Calibration of probe volume in fluorescence correlation spectroscopy
Calibration of probe volume in fluorescence correlation spectroscopy
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DOI：
10.1366/000370207781745883
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发表时间：
2007-09-01
影响因子：
3.5
通讯作者：
Geng, M. Lei
中科院分区：
文献类型：
--
作者：
Gao, Yi;Zhong, Zhenming;Geng, M. Lei
在荧光相关光谱（FCS）中，准确评估探针体积是正确解释实验数据和求解适当扩散模型的基础。在确定尺寸参数、光束半径Ω和沿探头体积的光轴的距离沿着l时，拟合收敛性差一直是一个问题。在这项工作中，在校准过程中的拟合的不稳定性进行了研究，通过检查卡（2）表面。我们证明了在w维中chi（2）的最小值对于收敛和发散数据都是定义良好的。拟合的困难来自于l维。由F统计量确定，l的不确定性可能显著大于Ω的不确定性。一个修改后的校准过程中，建议检查两种数据处理方法的基础上，结合几个短的数据集到一个单一的长期运行和平均的相关函数的几个短的数据集。它被发现，通过使用从短的数据集，而不是一个长时间的相关性的几个收敛的相关函数的平均值，更稳定和一致的尺寸参数被提取到定义的探针体积。
In fluorescence correlation spectroscopy (FCS), an accurate evaluation of the probe volume is the basis of correct interpretation of experimental data and solution of an appropriate diffusion model. Poor fitting convergence has been a problem in the determination of the dimensional parameters, the beam radius, omega, and the distance along the optical axis of the probe volume, l. In this work, the instability of fitting during the calibration process is investigated by examining the chi(2) surfaces. We demonstrate that the minimum of chi(2) in the w dimension is well defined for both converging and diverging data. The difficulty of fitting comes from the l dimension. The uncertainty in l could be significantly larger than that in omega, as determined by F-statistics. A modified calibration process is recommended based on examining two data treatment methods, combining several short data sets into a single long run and averaging the correlation functions of several short data sets. It is found that by using the mean of several converging correlation functions from short data sets instead of a long time correlation, more stable and consistent dimensional parameters are extracted to define the probe volume.