We optimized the spectral coverage of self-phase-modulation-enabled femtosecond fiber sources by careful investigations into the influence of input pulse width, fiber length, and fiber damage, and we have demonstrated a widely tunable source ranging from 740-1250 nm for two-photon microscopy applications. In addition, tens of milliwatt tunable near-UV/visible spectrum is easily obtained by a frequency-doubled conversion, and the gap between the fundamental and frequency-doubled spectra can be filled with the nonlinear wave breaking around 650 nm. A multi-modality microscopy incorporating two-photon microscopy and confocal fluorescence microscopy was also demonstrated to prove the versatility of our development for biomedical imaging.
We have realized video-rate three-photon fluorescence microscopy based on a simple 1320-nm femtosecond source, which is driven by a 24-MHz Cr:forsterite oscillator and fiber-optic nonlinear conversion. The development aims to find a robust pumping source for deep-tissue GFP imaging. We optimized the laser oscillator to deliver 40-nJ output pulse energy at around 1260 nm, and the matching of the laser spectrum and GFP excitation can be realized by precisely control the interplay between self-phase modulation and fiber dispersion. We have obtained clear video-rate three-photon imaging from fluorescent beads, and further optimization leading to deep-tissue GFP imaging will be discussed.
We have demonstrated a widely-tunable femtosecond fiber source between 770-1180 nm enabled by self-phase modulation, and the wide spectral coverage is suitable for most of two-photon fluorescence microscopy applications. Based on femtosecond Yb:fiber laser, we also compared spectral broadening in different dispersion regimes using different photonic crystal fibers. We managed to maximize the self-phase modulated feature from the broadened spectra while avoiding unwanted nonlinear temporal trapping. The optimization of fiber selection and laser input conditions led to a wide tunability down to below 800nm region, which is the most commonly used two-photon excitation wavelength for many intrinsic fluorescent labels in biological tissues. We believe this fiber-based femtosecond source can be a relatively cost-effective and robust solution for most of two-photon fluorescence microscopy applications.
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