A Feasible Add-On Upgrade on a Commercial Two-Photon FLIM Microscope for Optimal FLIM-FRET Imaging of CFP-YFP Pairs.

Fluorescence lifetime imaging microscopy (FLIM) based on time-correlated single photon counting (TCSPC) is a widely used method for fluorescence resonance energy transfer (FRET). Here we report a feasible add-on approach to upgrade a commercial two-photon FLIM microscope into a single-photon FLIM microscope which provides optimal FLIM-FRET imaging of FRET pairs consisting of cyan fluorescent proteins (CFPs) as the donor and yellow fluorescent proteins (YFPs) as the acceptor. The capability of the upgraded system is evaluated and discussed, and the imaging performance of the system is demonstrated using FLIM-FRET experiments with a representative CFP-YFP FRET pair (mCerulean-mCitrine). [ABSTRACT FROM AUTHOR]