FLIM and FCS measurements performed with a master oscillator fiber amplifier (MOFA) laser at 530 nm

Abstract

Upgrade kits towards time-resolved measurements for Confocal Microscopes allow new measurement modes like Fluorescence Lifetime Imaging (FLIM), time-resolved analysis of Fluorescence Correlation Spectroscopy (FCS) and Fluorescence Resonance Energy Transfer (FRET). Microscope users would typically like to use the same excitation wavelength for time-resolved measurements as for steady-state measurements, because their fluorophores are designed for the CW-laser wavelengths usually provided with the system. Pulsed diode lasers, which are ideally used for these upgrade kits are, however, not available for every spectral region of interest. Especially for "green" excitation around 530 nm this is still a problem, as there are no direct emitting laser diodes available.We present a new picosecond pulsed laser system for 530 nm emission with variable repetition rate and pulse energy, which is ideally suited for time-resolved measurements using Time-Correlated Single Photon Counting (TCSPC), and demonstrate its integration into a confocal microscope as well as first results of FLIM and FCS measurements.

Diode laser based light sources for biomedical applications

“…It becomes obvious that gainswitched diode lasers represent the most direct approach towards short picoseconds pulses and can fulfill the requirements in an efficient and compact manner. Therefore, they are well suited for such applications and different groups have already demonstrated gain-switched diode laser based FLIM or FT systems in the visible or near-infrared spectral range [202,208,[224][225][226][227]. The possibilities of subsequent amplification or frequency conversion enable higher peak powers and increase the number of available wavelengths, giving diode lasers a strong standing in the field of biomedical pulsed laser applications.…”