Three-dimensional deep tissue multiphoton frequency-domain fluorescence lifetime imaging microscopy via phase multiplexing and adaptive optics

Yide Zhang; Ian H. Guldner; Evan L. Nichols; David Benirschke; Cody J. Smith; Siyuan Zhang; Scott S. Howard

doi:10.1117/12.2510674

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22 February 2019 Three-dimensional deep tissue multiphoton frequency-domain fluorescence lifetime imaging microscopy via phase multiplexing and adaptive optics

Yide Zhang, Ian H. Guldner, Evan L. Nichols, David Benirschke, Cody J. Smith, Siyuan Zhang, Scott S. Howard

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Proceedings Volume 10882, Multiphoton Microscopy in the Biomedical Sciences XIX; 108822H (2019) https://doi.org/10.1117/12.2510674
Event: SPIE BiOS, 2019, San Francisco, California, United States

Abstract

We propose and demonstrate a novel multiphoton frequency-domain fluorescence lifetime imaging microscopy (MPM-FD-FLIM) system that is able to generate 3D lifetime images in deep scattering tissues. The imaging speed of FD-FLIM is improved using phase multiplexing, where the fluorescence signal is split and mixed with the reference signal from the laser in a multiplexing manner. The system allows for easy generation of phasor plots, which not only address multi-exponential decay problems but also clearly represent the dynamics of the fluorophores being investigated. Lastly, a sensorless adaptive optics setup is used for FLIM imaging in deep scattering tissues. The capability of the system is demonstrated in fixed and living animal models, including mice and zebrafish.

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Yide Zhang, Ian H. Guldner, Evan L. Nichols, David Benirschke, Cody J. Smith, Siyuan Zhang, and Scott S. Howard "Three-dimensional deep tissue multiphoton frequency-domain fluorescence lifetime imaging microscopy via phase multiplexing and adaptive optics", Proc. SPIE 10882, Multiphoton Microscopy in the Biomedical Sciences XIX, 108822H (22 February 2019); https://doi.org/10.1117/12.2510674

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