Visser A J, van Hoek A
J Biochem Biophys Methods. 1979 Aug;1(4):195-208. doi: 10.1016/0165-022x(79)90006-x.
A system is described consisting of a mode-locked Ar ion laser and time-resolved photon-counting electronics. The system is capable of measuring fluorescence lifetimes in the subnanosecond time domain. The Ar ion laser is suitable for the excitation of flavins, since the available laser wavelengths encompass the first absorption band of the yellow chromophore. Due to the high radiation density and the short pulse, both the time and wavelength resolution of the fluorescence of very weakly emitting compounds can be measured. Experiments have been described for flavin models exhibiting single and multiple modes of decay. In these examples lifetimes were determined both from deconvolved decay curves and from direct analysis of the tail of the curve, where no interference of the exciting pulse is encountered. Both determinations showed very good agreement. Due to the highly polarized laser light the decay of the emission anisotropy could be measured directly after the exciting pulse. In principle, fast rotational motions might be detected. An anisotropy measurement conducted with a flavoprotein with a noncovalently attached FAD is presented.
描述了一种由锁模氩离子激光器和时间分辨光子计数电子设备组成的系统。该系统能够在亚纳秒时域内测量荧光寿命。氩离子激光器适用于黄素的激发，因为可用的激光波长涵盖了黄色发色团的第一吸收带。由于高辐射密度和短脉冲，可以测量非常弱发射化合物荧光的时间和波长分辨率。已经描述了针对呈现单重和多重衰减模式的黄素模型的实验。在这些示例中，寿命既可以从去卷积衰减曲线确定，也可以从曲线尾部的直接分析确定，在曲线尾部不会遇到激发脉冲的干扰。两种测定结果显示出非常好的一致性。由于激光具有高度偏振性，发射各向异性的衰减可以在激发脉冲之后直接测量。原则上，可以检测到快速旋转运动。本文展示了对一种具有非共价连接FAD的黄素蛋白进行的各向异性测量。
