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High-Resolution Nonlinear Optical Imaging of Live Cells by Second Harmonic Generation
Open accessAbstract
By adapting a laser scanning microscope with a titanium sapphire femtosecond pulsed laser and transmission optics, we are able to produce live cell images based on the nonlinear optical phenomenon of second harmonic generation (SHG). Second harmonic imaging (SHIM) is an ideal method for probing membranes of living cells because it offers the high resolution of nonlinear optical microscopy with the potential for near-total avoidance of photobleaching and phototoxicity. The technique has been implemented on three cell lines labeled with membrane-staining dyes that have large nonlinear optical coefficients. The images can be obtained within physiologically relevant time scales. Both achiral and chiral dyes were used to compare image formation for the case of single- and double-leaflet staining, and it was found that chirality plays a significant role in the mechanism of contrast generation. It is also shown that SHIM is highly sensitive to membrane potential, with a depolarization of 25 mV resulting in an approximately twofold loss of signal intensity.
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Cites
Kleinfeld, D., Mitra, P. P., Helmchen, F., & Denk, W. (1998) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.95.26.15741 · Cited by 832
Cited by
Campagnola, P. J., & Loew, L. M. (2003) · Nature Biotechnology · doi:10.1038/nbt894 · Cited by 1,326
Self-citationPrinciples of two-photon excitation microscopy and its applications to neuroscience
Svoboda, K., & Yasuda, R. (2006) · Neuron · doi:10.1016/j.neuron.2006.05.019 · Cited by 1,027
Coherent Anti-Stokes Raman Scattering Microscopy: Instrumentation, Theory, and Applications
Cheng, J.-X., & Xie, X. S. (2004) · The Journal of Physical Chemistry B · doi:10.1021/jp035693v · Cited by 963
Zoumi, A., Yeh, A., & Tromberg, B. J. (2002) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.172368799 · Cited by 870
Laser-Scanning Coherent Anti-Stokes Raman Scattering Microscopy and Applications to Cell Biology
Cheng, J.-X., Jia, Y. K., Zheng, G., & Xie, X. S. (2002) · Biophysical Journal · doi:10.1016/s0006-3495(02)75186-2 · Cited by 371
Metadata
- Authors
- Campagnola, P. J., Wei, M.-D., Lewis, A., & Loew, L. M.
- Venue
- Biophysical Journal
- Publisher
- Elsevier BV
- Published
- 1999-12
- Volume
- 77
- Issue
- 6
- Pages
- 3341-3349
- ISSN
- 0006-3495 (ISSN-L)
- OpenAlex
- W1997225236
- Cited by
- 504
- References
- 1
- FWCI
- 9.20
- Type
- journal article
- Language
- en
- Open access
- bronze
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- Campagnola, P. J., Wei, M.-D., Lewis, A., & Loew, L. M. (1999). High-Resolution Nonlinear Optical Imaging of Live Cells by Second Harmonic Generation. Biophysical Journal. https://doi.org/10.1016/s0006-3495(99)77165-1
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Bibliographic data: Semantic Scholar, OpenAlex, Crossref.
Contains information from the Semantic Scholar Academic Graph Datasets, made available under the ODC Attribution License (ODC-BY 1.0).
Semantic Scholar’s data is described in: Kinney, R., et al. (2023). The Semantic Scholar Open Data Platform. arXiv:2301.10140.
