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Peer-reviewedCC BYAnalytical chemistry

Effects of Chemical Fixation on Stimulated Raman Scattering Microscopy.

  • Chiara Ceconello1,2
  • Wuji Han3
  • Bryce Manifold1
  • Dario Polli2,4
  • Aaron Streets1,3,5,6
6 affiliations
  1. 1Department of Bioengineering, University of California Berkeley, Berkeley, California94720, United States.
  2. 2Department of Physics, Politecnico di Milano, Piazza Leonardo da Vinci 32, Milano20133, Italy.
  3. 3Center for Computational Biology, University of California Berkeley, Berkeley, California94720, United States.
  4. 4CNR-Institute for Photonics and Nanotechnologies (IFN-CNR), Piazza Leonardo Da Vinci 32, Milano20133, Italy.
  5. 5Biophysics Graduate Group, University of California Berkeley, Berkeley, California94720, United States.
  6. 6Biohub, San Francisco, California94158, United States.
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Abstract

Stimulated Raman scattering (SRS) microscopy enables label-free imaging of cells and tissues in their native state with chemical specificity. However, there are often experimental advantages of chemical fixation of samples prior to imaging, which can introduce perturbations that may alter the native state of the samples and possibly impact the SRS signal. In this study, we systematically characterize the effects of common fixatives (paraformaldehyde, formalin, glutaraldehyde, methanol, ethanol) on the preservation of cellular integrity and molecular composition in a HeLa cell model as observed by SRS microscopy. We demonstrate how the different fixatives can influence lipid and protein content and overall cell morphology, with significant implications for the accuracy of quantitative SRS microscopy. Our findings indicate that formaldehyde-based fixatives impose minimal disruption to the spatial or spectral features of biomolecules compared to the other fixatives tested and suggest glutaraldehyde (GA) as a suitable alternative for SRS imaging. This study provides insights into the choice of optimal sample preparation, enabling more reliable SRS-based studies for the evaluation of cellular processes and disease mechanisms where fixation is used.

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Cite

BibTeXRIS
BibTeX
@article{ceconello2026effects,
  title = {{Effects of Chemical Fixation on Stimulated Raman Scattering Microscopy.}},
  author = {Chiara Ceconello and Wuji Han and Bryce Manifold and Dario Polli and Aaron Streets},
  journal = {Analytical chemistry},
  year = {2026},
  month = oct,
  doi = {10.1021/acs.analchem.6c01478},
  url = {https://europepmc.org/article/MED/42915705},
  note = {CC BY},
}
RIS
TY  - JOUR
TI  - Effects of Chemical Fixation on Stimulated Raman Scattering Microscopy.
AU  - Chiara Ceconello
AU  - Wuji Han
AU  - Bryce Manifold
AU  - Dario Polli
AU  - Aaron Streets
PY  - 2026
DA  - 2026/10/01
JO  - Analytical chemistry
DO  - 10.1021/acs.analchem.6c01478
UR  - https://europepmc.org/article/MED/42915705
AB  - Stimulated Raman scattering (SRS) microscopy enables label-free imaging of cells and tissues in their native state with chemical specificity. However, there are often experimental advantages of chemical fixation of samples prior to imaging, which can introduce perturbations that may alter the native state of the samples and possibly impact the SRS signal. In this study, we systematically characterize the effects of common fixatives (paraformaldehyde, formalin, glutaraldehyde, methanol, ethanol) on the preservation of cellular integrity and molecular composition in a HeLa cell model as observed by SRS microscopy. We demonstrate how the different fixatives can influence lipid and protein content and overall cell morphology, with significant implications for the accuracy of quantitative SRS microscopy. Our findings indicate that formaldehyde-based fixatives impose minimal disruption to the spatial or spectral features of biomolecules compared to the other fixatives tested and suggest glutaraldehyde (GA) as a suitable alternative for SRS imaging. This study provides insights into the choice of optimal sample preparation, enabling more reliable SRS-based studies for the evaluation of cellular processes and disease mechanisms where fixation is used.
SN  - 0003-2700
SN  - 1520-6882
ER  -