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Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organisms
Abstract
Although the nonlinear optical effect known as second-harmonic generation (SHG) has been recognized since the earliest days of laser physics and was demonstrated through a microscope over 25 years ago, only in the past few years has it begun to emerge as a viable microscope imaging contrast mechanism for visualization of cell and tissue structure and function. Only small modifications are required to equip a standard laser-scanning two-photon microscope for second-harmonic imaging microscopy (SHIM). Recent studies of the three-dimensional in vivo structures of well-ordered protein assemblies, such as collagen, microtubules and muscle myosin, are beginning to establish SHIM as a nondestructive imaging modality that holds promise for both basic research and clinical pathology. Thus far the best signals have been obtained in a transmitted light geometry that precludes in vivo measurements on large living animals. This drawback may be addressed through improvements in the collection of SHG signals via an epi-illumination microscope configuration. In addition, SHG signals from certain membrane-bound dyes have been shown to be highly sensitive to membrane potential. Although this indicates that SHIM may become a valuable tool for probing cell physiology, the small signal size would limit the number of photons that could be collected during the course of a fast action potential. Better dyes and optimized microscope optics could ultimately lead to the imaging of neuronal electrical a…
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“second harmonic generation (SHG) microscopy has emerged as a label-free imaging modality capable of visualizing collagen-rich structures within intact spheroids [259,260].”
From Modeling Tumor Microenvironment Complexity In Vitro: Spheroids as Physiologically Relevant Tumor Models and Strategies for Their Analysis (2025) · cites this paper as [260]
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Metadata
- Authors
- Campagnola, P. J., & Loew, L. M.
- Affiliations
- UConn Health (US)
- Corresponding author
- Loew, Leslie M
- Venue
- Nature Biotechnology
- Published
- 2003-10-31
- Volume
- 21
- Issue
- 11
- Pages
- 1356-1360
- ISSN
- 0733-222X (ISSN-L)
- 1087-0156
- 1546-1696
- OpenAlex
- W2077572701
- Cited by
- 1,326
- References
- 29
- Citations by year
- 2026: 37 · 2025: 42 · 2024: 43 · 2023: 50 · 2022: 55 · 2021: 55 · 2020: 67 · 2019: 79 · 2018: 55 · 2017: 67 · 2016: 73 · 2015: 79 · 2014: 61 · 2013: 77 · 2012: 72
- FWCI
- 3.10
- Type
- journal article
- Language
- en
- Open access
- closed
- Licence
- www.springer.com/tdm (tdm)
- Locations
- Nature Biotechnology (publishedVersion)
- PubMed (publishedVersion)
- Funders
- National Institutes of Health (5r21rr013472-02)
- Office of Naval Research
- National Institute of Biomedical Imaging and Bioengineering (R01EB00196)
- National Center for Research Resources (R21 RR13472)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Spectroscopy Techniques in Biomedical and Chemical Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Cell Image Analysis Techniques
- Keywords
- second harmonic generation, nonlinear optical microscopy, collagen imaging, microtubules, myosin, membrane potential, neuronal electrical activity, in vivo imaging
- MeSH terms
- Action Potentials, Cell Membrane, Cells, Image Enhancement, Membrane Potentials, Proteins, Microscopy, Confocal
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