Effect of pulse phase and shape on the efficiency of multiphoton processes: Implications for fluorescence microscopy
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The primary laser variables in ultrashort pulse excitation microscopy, which are used to optimize efficiency, were wavelength, average power and peak power. Two more parameters, pulse shape and phase (or chirp), are presented. The effects of spectral shape on two-photon fluorescence efficiency were investigated using an acoustooptic pulse shaper to modify femtosecond pulses from a Ti:sapphire laser. In the case of single-photon excitation of a fluorophore at high energy densities, the fluorescence shows a strong chirp or phase dependence.