OSA's Digital Library

Biomedical Optics Express

Biomedical Optics Express

  • Editor: Joseph A. Izatt
  • Vol. 2, Iss. 1 — Jan. 1, 2011
  • pp: 149–158

Imaging carious dental tissues with multiphoton fluorescence lifetime imaging microscopy

Po-Yen Lin, Hong-Chou Lyu, Chin-Ying Stephen Hsu, Chia-Seng Chang, and Fu-Jen Kao  »View Author Affiliations

Biomedical Optics Express, Vol. 2, Issue 1, pp. 149-158 (2011)

View Full Text Article

Enhanced HTML    Acrobat PDF (2263 KB)

Browse Journals / Lookup Meetings

Browse by Journal and Year


Lookup Conference Papers

Close Browse Journals / Lookup Meetings

Article Tools



In this study, multiphoton excitation was utilized to image normal and carious dental tissues noninvasively. Unique structures in dental tissues were identified using the available multimodality (second harmonic, autofluorescence, and fluorescence lifetime analysis) without labeling. The collagen in dentin exhibits a strong second harmonic response. Both dentin and enamel emit strong autofluorescence that reveals in detail morphological features (such as dentinal tubules and enamel rods) and, despite their very similar spectral profiles, can be differentiated by lifetime analysis. Specifically, the carious dental tissue exhibits a greatly reduced autofluorescence lifetime, which result is consistent with the degree of demineralization, determined by micro-computed tomography. Our findings suggest that two-photon excited fluorescence lifetime imaging may be a promising tool for diagnosing and monitoring dental caries.

© 2010 OSA

OCIS Codes
(170.1850) Medical optics and biotechnology : Dentistry
(170.3880) Medical optics and biotechnology : Medical and biological imaging
(170.6920) Medical optics and biotechnology : Time-resolved imaging
(170.6935) Medical optics and biotechnology : Tissue characterization

ToC Category:
Dentistry Applications

Original Manuscript: November 2, 2010
Revised Manuscript: December 7, 2010
Manuscript Accepted: December 16, 2010
Published: December 17, 2010

Po-Yen Lin, Hong-Chou Lyu, Chin-Ying Stephen Hsu, Chia-Seng Chang, and Fu-Jen Kao, "Imaging carious dental tissues with multiphoton fluorescence lifetime imaging microscopy," Biomed. Opt. Express 2, 149-158 (2011)

Sort:  Author  |  Year  |  Journal  |  Reset  


  1. A. R. Ten Cate, Oral Histology 3rd ed. (Mosby, St. Louis, 1998), Chap. 5.
  2. F. J. Kao, “The use of optical parametric oscillator for harmonic generation and two-photon UV fluorescence microscopy,” Microsc. Res. Tech. 63(3), 175–181 (2004). [CrossRef] [PubMed]
  3. S. Y. Chen, C. Y. Hsu, and C. K. Sun, “Epi-third and second harmonic generation microscopic imaging of abnormal enamel,” Opt. Express 16(15), 11670–11679 (2008). [PubMed]
  4. L. Bachmann, D. M. Zezell, A. da Costa Ribeiro, L. Gomes, and A. S. Ito, “Fluorescence spectroscopy of biological tissue-A review,” Appl. Spectrosc. Rev. 41(6), 575–590 (2006). [CrossRef]
  5. A. Zoumi, A. Yeh, and B. J. Tromberg, “Imaging cells and extracellular matrix in vivo by using second-harmonic generation and two-photon excited fluorescence,” Proc. Natl. Acad. Sci. U.S.A. 99(17), 11014–11019 (2002). [CrossRef] [PubMed]
  6. W. R. Zipfel, R. M. Williams, R. Christie, A. Y. Nikitin, B. T. Hyman, and W. W. Webb, “Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation,” Proc. Natl. Acad. Sci. U.S.A. 100(12), 7075–7080 (2003). [CrossRef] [PubMed]
  7. P. J. Campagnola and L. M. Loew, “Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organisms,” Nat. Biotechnol. 21(11), 1356–1360 (2003). [CrossRef] [PubMed]
  8. P. J. Campagnola, A. C. Millard, M. Terasaki, P. E. Hoppe, C. J. Malone, and W. A. Mohler, “Three-dimensional high-resolution second-harmonic generation imaging of endogenous structural proteins in biological tissues,” Biophys. J. 82(1), 493–508 (2002). [CrossRef] [PubMed]
  9. M. H. Chen, W. L. Chen, Y. Sun, P. T. Fwu, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the tooth,” J. Biomed. Opt. 12(6), 064018 (2007). [CrossRef] [PubMed]
  10. K. Koenig and H. Schneckenburger, “Laser-induced autofluorescence for medical diagnosis,” J. Fluoresc. 4(1), 17–40 (1994). [CrossRef]
  11. V. V. Ghukasyan and F. J. Kao, “Monitoring cellular metabolism with fluorescence lifetime of reduced nicotinamide adnine dinucleotide,” J. Phys. Chem. 113, 11532–11549 (2009).
  12. K. Koenig, R. Hibst, G. Flemming, and H. Schneckenburger, “Laser-induced autofluorescence of caries,” Proc. SPIE 1880, 125–131 (1993). [CrossRef]
  13. R. Hibst, R. Paulus, and A. Lussi, “Detection of occlusal caries by laser fluorescence: basic and clinical investigations,” Med. Laser Appl. 16(3), 205–213 (2001). [CrossRef]
  14. E. Borisova, T. Uzunov, and L. Avramov, “Laser-induced autofluorescence study of caries model in vitro,” Lasers Med. Sci. 21(1), 34–41 (2006). [CrossRef] [PubMed]
  15. R. R. Gallagher, S. G. Demos, M. Balooch, G. W. Marshall, and S. J. Marshall, “Optical spectroscopy and imaging of the dentin-enamel junction in human third molars,” J. Biomed. Mater. Res. A 64A(2), 372–377 (2003). [CrossRef] [PubMed]
  16. W. Becker, A. Bergmann, M. A. Hink, K. König, K. Benndorf, and C. Biskup, “Fluorescence lifetime imaging by time-correlated single-photon counting,” Microsc. Res. Tech. 63(1), 58–66 (2004). [CrossRef] [PubMed]
  17. J. R. Lakowicz, Principles of fluorescence spectroscopy, 3rd edition. (New.York: Plenum Press, 2006)
  18. V. V. Ghukasyan, Y. Y. Hsu, S. H. Kung, and F. J. Kao, “Application of fluorescence resonance energy transfer resolved by fluorescence lifetime imaging microscopy for the detection of enterovirus 71 infection in cells,” J. Biomed. Opt. 12(2), 024016 (2007). [CrossRef] [PubMed]
  19. 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]
  20. K. M. Hanson, M. J. Behne, N. P. Barry, T. M. Mauro, E. Gratton, and R. M. Clegg, “Two-photon fluorescence lifetime imaging of the skin stratum corneum pH gradient,” Biophys. J. 83(3), 1682–1690 (2002). [CrossRef] [PubMed]
  21. H. C. Gerritsen, R. Sanders, A. Draaijer, C. Ince, and Y. K. Levine, “Fluorescence lifetime imaging of oxygen in living cells,” J. Fluoresc. 7(1), 11–15 (1997). [CrossRef]
  22. S. E. D. Webb, S. Leveque-Fort, D. S. Elson, J. Siegel, T. Watson, M. J. Lever, M. Booth, R. Juskaitis, M. A. A. Neil, L. O. Sucharov, T. Wilson, and P. M. W. French, “Wavelength-resolved 3-dimensional fluorescence lifetime imaging,” J. Fluoresc. 12(2), 279–283 (2002). [CrossRef]
  23. G. McConnell, J. M. Girkin, S. M. Ameer-Beg, P. R. Barber, B. Vojnovic, T. Ng, A. Banerjee, T. F. Watson, and R. J. Cook, “Time-correlated single-photon counting fluorescence lifetime confocal imaging of decayed and sound dental structures with a white-light supercontinuum source,” J. Microsc. 225(2), 126–136 (2007). [CrossRef] [PubMed]
  24. F. Ferretti de Oliveira, A. S. Ito, and L. Bachmann, “Time-resolved fluorescence spectroscopy of white-spot caries in human enamel,” Appl. Opt. 49(12), 2244–2249 (2010). [CrossRef] [PubMed]
  25. C. L. Darling and D. Fried, “Real-time near IR (1310 nm) imaging of CO2 laser ablation of enamel,” Opt. Express 16(4), 2685–2693 (2008). [CrossRef] [PubMed]
  26. G. K. Stookey, “Quantitative light fluorescence: a technology for early monitoring of the caries process,” Dent. Clin. North Am. 49(4), 753–770, vi (2005). [CrossRef] [PubMed]
  27. P. T. C. So, C. Y. Dong, B. R. Masters, and K. M. Berland, “Two-photon excitation fluorescence microscopy,” Annu. Rev. Biomed. Eng. 2(1), 399–429 (2000). [CrossRef] [PubMed]
  28. F. Helmchen and W. Denk, “Deep tissue two-photon microscopy,” Nat. Methods 2(12), 932–940 (2005). [CrossRef] [PubMed]
  29. U. Hafström-Björkman, F. Sundström, and J. J. ten Bosch, “Fluorescence in dissolved fractions of human enamel,” Acta Odontol. Scand. 49(3), 133–138 (1991). [CrossRef] [PubMed]
  30. W. Buchalla, “Comparative fluorescence spectroscopy shows differences in noncavitated enamel lesions,” Caries Res. 39(2), 150–156 (2005). [CrossRef] [PubMed]
  31. K. König, H. Schneckenburger, and R. Hibst, “Time-gated in vivo autofluorescence imaging of dental caries,” Cell. Mol. Biol. (Noisy-le-grand) 45(2), 233–239 (1999). [PubMed]
  32. A. Banerjee and A. Boyde, “Autofluorescence and mineral content of carious dentine: scanning optical and backscattered electron microscopic studies,” Caries Res. 32(3), 219–226 (1998). [CrossRef] [PubMed]
  33. W. Buchalla, “Comparative fluorescence spectroscopy shows differences in noncavitated enamel lesions,” Caries Res. 39(2), 150–156 (2005). [CrossRef] [PubMed]
  34. C. Robinson, R. C. Shore, S. J. Brookes, S. Strafford, S. R. Wood, and J. Kirkham, “The chemistry of enamel caries,” Crit. Rev. Oral Biol. Med. 11(4), 481–495 (2000). [CrossRef] [PubMed]
  35. M. V. Swain and J. Xue, “State of the art of Micro-CT applications in dental research,” Int. J. Oral Sci. 1(4), 177–188 (2009). [CrossRef] [PubMed]
  36. T. T. Huang, A. S. Jones, L. H. He, M. A. Darendeliler, and M. V. Swain, “Characterisation of enamel white spot lesions using X-ray micro-tomography,” J. Dent. 35(9), 737–743 (2007). [CrossRef] [PubMed]
  37. N. Efeoglu, D. Wood, and C. Efeoglu, “Microcomputerised tomography evaluation of 10% carbamide peroxide applied to enamel,” J. Dent. 33(7), 561–567 (2005). [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 Fig. 5

« Previous Article  |  Next Article »

OSA is a member of CrossRef.

CrossCheck Deposited