Fluorescence measurement by a streak camera in a single-photon-counting mode

Abstract

We describe here a recently developed fluorescence measurement system that uses a streak camera to detect fluorescence decay in a single photon-counting mode. This system allows for easy measurements of various samples and provides 2D images of fluorescence in the wavelength and time domains. The great advantage of the system is that the data can be handled with ease; furthermore, the data are amenable to detailed analysis. We describe the picosecond kinetics of fluorescence in spinach Photosystem (PS) II particles at 4-77 K as a typical experimental example. Through the global analysis of the data, we have identified a new fluorescence band (F689) in addition to the already established F680, F685, and F695 emission bands. The blue shift of the steady-state fluorescence spectrum upon cooling below 77 K can be interpreted as an increase of the shorter-wavelength fluorescence, especially F689, due to the slowdown of the excitation energy transfer process. The F685 and F695 bands seem to be thermally equilibrated at 77 K but not at 4 K. The simple and efficient photon accumulation feature of the system allows us to measure fluorescence from leaves, solutions, single colonies, and even single cells. The 2D fluorescence images obtained by this system are presented for isolated spinach PS II particles, intact leaves of Arabidopsis thaliana, the PS I super-complex of a marine centric diatom, Chaetoceros gracilis, isolated membranes of a purple photosynthetic bacterium, Acidiphilium rubrum, which contains Zn-BChl a, and a coral that contains a green fluorescent protein and an algal endosymbiont, Zooxanthella.