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Two-photon fluorescence imaging using a tunable spectral window based on fiber supercontinuum

Zhongyun Chen, Jiangfeng Huang, Xinyuan Huang, Xiujuan Gao, Yifu Zhou, and Ling Fu

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Abstract

Two-photon excitation fluorescence (TPEF) microscopy has evolved into a versatile tool in biological research. However, the multiplexing capability of TPEF microscopy is limited by the narrow spectral bandwidth of the light source. In this study, we apply a photonic crystal fiber in TPEF microscopy to broaden the excitation source bandwidth. We tuned the spectral window using a spatial light modulator as a programmable diffraction grating that was placed behind a prism pair. In addition, we combined a grating pair to compensate for dispersion to improve the two-photon excitation efficiency. The combination of a broad spectrum and a programmable grating enabled fast spectral window tuning rate on a time scale of tens of milliseconds. We demonstrate the performance of our method by imaging live B16 cells labeled with four emission spectrum overlapped fluorescent proteins.

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Supplementary Material (1)

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Supplement 1       The supplementary detail about the experiments

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The data underlying the results presented in this study are not publicly available at this time but may be obtained from the authors upon reasonable request.

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