Investigation of Synchronous Fluorescence Method in Multicomponent Analysis in Tissue

We investigate the advantages and disadvantages of the synchronous fluorescence (SF) scan method as compared with the fixed-excitation or fixed-emission fluorescence methods in analyzing biological tissue samples or tissue-like synthetic phantoms by quantitatively comparing the accuracy of multicomponent analysis in nonscattering phantoms and qualitatively comparing the spectral profiles of fluorescence spectra measured from tissue-like phantoms and ex vivo rat brains. The results of experiments in nonscattering phantoms indicate that the synchronous scan method with an appropriate wavelength interval (Δλ, i.e., the difference between the excitation wavelength and emission wavelength) always yields the highest overall quantification accuracy in a single scan. The results of experiments in tissue-like phantoms and ex vivo rat brains suggest that the synchronous scan effectively examines the fluorescence peaks of multiple fluorophores in one single scan, reduce the overlap between fluorescence peaks of different endogenous fluorophores, and express/suppress the effect of specific absorption peaks on the shapes of fluorescence spectra. These features greatly facilitate the qualitative interpretation of fluorescence spectra from tissue samples or tissue-like phantoms. Finally, it is worth pointing out that background signals could exert a greater impact on the spectral shape for small wavelength intervals (Δλ <; 40 nm) compared to excitation or emission scans.