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Advanced vibrational microscopes for life science

Nature Methods volume 22, pages 912–927 (2025)Cite this article

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Abstract

Providing molecular fingerprint information, vibrational spectroscopic imaging opens a new window to decipher the function of biomolecules in living systems. While classic vibrational microscopes based on spontaneous Raman scattering or mid-infrared absorption offer rich insights into sample composition, they have very small cross sections or poor spatial resolution. Nonlinear vibrational microscopy, based on coherent Raman scattering or optical photothermal detection of vibrational absorption, overcomes these barriers and enables high-speed and high-sensitivity imaging of chemical bonds in live cells and tissues. Here, we introduce various modalities, including their principles, strengths, weaknesses and data mining methods to the life sciences community. We further provide a guide for prospective users and an outlook on future technological advances.

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Fig. 1: Energy diagrams for advanced vibrational microscopy.
Fig. 2: Various modalities developed for CARS imaging.
Fig. 3: Various modalities developed for SRS imaging.
Fig. 4: Contrast mechanisms of MIP imaging.
Fig. 5: MIP imaging implementations.
Fig. 6: Broad applications of advanced vibrational microscopy to life sciences.

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Acknowledgements

We acknowledge grants from the National Institutes of Health (R35 GM136223, R01 EB032391, R01 EB035429, R01 AI141439, R01 CA224275 and R33CA287046) and the Chan Zuckerberg Initiative DAF (2023-321163), an advised fund of Silicon Valley Community Foundation.

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Authors and Affiliations

  1. Department of Electrical and Computer Engineering, Boston University, Boston, MA, USA

    Ji-Xin Cheng, Yuhao Yuan, Hongli Ni, Jianpeng Ao, Qing Xia & Xiaowei Ge

  2. Department of Biomedical Engineering, Boston University, Boston, MA, USA

    Ji-Xin Cheng

  3. Department of Chemistry, Boston University, Boston, MA, USA

    Ji-Xin Cheng & Rylie Bolarinho

  4. Photonics Center, Boston University, Boston, MA, USA

    Ji-Xin Cheng

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  1. Ji-Xin Cheng
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  5. Qing Xia
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  7. Xiaowei Ge
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Contributions

J.-X.C. drafted the outline, introduction, outlook and two boxes. Y.Y. drafted the CARS section. H.N. drafted the SRS and SWIP sections. J.A. drafted the data science section. R.B. and Q.X. drafted the MIP section. X.G. and H.N. drafted the SRP and SWIP section. Q.X., J.A. and Y.Y. drafted the applications and user guide section. J.A., Q.X. and Y.Y. contributed to manuscript revision.

Corresponding author

Correspondence to Ji-Xin Cheng.

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Competing interests

J.-X.C. declares competing interests with VibroniX and Photothermal Spectroscopy, which did not fund this work. The remaining authors declare no competing interests.

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Nature Methods thanks Caitlin Davis, Vladislav Yakovlev, and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Primary Handling Editor: Rita Strack, in collaboration with the Nature Methods team.

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Supplementary Notes and Supplementary Tables 1–5

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Cheng, JX., Yuan, Y., Ni, H. et al. Advanced vibrational microscopes for life science. Nat Methods 22, 912–927 (2025). https://doi.org/10.1038/s41592-025-02655-w

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