Simultaneous time- and spectrum-resolved multifocal multiphoton microscopy

We present a novel multifocal multiphoton microscope that is based on a sampling imaging technique using a microlens array, a prism for two-dimensionalspectral dispersion, and a specially designed streak camera to provide simultaneous time- and spectrum-resolved fluorescence microscopy. We split the near-infrared light of a mode-locked titanium:sapphire femtosecond laser into an array of beams that are transformed into an array of high-aperture foci at the sample. A time- and spectrum-resolved image of 3x3 foci on the sample can be obtained with a snapshot. By translating the sample stage laterally and axially and implementing a dedicated image reconstruction algorithm in the control system of the instrument, we demonstrate the acquisition of a five-dimensional data set combining lifetime and spectral resolutions in biological fluorescence imaging.