两个极端方向上收集的衰减迹的组合足以用于参数恢复。对于具有多指数r(t)的单一复合系统，建议使用不同的激发波长。当各向异性衰减中的弛豫时间差异很大时，必须采用不同的定时校准。对于混合物，使用不同的激发/发射波长可获得最佳结果。作为后者的一个例子，间鲁精和甲酚紫的混合物在20℃的1-丙醇中进行了参考卷积分析。
The performance of the simultaneous analysis of unmatched polarized fluorescence decay curves collected under various experimental conditions, including reference convolution, is investigated systematically for a number of nonassociative (homogeneous) and associative (heterogeneous) model cases with zero limiting anisotropy at long times. The study has been carried out using an improved and rapidly converging algorithm based on the constraint for equal area under the observed and fitted curves. The efficiency of changing experimental conditions such as the orientation of the emission polarizer, the emission/excitation wavelength, and the timing calibration is discussed with respect to the parameter recovery and the power to discriminate between alternative models for the fluorescence anisotropy. When the decay of the total fluorescence and the anisotropy are monoexponential, the combination of decay traces collected at the two extreme orientations of the emission polarizer is sufficient for the parameter recovery when the rotational relaxation time is not too much different from the fluorescence relaxation time and is larger than the timing calibration of the channels. For a single compound system with a multiexponential r(t) the use of different excitation wavelengths is recommended. Different timing calibrations have to be utilized when the relaxation times in the anisotropy decay are widely different. For a mixture of compounds the best results are obtained by using different excitation/emission wavelengths. As an example of the latter a mixture of resorufine and cresyl violet in 1-propanol at 20-degrees-C has been analyzed using reference convolution.