University of Delaware, Dept. of Chemistry and Biochemistry, Brown Laboratory, 163 The Green, Newark, Delaware 19716, USA.
Appl Spectrosc. 2010 Oct;64(10):1145-53. doi: 10.1366/000370210792973479.
Fluorescence correlation spectroscopy (FCS) uses fluctuations in the fluorescence collected from a small illuminated volume to measure dynamic processes of fluorophores. In traditional FCS, spectral overlap produces cross-talk in dedicated detector channels, undermining the accuracy of measurements of molecular interactions. Here, the experimental realization of full-spectrum fluorescence correlation spectroscopy is described and coupled with multivariate data analysis to numerically correct detector cross-talk, isolating spectra and fluctuation traces of mixture components in spite of overlap. Application of this methodology is illustrated using the measurement of the diffusion constant of labeled polystyrene in hydroxypropyl cellulose in the presence of a persistent dye. Additionally, the results show that full-spectrum FCS with multivariate analysis can isolate and characterize signals from unanticipated sample components.
荧光相关光谱学（FCS）利用从小体积照明区域收集的荧光强度波动来测量荧光团的动态过程。在传统的 FCS 中，光谱重叠会在专用检测器通道中产生串扰，从而降低分子相互作用测量的准确性。本文描述了全谱荧光相关光谱学的实验实现，并与多元数据分析相结合，以数值方式校正检测器串扰，从而在存在重叠的情况下分离混合物成分的光谱和波动轨迹。该方法的应用通过标记的聚苯乙烯在羟丙基纤维素中扩散常数的测量来说明，同时存在持久染料。此外，结果表明，具有多元分析的全谱 FCS 可以分离和表征意外样品成分的信号。
