OSA's Digital Library

Virtual Journal for Biomedical Optics

Virtual Journal for Biomedical Optics

| EXPLORING THE INTERFACE OF LIGHT AND BIOMEDICINE

  • Editor: Gregory W. Faris
  • Vol. 4, Iss. 6 — May. 26, 2009

Enhanced fluorescence signal in nonlinear microscopy through supplementary fiber-optic light collection

Christoph J. Engelbrecht, Werner Göbel, and Fritjof Helmchen  »View Author Affiliations


Optics Express, Vol. 17, Issue 8, pp. 6421-6435 (2009)
http://dx.doi.org/10.1364/OE.17.006421


View Full Text Article

Enhanced HTML    Acrobat PDF (1597 KB) Open Access





Browse Journals / Lookup Meetings

Browse by Journal and Year


   


Lookup Conference Papers

Close Browse Journals / Lookup Meetings

Article Tools

Share
Citations

Abstract

Nonlinear microscopy techniques crucially rely on efficient signal detection. Here, we present a ring of large-core optical fibers for epi-collection of fluorescence photons that are not transmitted through the objective and thus normally wasted. Theoretical treatments indicated that such a supplementary fiber-optic light collection system (SUFICS) can provide an up to 4-fold signal gain. In typical in vivo imaging experiments, the fiber-ring channel was brighter than the objective channel down to 800 μm depth, thus providing a gain >2. Moreover, SUFICS reduced noise levels in calcium imaging experiments by about 23%. We recommend SUFICS as a generally applicable, effective add-on to nonlinear microscopes for enhancing fluorescence signals.

© 2009 Optical Society of America

OCIS Codes
(110.4280) Imaging systems : Noise in imaging systems
(170.3660) Medical optics and biotechnology : Light propagation in tissues
(180.2520) Microscopy : Fluorescence microscopy
(180.5810) Microscopy : Scanning microscopy
(190.4180) Nonlinear optics : Multiphoton processes
(180.4315) Microscopy : Nonlinear microscopy

ToC Category:
Microscopy

History
Original Manuscript: February 17, 2009
Revised Manuscript: March 29, 2009
Manuscript Accepted: March 30, 2009
Published: April 2, 2009

Virtual Issues
Vol. 4, Iss. 6 Virtual Journal for Biomedical Optics

Citation
Christoph J. Engelbrecht, Werner Göbel, and Fritjof Helmchen, "Enhanced fluorescence signal in nonlinear microscopy through supplementary fiber-optic light collection," Opt. Express 17, 6421-6435 (2009)
http://www.opticsinfobase.org/vjbo/abstract.cfm?URI=oe-17-8-6421


Sort:  Author  |  Year  |  Journal  |  Reset  

References

  1. W. R. Zipfel, R. M. Williams, and W. W. Webb, "Nonlinear magic: multiphoton microscopy in the biosciences," Nat. Biotechnol. 21, 1369-1377 (2003). [CrossRef] [PubMed]
  2. J. Mertz, "Nonlinear microscopy: new techniques and applications," Curr. Opin. Neurobiol. 14, 610-616 (2004). [CrossRef] [PubMed]
  3. W. Denk, J. H. Strickler, and W. W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990). [CrossRef] [PubMed]
  4. F. Helmchen and W. Denk, "Deep tissue two-photon microscopy," Nat. Methods 2, 932-940 (2005). [CrossRef] [PubMed]
  5. K. Svoboda and R. Yasuda, "Principles of two-photon excitation microscopy and its applications to neuroscience," Neuron 50, 823-839 (2006). [CrossRef] [PubMed]
  6. W. Denk, D. W. Piston, and W. W. Webb, "Multi-photon molecular excitation in laser-scanning microscopy," in Handbook of Biological Confocal Microscopy, J. B. Pawley, ed., (Plenum Press, New York, 2005).
  7. M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, "Two-photon microscopy in brain tissue: parameters influencing the imaging depth (Vol. 111, pg 29, 2001)," J. Neurosci. Meth. 112, 205 (2001). [CrossRef]
  8. E. Beaurepaire and J. Mertz, "Epifluorescence collection in two-photon microscopy," Appl. Opt. 41, 5376-5382 (2002). [CrossRef] [PubMed]
  9. H. J. Koester, D. Baur, R. Uhl, and S. W. Hell, "Ca2+ fluorescence imaging with pico- and femtosecond two-photon excitation: signal and photodamage," Biophys. J. 77, 2226-2236 (1999). [CrossRef] [PubMed]
  10. Z. F. Mainen, M. Maletic-Savatic, S. H. Shi, Y. Hayashi, R. Malinow, and K. Svoboda, "Two-photon imaging in living brain slices," Methods 18, 231-239 (1999). [CrossRef] [PubMed]
  11. D. Vucinic, T. M. Bartol, and T. J. Sejnowski, "Hybrid reflecting objectives for functional multiphoton microscopy in turbid media," Opt. Lett. 31, 2447-2449 (2006). [CrossRef] [PubMed]
  12. C. A. Combs, A. V. Smirnov, J. D. Riley, A. H. Gandjbakhche, J. R. Knutson, and R. S. Balaban, "Optimization of multiphoton excitation microscopy by total emission detection using a parabolic light reflector," J. Microsc. 228, 330-337 (2007). [CrossRef] [PubMed]
  13. B. A. Flusberg, E. D. Cocker, W. Piyawattanametha, J. C. Jung, E. L. M. Cheung, and M. J. Schnitzer, "Fiber-optic fluorescence imaging," Nat. Methods 2, 941-950 (2005). [CrossRef] [PubMed]
  14. C. J. Engelbrecht, R. S. Johnston, E. J. Seibel, and F. Helmchen, "Ultra-compact fiber-optic two-photon microscope for functional fluorescence imaging in vivo," Opt. Express 16, 5556-5564 (2008). [CrossRef] [PubMed]
  15. W. Göbel, B. M. Kampa, and F. Helmchen, "Imaging cellular network dynamics in three dimensions using fast 3D laser scanning," Nat. Methods 4, 73-79 (2007). [CrossRef]
  16. C. Stosiek, O. Garaschuk, K. Holthoff, and A. Konnerth, "In vivo two-photon calcium imaging of neuronal networks," Proc. Natl. Acad. Sci. U.S.A. 100, 7319-7324 (2003). [CrossRef] [PubMed]
  17. J. N. D. Kerr, D. Greenberg, and F. Helmchen, "Imaging input and output of neocortical networks in vivo " Proc. Natl. Acad. Sci. U.S.A. 102, 14063-14068 (2005). [CrossRef] [PubMed]
  18. W. Göbel and F. Helmchen, "In vivo calcium imaging of neural network function," Physiology (Bethesda) 22, 358-365 (2007). [CrossRef]
  19. L. Wang, S. L. Jacques, and L. Zheng, "MCML--Monte Carlo modeling of light transport in multi-layered tissues," Comput. Methods Programs Biomed. 47, 131-146 (1995). [CrossRef] [PubMed]
  20. L. G. Henyey and J. L. Greenstein, "Diffuse radiation in the galaxy," Astrophys. J. 93, 70-83 (1941). [CrossRef]
  21. S. L. Jacques and S. A. Prahl, "Steady-state Monte Carlo: A minimal program, "mc321.c"." (1998), http://omlc.ogi.edu/classroom/ece532/class4/ssmc/roulette.html.
  22. P. Theer and W. Denk, "On the fundamental imaging-depth limit in two-photon microscopy," J. Opt. Soc. Am. A Opt. Image Sci. Vis. 23, 3139-3149 (2006). [CrossRef] [PubMed]
  23. A. Leray, C. Odin, and Y. Le Grand, "Out-of-focus fluorescence collection in two-photon microscopy of scattering media," Opt. Commun. 281, 6139-6144 (2008). [CrossRef]
  24. E. J. Seibel, R. S. Johnston, and C. D. Melville, "A full-color scanning fiber endoscope," Optical Fibers and Sensors for Medical Diagnostics and Treatment Applications VI.Proc. SPIE. 6083, 9-16 (2006).
  25. J. D. McMullen and W. Zipfel, "A Scheme for Increasing the Collection Efficiency of Multiphoton Microscopy," Biophys. J. 96, 639a (2009). [CrossRef]
  26. A. Hopt and E. Neher, "Highly nonlinear photodamage in two-photon fluorescence microscopy," Biophys. J. 80, 2029-2036 (2001). [CrossRef] [PubMed]
  27. G. Donnert, C. Eggeling, and S. W. Hell, "Major signal increase in fluorescence microscopy through dark-state relaxation," Nat. Methods 4, 81-86 (2007). [CrossRef]
  28. N. Ji, J. C. Magee, and E. Betzig, "High-speed, low-photodamage nonlinear imaging using passive pulse splitters," Nat. Methods 5, 197-202 (2008). [CrossRef] [PubMed]
  29. D. Debarre, N. Olivier, and E. Beaurepaire, "Signal epidetection in third-harmonic generation microscopy of turbid media," Opt. Express 15, 8913-8924 (2007). [CrossRef] [PubMed]
  30. J. X. Cheng, Y. K. Jia, G. Zheng, and X. S. Xie, "Laser-scanning coherent anti-Stokes Raman scattering microscopy and applications to cell biology," Biophys. J. 83, 502-509 (2002). [CrossRef] [PubMed]
  31. E. W. Weisstein, "Circle-Circle Intersection," http://mathworld.wolfram.com/Circle-CircleIntersection.html.

Cited By

Alert me when this paper is cited

OSA is able to provide readers links to articles that cite this paper by participating in CrossRef's Cited-By Linking service. CrossRef includes content from more than 3000 publishers and societies. In addition to listing OSA journal articles that cite this paper, citing articles from other participating publishers will also be listed.


« Previous Article  |  Next Article »

OSA is a member of CrossRef.

CrossCheck Deposited