Ultrafast single-molecule fluorescence measured by femtosecond double-pulse excitation photon antibunching

Most measurements of fluorescence lifetimes on the single-molecule level are carried out using avalanche photon diodes (APDs). These single-photon counters are inherently slow and their response shows a strong dependence on photon energy, which can make deconvolution of the instrument response function (IRF) challenging. An ultrafast time resolution in single-molecule fluorescence is crucial, e.g., in determining donor lifetimes in donor-acceptor couples which undergo energy transfer, or in plasmonic antenna structures, where the radiative rate is enhanced. We introduce a femtosecond double-excitation (FeDEx) photon correlation technique, which measures the degree of photon antibunching as a function of time delay between two excitation pulses. In this boxcar integration, the time resolution of fluorescence transients is limited solely by the laser pulse length and is independent of the detector IRF. The versatility of the technique is demonstrated with a custom-made donor acceptor complex with one donor ...