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In multiphoton microscopy, measurements and compression of the laser pulsewidth at the setup plays an important role in optimizing the fluorescenceoutput with minimal laser power applications. For example, in deep two andthree-photon intravital microscopy it is very crucial to have shortest laserpulses for efficient fluorescence excitation with lowest possible laser powerat the sample. There are several published experimental methods and techniquesfor measuring the group delay dispersion and pulse width. Yet, there arepractical limitations for their implementation and beam optimization atintravital setups in individual labs or microscope facility. Here, we present amethod to construct a portable optical collinear autocorrelator, to probe thebeam directly after the objective and at desired place in the beam path tomeasure the pulse width, and to optimize the pulse width with a pulsecompressor. Finally, we demonstrate that the method increases fluorescenceexcitation efficiency in three-photon microscopy.

Measuring ultra-short pulse widths before and after the objective with a home built autocorrelator