Ramamurthy Kannan, Ponnusamy Karunakaran, Chellappan Selvaraju
National Centre for Ultrafast Processes, University of Madras Chennai 600 113 Tamil Nadu India
RSC Adv. 2020 Jan 3;10(2):998-1006. doi: 10.1039/c9ra10154c. eCollection 2020 Jan 2.
Excitation-resolved area-normalized emission spectroscopy (ERANES) is proposed as a new steady-state fluorescence technique for the investigation of heterogeneous fluorescence (HGF) from a mixture of fluorophores and fluorophores present in various environments and proteins. The presence of a single isoemissive point was used to confirm the presence of two absorbing and emitting species in the system. The isoemissive point was found to occur at the wavelength where the ratio of wavelength dependent fluorescence quantum yield of the emissive species equals to the ratio of their total fluorescence quantum yield. The application of the ERANES method for resolving HGF from a mixture of fluorophores having similar or different fluorescence lifetimes with a relatively high degree of fluorescence spectral overlap was demonstrated. When compared to excitation fluorescence (EF) matrix and time-resolved methods, ERANES was found to be a simple analytical method for analyzing HGF from a mixture of fluorophores, and from fluorophores present in heterogeneous media, such as cells, membranes, , and for analyzing protein fluorescence, without the requirement for sophisticated instrumentation and data analysis.
激发分辨面积归一化发射光谱法（ERANES）被提议作为一种新的稳态荧光技术，用于研究来自荧光团混合物以及存在于各种环境中的荧光团和蛋白质的非均相荧光（HGF）。利用单一等发射点的存在来确认系统中存在两种吸收和发射物种。发现等发射点出现在发射物种的波长依赖性荧光量子产率之比等于其总荧光量子产率之比的波长处。证明了ERANES方法在解析具有相似或不同荧光寿命且荧光光谱重叠程度较高的荧光团混合物中的HGF方面的应用。与激发荧光（EF）矩阵法和时间分辨方法相比，发现ERANES是一种用于分析荧光团混合物、存在于非均相介质（如细胞、膜等）中的荧光团以及分析蛋白质荧光的简单分析方法，无需复杂的仪器和数据分析。
