OSA's Digital Library

Optics Express

Optics Express

  • Editor: Andrew M. Weiner
  • Vol. 22, Iss. 9 — May. 5, 2014
  • pp: 10728–10734

TIRF microscopy with ultra-short penetration depth

Hao Shen, Eric Huang, Tapaswini Das, Hongxing Xu, Mark Ellisman, and Zhaowei Liu  »View Author Affiliations

Optics Express, Vol. 22, Issue 9, pp. 10728-10734 (2014)

View Full Text Article

Enhanced HTML    Acrobat PDF (1189 KB)

Browse Journals / Lookup Meetings

Browse by Journal and Year


Lookup Conference Papers

Close Browse Journals / Lookup Meetings

Article Tools



Total internal reflection fluorescence microscopy (TIRF), in both commercial and custom-built configurations, is widely used for high signal-noise ratio imaging. The imaging depth of traditional TIRF is sensitive to the incident angle of the laser, and normally limited to around 100 nm. In our paper, using a high refractive index material and the evanescent waves of various waveguide modes, we propose a compact and tunable ultra-short decay length TIRF system, which can reach decay lengths as short as 19 nm, and demonstrate its application for imaging fluorescent dye-labeled F-actin in HeLa cells.

© 2014 Optical Society of America

OCIS Codes
(110.0180) Imaging systems : Microscopy
(180.2520) Microscopy : Fluorescence microscopy
(240.6690) Optics at surfaces : Surface waves

ToC Category:

Original Manuscript: March 13, 2014
Revised Manuscript: April 17, 2014
Manuscript Accepted: April 17, 2014
Published: April 25, 2014

Virtual Issues
Vol. 9, Iss. 7 Virtual Journal for Biomedical Optics

Hao Shen, Eric Huang, Tapaswini Das, Hongxing Xu, Mark Ellisman, and Zhaowei Liu, "TIRF microscopy with ultra-short penetration depth," Opt. Express 22, 10728-10734 (2014)

Sort:  Author  |  Year  |  Journal  |  Reset  


  1. E. M. Slayter, Optical Methods in Biology (R. E. Krieger, 1976).
  2. S. Kudo, Y. Magariyama, S.-I. Aizawa, “Abrupt changes in flagellar rotation observed by laser dark-field microscopy,” Nature 346, 677–680 (1990). [CrossRef] [PubMed]
  3. J. B. Pawley, Handbook of Biological Confocal Microscopy (Springer, 1995).
  4. T. Wilson, Confocal Microscopy (Academic, 1990).
  5. M. Bates, B. Huang, G. T. Dempsey, X. Zhuang, “Multicolor super-resolution imaging with photo-switchable fluorescent probes,” Science 317, 1749–1753 (2007). [CrossRef] [PubMed]
  6. M. J. Rust, M. Bates, X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods 3, 793–796 (2006). [CrossRef] [PubMed]
  7. S. W. Hell, J. Wichmann, “Breaking the diffraction resolution limit by stimulated emission: stimulated-emission-depletion fluorescence microscopy,” Opt. Lett. 19, 780–782 (1994). [CrossRef] [PubMed]
  8. T. A. Klar, S. W. Hell, “Subdiffraction resolution in far-field fluorescence microscopy,” Opt. Lett. 24, 954–956 (1999). [CrossRef]
  9. B. Bailey, D. L. Farkas, D. L. Taylor, F. Lanni, “Enhancement of axial resolution in fluorescence microscopy by standing-wave excitation,” Nature 366, 44–48 (1993). [CrossRef] [PubMed]
  10. M. G. L. Gustafsson, “Surpassing the lateral resolution limit by a factor of two using structured illumination microscopy,” J. Microsc. 198, 82–87 (2000). [CrossRef] [PubMed]
  11. F. Wei, Z. Liu, “Plasmonic structured illumination microscopy,” Nano Lett. 10, 2531–2536 (2010). [CrossRef] [PubMed]
  12. F. W. Rost, Fluorescence Microscopy (Cambridge University, 1995), Vol. 1–3.
  13. T. Funatsu, Y. Harada, M. Tokunaga, K. Saito, T. Yanagida, “Imaging of single fluorescent molecules and individual ATP turnovers by single myosin molecules in aqueous solution,” Nature 374, 555–559 (1995). [CrossRef] [PubMed]
  14. J. E. Hobbie, R. J. Daley, S. Jasper, “Use of nuclepore filters for counting bacteria by fluorescence microscopy,” Appl. Environ. Microbiol. 33, 1225–1228 (1977). [PubMed]
  15. R. Pepperkok, J. Ellenberg, “High-throughput fluorescence microscopy for systems biology,” Nat. Rev. Mol. Cell Biol. 7, 690–696 (2006). [CrossRef] [PubMed]
  16. J. M. Prober, G. L. Trainor, R. J. Dam, F. W. Hobbs, C. W. Robertson, R. J. Zagursky, A. J. Cocuzza, M. A. Jensen, K. Baumeister, “A system for rapid DNA sequencing with fluorescent chain-terminating dideoxynu-cleotides,” Science 238, 336–341 (1987). [CrossRef] [PubMed]
  17. G. A. Wagnieres, W. M. Star, B. C. Wilson, “In vivo fluorescence spectroscopy and imaging for oncological applications,” Photochem. Photobiol. 68, 603–632 (1998). [CrossRef] [PubMed]
  18. D. Axelrod, “Total internal reflection fluorescence microscopy,” Method. Cell Biol. 89, 169–221 (2008). [CrossRef]
  19. D. Axelrod, “Cell-substrate contacts illuminated by total internal-reflection fluorescence,” J. Cell Biol. 89, 141–145 (1981). [CrossRef] [PubMed]
  20. D. Breitsprecher, A. K. Kiesewetter, J. Linkner, J. Faix, “Analysis of actin assembly by in vitro TIRF microscopy,” in Chemotaxis (Springer, 2009), pp. 401–415. [CrossRef]
  21. B. Lassen, M. Malmsten, “Competitive protein adsorption studied with TIRF and ellipsometry,” J. Colloid Interface Sci. 179, 470–477 (1996). [CrossRef]
  22. M. Ohara-Imaizumi, C. Nishiwaki, T. Kikuta, S. Nagai, Y. Nakamichi, S. Nagamatsu, “TIRF imaging of docking and fusion of single insulin granule motion in primary rat pancreatic beta-cells: different behaviour of granule motion between normal and Goto-Kakizaki diabetic rat beta-cells,” Biochem. J. 381, 13–18 (2004). [CrossRef] [PubMed]
  23. M. Tokunaga, K. Kitamura, K. Saito, A. H. Iwane, T. Yanagida, “Single molecule imaging of fluorophores and enzymatic reactions achieved by objective-type total internal reflection fluorescence microscopy,” Biochem. Biophys. Res. Commun. 235, 47–53 (1997). [CrossRef] [PubMed]
  24. C. M. Ajo-Franklin, L. Kam, S. G. Boxer, “High refractive index substrates for fluorescence microscopy of biological interfaces with high z contrast,” Proc. Natl. Acad. Sci. U. S. A. 98, 13643–13648 (2001). [CrossRef] [PubMed]
  25. D. S. Johnson, J. K. Jaiswal, S. Simon, “Total internal reflection fluorescence (TIRF) microscopy illuminator for improved imaging of cell surface events,” in Current Protocols in Cytometry (John Wiley, 2012). [CrossRef]
  26. L. Wang, C. Vasilev, D. P. Canniffe, L. R. Wilson, C. N. Hunter, A. J. Cadby, “Highly confined surface imaging by solid immersion total internal reflection fluorescence microscopy,” Opt. Express 20, 3311–3324 (2012). [CrossRef] [PubMed]

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.


Fig. 1 Fig. 2 Fig. 3
Fig. 4

« Previous Article  |  Next Article »

OSA is a member of CrossRef.

CrossCheck Deposited