ANALYSIS OF FLUORESCENCE DECAY KINETICS BY METHOD OF LEAST-SQUARES
ANALYSIS OF FLUORESCENCE DECAY KINETICS BY METHOD OF LEAST-SQUARES
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DOI：
10.1016/0003-2697(74)90312-1
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发表时间：
1974-01-01
影响因子：
2.9
通讯作者：
STEINBER.IZ
中科院分区：
文献类型：
--
作者：
GRINVALD, A;STEINBER.IZ
荧光衰变动力学分析的目的是确定描述衰变过程的相关参数的解析表达式和数值。在众所周知的最小二乘方法中，人们为衰减数据假设一种合理的函数形式，并调整参数的值，直到在数据和计算的衰减函数之间获得统计上的最佳拟合，即直到残差的加权平方和最小。结果表明，残差平方的适当加权可以显着提高分析质量。这种加权需要有关实验噪声特征的信息，这些信息通常是可用的，例如，当噪声是由于光子计数技术中的计数误差引起时。此外，通过使用有关所研究系统的可用辅助信息，通常可以实现分析准确性的显着提高。例如，发色团混合物（例如蛋白质分子中的色氨酸残基）的多指数荧光衰减中的前指数有时可以独立估计；通过分析衰变动力学，可以获得更高的衰变寿命精度。提出加权残差的自相关函数的形状可以作为实验数据与分析得到的衰减函数之间的拟合质量的便捷标准。上述结论是通过计算机模拟实验分析得出的，并说明了该方法的实用性。强调了说明衰变动力学分析中固有的估计参数的不确定性的重要性。
Analysis of fluorescence decay kinetics aims at the determination of the analytic expression and the numerical values of the pertinent parameters which describe the decay process. In the well-known method of least-squares, one assumes a plausible functional form for the decay data and adjusts the values of the parameters until the statistically best fit is obtained between the data and the calculated decay function, i.e., until the sum of the weighted squares of the residuals is at a minimum. It is shown that proper weighting of the squares of the residuals may markedly improve the quality of the analysis. Such weighting requires information about the character of the experimental noise, which is often available, e.g., when the noise is due to counting error in photon-counting techniques. Furthermore, dramatic improvements in the accuracy of the analysis may often be achieved by use of auxiliary information available about the system studied. For example, the preexponents in a multiexponential fluorescence decay of a mixture of chromophores (such as tryptophan residues in a protein molecule) may sometimes be estimated independently; much higher accuracy can then be attained for the decay lifetimes by analysis of the decay kinetics. It is proposed that the shape of the autocorrelation function of the weighted residuals may serve as a convenient criterion for the quality of fit between the experimental data and the decay function obtained by analysis. The above conclusions were reached by analysis of computer-simulated experiments, and the usefulness of this approach is illustrated. The importance of stating the uncertainties in the estimated parameters inherent in the analysis of decay kinetics is stressed.