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Compact multiphoton/single photon laser scanning microscope for spectral imaging and fluorescence lifetime imaging

: Ulrich, V.; Fischer, P.; Riemann, I.; Konig, K.


Scanning 26 (2004), Nr.5, S.217-225
ISSN: 0161-0457
ISSN: 1932-8745
Fraunhofer IBMT ()

An inverted fluorescence microscope was upgraded into a compact three-dimensional laser scanning microscope (LSM) of 65 x 62 x 48 cm dimensions by means of a fast kHz galvoscanner unit, a piezodriven z-stage, and a picosecond (ps) 50 MHz laser diode at 405 nm. In addition, compact turn-key near infrared femtosecond lasers have been employed to perform multiphoton fluorescence and second harmonic generation (SHG) microscopy. To expand the features of the compact LSM, a time-correlated single photon counting unit as well as a Sagnac interferometer have been added to realize fluorescence lifetime imaging (FLIM) and spectral imaging. Using this unique five-dimensional microscope, TauMap, single-photon excited (SPE), and two-photon excited (TPE) cellular fluorescence as well as intratissue autofluorescence of water plant leaves have been investigated with submicron spatial resolution, < 270 ps temporal resolution, and 10 nm spectral resolution.