In narrow-bandwidth coherent Raman scattering (CRS) microscopy, efficient signal generation is accomplished with two-color laser sources providing synchronized picosecond pulses whose frequency difference and spectral widths match the molecular Raman frequency and bandwidth, respectively. With vibrational bandwidths of typically 10 cm-1, the optimum laser pulse durations thus correspond to about 2 ps. Here, we present a new light source consisting of an amplified Yb-fiber oscillator providing 2-ps pulses at 1031 nm and a synchronously green-pumped optical parametric oscillator (OPO). The OPO slightly shortens the pulses to < 2 ps while maintaining a bandwidth of 10 cm-1. Output power levels of 1 W in both the 1031-nm and the OPO-branch with continuously tunable frequency differences between the two beams covering a broad range from 700 to 4500 cm-1 are achieved. In addition to CARS microscopy, this light source allows for SRS imaging via an integrated electro-optical modulation of the 1031-nm beam at 20 MHz with a depth of >95%, locked to the laser repetition rate of 80 MHz. The OPO noise at 20 MHz was found to be only 60% above the combined detector and laser noise of a conventional Nd:YVO pump source. This represents a significant reduction in laser noise when compared to other fiber-based laser sources previously proposed for SRS microscopy. When SRS imaging with this new light is compared with a Nd:YVO pumped OPO (delivering 7 ps and 5 ps pulses, respectively), a 5- to 6-fold increase in SRS signal strength and signal-to-noise ratio has been achieved. Video-rate SRS and the capability of multi-spectral SRS imaging are demonstrated.
We demonstrate the application of three photon excitation to fluorescence probe studies using time-correlated single- photon counting. By exciting with 120 fs Ti:sapphire laser pulses at 800 nm we have observed fluorescence emission from the scintillators p-Terphenyl (PT) and 2,1 Naphthyl, 5- Phenyloxazole ((alpha) -NPO). For solutions of (alpha) -NPO in cyclohexane and PT in propylene glycol the laser power dependence of the fluorescence is consistent with the emission being due to three-photon excitation of the same emitting S1 state which is populated with one-photon excitation at 267 nm. However, for (alpha) -NPO in alcohols some evidence for a mixture of three and two photon excitation is observed. This solute dependence correlates with the red edge of the one-photon absorption spectra. The observation of excimer emission and fluorescence anisotropy of (alpha) -NPO in small unilamellar vesicles (SUVs) of L- (alpha) -dipalmitoylphosphatidylcholine excited at 800 nm provides a clear illustration of the potential for using three photon excitation in fluorescence probe studies of microheterogeneous media. In SUVs the time-resolved data is consistent with a heterogeneous distribution of (alpha) -NPO molecules between isolated sites and ground state clusters in a similar manner to that which we reported previously for 2,5-diphenyloxazole.
Some photophysical properties (steady-state absorption, fluorescence and phosphorescence, fluorescence decay times and singlet oxygen quantum yields) of silicon phthalocyanines with methoxypolyethylene glycol (MPEG)-substituents of various chain length at the silicon atom [SiPc(OCH2CH2)n-OCH3; n equals 1,2,3,12], silicon phthalocyanine covalently bound to the water soluble polymer methoxypolyethylene glycol [SiPc(-O-MPEG 5000)2] and silicon naphthalocyanine [SiNc(CH2CH2OCH3)2] have been studied. The aim of these investigations was to get information about the influence of methoxypolyethylene-glycol-coupling on photophysical sensitizer parameters.
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