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Virtual Journal for Biomedical Optics

Virtual Journal for Biomedical Optics


  • Editors: Andrew Dunn and Anthony Durkin
  • Vol. 7, Iss. 7 — Jun. 25, 2012

Long working distance fluorescence lifetime imaging with stimulated emission and electronic time delay

Po-Yen Lin, Shin-Shian Lee, Chia-Seng Chang, and Fu-Jen Kao  »View Author Affiliations

Optics Express, Vol. 20, Issue 10, pp. 11445-11450 (2012)

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In this work, long working distance fluorescence lifetime imaging is realized with stimulated emission in combination with electronic time delay control. Spatial coherence, as a result of stimulated emission, supports unattenuated fluorescence detection at extended distance, using low NA optics. An electronic time delayed trigger provides an advantageous way in adjusting the pulse separation and probing the fluorescence lifetime in the nanosecond ranges. The fluorescence lifetime of selected fluorophores is accurately determined through the pump-probe configuration. The characteristics and applications in fluorescence lifetime measurement of stimulated emission are investigated and summarized succinctly here.

© 2012 OSA

OCIS Codes
(170.0110) Medical optics and biotechnology : Imaging systems
(170.1650) Medical optics and biotechnology : Coherence imaging
(170.6920) Medical optics and biotechnology : Time-resolved imaging

ToC Category:
Medical Optics and Biotechnology

Original Manuscript: February 14, 2012
Revised Manuscript: April 19, 2012
Manuscript Accepted: April 27, 2012
Published: May 4, 2012

Virtual Issues
Vol. 7, Iss. 7 Virtual Journal for Biomedical Optics

Po-Yen Lin, Shin-Shian Lee, Chia-Seng Chang, and Fu-Jen Kao, "Long working distance fluorescence lifetime imaging with stimulated emission and electronic time delay," Opt. Express 20, 11445-11450 (2012)

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  1. R. Y. Tsien, “Imagining imaging’s future,” Nat. Rev. Mol. Cell Biol.4(Suppl), SS16–SS21 (2003). [PubMed]
  2. A. Miyawaki, A. Sawano, and T. Kogure, “Lighting up cells: labeling proteins with fluorophores,” Nat. Cell Biol.5, S1–S7 (2003).
  3. A. Periasamy and R. M. Clegg, “FLIM applications in the biomedical sciences,” in FLIM Microscopy in Biology and Medicine, eds. A. Periasamy and R. M Clegg (CRC Press 2009), p. 385.
  4. J. R. Lakowicz, H. Szmacinski, K. Nowaczyk, and M. L. Johnson, “Fluorescence lifetime imaging of free and protein-bound NADH,” Proc. Natl. Acad. Sci. U.S.A.89(4), 1271–1275 (1992). [CrossRef] [PubMed]
  5. G. McConnell, “Confocal laser scanning fluorescence microscopy with a visible continuum source,” Opt. Express12(13), 2844–2850 (2004). [CrossRef] [PubMed]
  6. W. Min, S. Lu, S. Chong, R. Roy, G. R. Holtom, and X. S. Xie, “Imaging chromophores with undetectable fluorescence by stimulated emission microscopy,” Nature461(7267), 1105–1109 (2009). [CrossRef] [PubMed]
  7. T. Dellwig, M. R. Foreman, and F.-J. Kao, “Coherent long-distance signal detection using stimulated emission: a feasibility study,” Chin. J. Physiol.48, 873–884 (2010).
  8. T. Dellwig, P.-Y. Lin, and F.-J Kao, “Long-distance fluorescence lifetime imaging using stimulated emission,” J. Biomed. Opt. 17, (in press)
  9. E. B. van Munster and T. W. J. Gadella, “Fluorescence lifetime imaging microscopy (FLIM),” Adv. Biochem. Eng. Biotechnol.95, 143–175 (2005). [PubMed]
  10. J. R. Lakowicz, Principles of Fluorescence Spectroscopy, 3rd ed. (New.York: Plenum Press, 2006)
  11. H. Wallrabe and A. Periasamy, “Imaging protein molecules using FRET and FLIM microscopy,” Curr. Opin. Biotechnol.16(1), 19–27 (2005). [CrossRef] [PubMed]
  12. D. Li, W. Zheng, and J. Y. Qu, “Time-resolved spectroscopic imaging reveals the fundamentals of cellular NADH fluorescence,” Opt. Lett.33(20), 2365–2367 (2008). [CrossRef] [PubMed]
  13. C. Hille, M. Lahn, H.-G. Löhmannsröben, and C. Dosche, “Two-photon fluorescence lifetime imaging of intracellular chloride in cockroach salivary glands,” Photochem. Photobiol. Sci.8(3), 319–327 (2009). [CrossRef] [PubMed]
  14. M. Y. Berezin, J. Kao, and S. Achilefu, “pH-Dependent Optical Properties of Synthetic Fluorescent Imidazoles,” Chemistry15(14), 3560–3566 (2009). [CrossRef] [PubMed]
  15. P.-Y. Lin, H.-C. Lyu, C.-Y. S. Hsu, C.-S. Chang, and F.-J. Kao, “Imaging carious dental tissues with multiphoton fluorescence lifetime imaging microscopy,” Biomed. Opt. Express2(1), 149–158 (2011). [CrossRef] [PubMed]
  16. C.-Y. Dong, P. T. C. So, T. French, and E. Gratton, “Fluorescence lifetime imaging by asynchronous pump-probe microscopy,” Biophys. J.69(6), 2234–2242 (1995). [CrossRef] [PubMed]
  17. S. Lu, W. Min, S. Chong, G. R. Holtom, and X. S. Xie, “Label-free imaging of heme proteins with two-photon excited photothermal lens microscopy,” Appl. Phys. Lett.96(11), 113701 (2010). [CrossRef]
  18. E. Rittweger, B. R. Rankin, V. Westphal, and S. W. Hell, “Fluorescence depletion mechanisms in super-resolving STED microscopy,” Chem. Phys. Lett.442(4-6), 483–487 (2007). [CrossRef]
  19. V. Westphal and S. W. Hell, “Nanoscale resolution in the focal plane of an optical microscope,” Phys. Rev. Lett.94(14), 143903 (2005). [CrossRef] [PubMed]
  20. K. Fujita, M. Kobayashi, S. Kawano, M. Yamanaka, and S. Kawata, “High-resolution confocal microscopy by saturated excitation of fluorescence,” Phys. Rev. Lett.99(22), 228105 (2007). [CrossRef] [PubMed]
  21. C. W. Freudiger, W. Min, B. G. Saar, S. Lu, G. R. Holtom, C. W. He, J. C. Tsai, J. X. Kang, and X. S. Xie, “Label-free biomedical imaging with high sensitivity by stimulated Raman scattering microscopy,” Science322(5909), 1857–1861 (2008). [CrossRef] [PubMed]
  22. J. Lin, F. Lu, W. Zheng, S. Xu, D. Tai, H. Yu, and Z. Huang, “Assessment of liver steatosis and fibrosis in a bile duct ligation rat model using integrated coherent anti-Stokes Raman scattering and multiphoton imaging technique,” J. Biomed. Opt.16, 116024 (2011). [CrossRef] [PubMed]
  23. I. Gryczynski, S. W. Hell, and J. R. Lakowicz, “Light quenching of pyridine2 fluorescence with time-delayed pulses,” Biophys. Chem.66(1), 13–24 (1997). [CrossRef] [PubMed]
  24. J. Bückers, D. Wildanger, G. Vicidomini, L. Kastrup, and S. W. Hell, “Simultaneous multi-lifetime multi-color STED imaging for colocalization analyses,” Opt. Express19(4), 3130–3143 (2011). [CrossRef] [PubMed]
  25. K. Kolmakov, V. N. Belov, J. Bierwagen, C. Ringemann, V. Müller, C. Eggeling, and S. W. Hell, “Red-emitting rhodamine dyes for fluorescence microscopy and nanoscopy,” Chemistry16(1), 158–166 (2010). [CrossRef] [PubMed]

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