JBO Letters

Integrated femtosecond stimulated Raman scattering and two-photon fluorescence imaging of subcellular lipid and vesicular structures

[+] Author Affiliations
Xuesong Li, Qiqi Sun, Sicong He

Hong Kong University of Science and Technology, Department of Electronic and Computer Engineering, Biophotonics Research Laboratory, Clear Water Bay, Kowloon, Hong Kong SAR, China

Wen Jiun Lam, Zhe Cao, Yan Hao, Ho Yi Mak

Hong Kong University of Science and Technology, Division of Life Science, Clear Water Bay, Kowloon, Hong Kong SAR, China

Jianan Y. Qu

Hong Kong University of Science and Technology, Department of Electronic and Computer Engineering, Biophotonics Research Laboratory, Clear Water Bay, Kowloon, Hong Kong SAR, China

Hong Kong University of Science and Technology, School of Science and Institute for Advanced Study, Center of Systems Biology and Human Health, Clear Water Bay, Kowloon, Hong Kong SAR, China

J. Biomed. Opt. 20(11), 110501 (Nov 18, 2015). doi:10.1117/1.JBO.20.11.110501
History: Received August 28, 2015; Accepted October 16, 2015
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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.

Figures in this Article
© 2015 Society of Photo-Optical Instrumentation Engineers

Citation

Xuesong Li ; Wen Jiun Lam ; Zhe Cao ; Yan Hao ; Qiqi Sun, et al.
"Integrated femtosecond stimulated Raman scattering and two-photon fluorescence imaging of subcellular lipid and vesicular structures", J. Biomed. Opt. 20(11), 110501 (Nov 18, 2015). ; http://dx.doi.org/10.1117/1.JBO.20.11.110501


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