Simple approach to three-color two-photon microscopy by a fiber-optic wavelength convertor

A simple approach to multi-color two-photon microscopy of the red, green, and blue fluorescent indicators was reported based on an ultra-compact 1.03-mu m femtosecond laser and a nonlinear fiber. Inside the nonlinear fiber, the 1.03-mu m laser pulses were simultaneously blue-shifted to 0.6 similar to 0.8 mu m and red-shifted to 1.2 similar to 1.4 mu m region by the Cherenkov radiation and fiber Raman gain effects. The wavelength-shifted 0.6 similar to 0.8 mu m and 1.2 similar to 1.4 mu m radiations were co-propagated with the residual non-converted 1.03-mu m pulses inside the same nonlinear fiber to form a fiber-output three-color femtosecond source. The application of the multi-wavelength sources on multi-color two-photon fluorescence microscopy were also demonstrated. Overall, due to simple system configuration, convenient wavelength conversion, easy wavelength tunability within the entire 0.7 similar to 1.35 mu m biopenetration window and less requirement for high power and bulky light sources, the simple approach to multi-color two-photon microscopy could be widely applicable as an easily implemented and excellent research tool for future biomedical and possibly even clinical applications. (C) 2016 Optical Society of America