Portable and wavelength-tunable two-photon microscopy

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DESCRIPTION (provided by applicant): In recent years, efforts to achieve a detailed understanding of biological systems at the cellular and molecular levels have led to increasing requirements for advances in imaging and microscopy. Compared to standard confocal microscopy, two-photon excitation offers distinct advantages including better resolution, less background noise, deeper tissue penetration and less photo-damage. However, a high peak-power Ti: Sapphire laser has until now been required as the excitation source. The high cost and cumbersome size of the Ti:Sapphire laser and its pumping system, along with the difficulty of optical coupling of femtosecond pulses from the source to the microscope, severely limit practical applications for two-photon microscopy. During the last decede, rapid advances in fiber-optic communications have made fiber-based optical devices widely available and affordable. A femtosecond fiber laser the size of a small textbook can deliver >8kW peak power of optical pulses. In this project, we will demonstrate a wavelength-tunable near-infrared light source, called TP-FLEX, for two-photon microscopy. The system is based on the integration of an ultrafast fiber laser, a soliton-based tunable fiber-optic wavelength shifter and a fluorescence microscope. In addition to a substantially simpler generation of excitation optical pulses, rapid wavelength tunability provided by TP-FLEX will significantly enhance the capability of two-photon microscopy. Multi-color labeling of biological samples combined with the ability of wavelength-tunable excitation will enable rapid multi-color imaging through time-domain synchronization between the detector and the excitation source. Moreover, the TP-FLEX tunable laser system will be compact and the wavelength shifting fiber will serve to flexibly interconnect between the fiber laser and the microscope, allowing portability of the light source. The successful demonstration of this system will result in increased accessibility of multi-photon microscopy for a wide range of biological, chemical, and analytical applications. The interdisciplinary research project proposed here brings together three faculty members with expertise in fiber-optic devices and systems, biophysics and spectroscopy and cell biology and microscopy. This integrated effort is critical for the development of an innovative methodology for advanced biological imaging.