Pulse fluorometry using simultaneous acquisition of fluorescence and excitation

We report a new method of measuring fluorescence lifetimes which uses the single-photon technique and two detection channels with matched impulse response for simultaneous acquisition of fluorescence and excitation (SAFE). This differential arrangement is shown to correct automatically for variations in the optical pulse profile during the measurement, thus eliminating a common source of error. It can be used to improve precision and sensitivity with any pulsed light source such as a flashlamp, laser, or synchrotron. A routing system separates photomultiplier coincidences from the dual detection channels into different memory segments of a multichannel analyzer (MCA) using a single time-to-amplitude converter (TAC). Comprehensive data are presented on tuning the single-photon response of the Philips range of XP2020Q photomultipliers. Results obtained using a coaxial flashlamp to excite a dilute solution of PPO in ethanol give a lifetime of 1.63±0.02 ns in good agreement with that obtained using conventional fluorometry. The method is also useful in the study of dual emissions such as in monomer–excimer systems and in the measurement of time-resolved emission anisotropy.