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18 November 2015 Integrated femtosecond stimulated Raman scattering and two-photon fluorescence imaging of subcellular lipid and vesicular structures
Xuesong Li, Wen Jiun Lam, Zhe Cao, Yan Hao, Qiqi Sun, Sicong He, Ho Yi Mak, Jianan Y. Qu
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Journal of Biomedical Optics, Vol. 20, Issue 11, 110501 (November 2015). https://doi.org/10.1117/1.JBO.20.11.110501
Abstract
The primary goal of this study is to demonstrate that stimulated Raman scattering (SRS) as a new imaging modality can be integrated into a femtosecond (fs) nonlinear optical (NLO) microscope system. The fs sources of high pulse peak power are routinely used in multimodal nonlinear microscopy to enable efficient excitation of multiple NLO signals. However, with fs excitations, the SRS imaging of subcellular lipid and vesicular structures encounters significant interference from proteins due to poor spectral resolution and a lack of chemical specificity, respectively. We developed a unique NLO microscope of fs excitation that enables rapid acquisition of SRS and multiple two-photon excited fluorescence (TPEF) signals. In the in vivo imaging of transgenic C. elegans animals, we discovered that by cross-filtering false positive lipid signals based on the TPEF signals from tryptophan-bearing endogenous proteins and lysosome-related organelles, the imaging system produced highly accurate assignment of SRS signals to lipid. Furthermore, we demonstrated that the multimodal NLO microscope system could sequentially image lipid structure/content and organelles, such as mitochondria, lysosomes, and the endoplasmic reticulum, which are intricately linked to lipid metabolism.
© 2015 Society of Photo-Optical Instrumentation Engineers (SPIE) 1083-3668/2015/$25.00 © 2015 SPIE
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Xuesong Li, Wen Jiun Lam, Zhe Cao, Yan Hao, Qiqi Sun, Sicong He, Ho Yi Mak, and Jianan Y. Qu "Integrated femtosecond stimulated Raman scattering and two-photon fluorescence imaging of subcellular lipid and vesicular structures," Journal of Biomedical Optics 20(11), 110501 (18 November 2015). https://doi.org/10.1117/1.JBO.20.11.110501
Published: 18 November 2015
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Cited by 11 scholarly publications.
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KEYWORDS
Green fluorescent protein
Imaging systems
Proteins
Nonlinear optics
Luminescence
Microscopes
Raman scattering
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