PHOTON FLUORESCENCE SPECT & DYNAMICS OF ENHANCED GREEN FLUORESCENT PROTEIN

单光子荧光相关光谱（FCS）已被 进行了一些荧光染料的光漂白动力学 特性进行了实验研究。 光漂白 量子产率（？B）罗丹明绿色、cy 2和荧光素， 单光子激发（100-fs脉冲和CW），已被表征。 虽然罗丹明绿色在脉冲激发下不漂白， 荧光素光漂白（？B=8.4x10-6和1.0x10-5） 分别在CW激励和脉冲激励之后。 类似 已经对Cy 3，别藻蓝蛋白， B-藻红蛋白、R-藻红蛋白、R-藻蓝蛋白和玫瑰红， CW单光子FCS 比较之间的测量？B in 这份工作和其他任何可能的。 例如，？B~7.0x10-5 R-藻红蛋白与Whiet和Stryer（1998）的结果一致 使用不同的技术（1.1x10-5）。 虽然Cy 3的漂白 不依赖于单线态氧，没有观察到光漂白 在脱氧的玫瑰红中， 态和单线态氧的光漂白动力学。 初步 突变型绿色荧光蛋白（EGFP）的测量作为一个函数 激发波长（405和488 nm）以及pH值（5.8和 11.0）的缓冲液进行。 三重态寿命 (? T）和这些荧光团的系统间交叉率（kISC 还使用强度依赖性FCS结果进行了分析。

One-photon fluorescence correlation spectroscopy (FCS) has been conducted on a number of fluorescent dyes for photobleaching dynamics characterization has been studied experimentally. Photobleaching quantum yield (?b) of rhodamine green, cy2, and fluorescein, following one-photon excitation ( 100-fs pulses and CW), has been characterized. While rhodamine green did not bleach under pulsed excitation, fluorescein underwent photobleaching (?b=8.4x10-6 and 1.0x10-5) following the CW- and pulsed-excitation; respectively. Similar measurements has been carried out on cy3, allophycocyanin, B-phycoerythrin, R-phycoerythrin, R-phycocyanin and rose bengal using CW-one-photon FCS. Comparison was made between the measured ?b in this work and others whenever possible. For example, ?b~7.0x10-5 of R-phycoerythrin is in a good agreement with Whiet and Stryer (1998) using a different technique (1.1x10-5). While the bleaching of cy3 was independent of the singlet oxygen, no photobleaching was obser ved in de-oxygenated rose bengal suggesting the involvement of triplet state and singlet oxygen in photobleaching dynamics. Preliminary measurements on mutant green fluorescent protein (EGFP) as a function of excitation wavelength (405 and 488 nm) as well as pH-value (5.8 and 11.0) of the buffer were also carried out. The triplet state lifetime (?T) and the intersystem crossing rate (kISC) of those fluorophores were also analyzed using the intensity-dependent FCS results.