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Virtual Journal for Biomedical Optics

Virtual Journal for Biomedical Optics

| EXPLORING THE INTERFACE OF LIGHT AND BIOMEDICINE

  • Editor: Gregory W. Faris
  • Vol. 1, Iss. 7 — Jul. 17, 2006
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In vivo optical biopsy of hamster oral cavity with epi-third-harmonic-generation microscopy

Shih-Peng Tai, Wen-Jeng Lee, Dar-Bin Shieh, Ping-Ching Wu, Hsin-Yi Huang, Che-Hang Yu, and Chi-Kuang Sun  »View Author Affiliations


Optics Express, Vol. 14, Issue 13, pp. 6178-6187 (2006)
http://dx.doi.org/10.1364/OE.14.006178


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Abstract

The first in vivo optical virtual biopsy based on epi-third-harmonic-generation (THG) microscopy is successfully demonstrated using Syrian hamster oral mucosa as a model system. Without complex physical biopsy procedures, epi-THG microscopy can provide high spatial resolution dynamic images of oral mucosa and sub-mucosa in all three dimensions. The demonstrated intra-vital epi-THG microscopy provide high resolution observation of blood flow in the capillary and could be a promising tool to image angiogenesis, which is an important feature for many human diseases including malignancies. The system setup of epi-THG microscopy can be easily integrated with other nonlinear optical microscopy such as second-harmonic generation and multi-photon fluorescence microscopy by using the same laser system to provide better integrated molecular and structural information for future clinical diagnosis. By adding 6% acetic acid solution on the mucosa, THG contrast on the borders of nuclei was found to be greatly enhanced due to the alterations of their linear and nonlinear THG susceptibilities. With a virtual-transition-based technology without using fluorescence, the optical epi-THG biopsy we demonstrated shows promise for future noninvasive in vivo diseases examinations.

© 2006 Optical Society of America

1. Introduction

In clinical oral diseases diagnosis, physical biopsy is traditionally recognized as the final standard tool. However, physical biopsy requires the removal, fixation, and staining of tissues, cells, or fluids from the lesions of patients. Such histological procedures are not only time-consuming but also invasive and painful. In addition, traditional physical biopsy procedures per se may potentially put patients in the risk of spreading tumor cells. Moreover, unless tedious serial sectionings are performed, there is no guarantee for a missing diagnosis due to a local invasion not present in the given histological sectioning examined by the pathologists. Therefore, a non-invasive in vivo optical virtual biopsy, which can provide highly penetrative three-dimensional (3D) images with a sub-micron spatial resolution to assist the real physical biopsy, is highly desired.

Recently, third-harmonic-generation (THG) has been emerged as an important imaging modality in biological researches with the advantages including no energy release due to the characteristic of virtual-state-transition and intrinsic optical sectioning capability due to the nature of third-order nonlinearity. THG imaging has been applied to observe various structures in bio-tissues [1

1. D. Yelin and Y. Silberberg, “Laser scanning third-harmonic-generation microscopy in biology,” Opt. Express 5, 169–175 (1999. [CrossRef] [PubMed]

7

7. D. Debarre, W. Supatto, A.-M. Pena, A. Fabre, T. Tordjmann, L. Combettes, M.-C. Schanne-Klein, and E. Beaurepaire, “Imaging lipid bodies in cells and tissues using third-harmonic generation microscopy,” Nat. Methods 3, 47–53 (2006). [CrossRef]

] and embryos [8

8. C.-K. Sun, S.-W. Chu, S.-Y. Chen, T.-H. Tsai, T.-M. Liu, C.-Y. Lin, and H.-J. Tsai, “Higher harmonic generation microscopy for developmental biology,” J. Struct. Biol. 147, 19–30 (2004). [CrossRef] [PubMed]

], and to visualize intracellular Ca2+ dynamics [9

9. L. Canioni, S. Rivet, L. Sarger, R. Barille, P. Vacher, and P. Voisin, “Imaging of Ca2+ intracellular dynamics with a third-harmonic generation microscope,” Opt. Lett. 26, 515–517 (2001).). [CrossRef]

]. Although the third order nonlinearity exists theoretically in all materials, the Gouy phase shift effect substantially limits THG to be observed in the vicinity of interfaces where the first or the third order susceptibilities discontinue. Therefore, THG is an ideal optical biopsy tool to visualize the morphological structures in the bio-tissues because of its interface-sensitive nature.

In this paper, the first in vivo optical biopsy based on epi-THG microscopy is demonstrated by using live Syrian hamsters as an animal model before human subjects. We confirm the fact that abundant epi-THG signals can be collected from mucosa and sub-mucosa in the hamster oral cavity. Just like forward propagating THG, epi-THG can provide morphological images in both mucosa and sub-mucosa layers with a high spatial resolution, thus providing detailed histological information including the sizes, shapes, and distributions of basal cells. Besides, the movement of red blood cells in the capillary can also be clearly resolved by dynamic epi-THG microscopy. In order to study the origin of the collected epi-THG signals, acetic acid was added into the mucosa to enhance backward scatterings. Our study indicates that the contribution from the direct backward THG emissions should not be neglected, while enhanced epi-THG of nuclear borders can be clearly observed. With a virtual-transition-based technology without using fluorescence, the optical epi-THG biopsy we demonstrated shows promises for noninvasive in vivo diseases examinations. Combining with the acetic acid that serves as the THG contrast agent for nuclei, the functional epi-THG microscopy we developed is especially ideal for diagnosing those diseases associated with the nuclear morphology alterations.

2. Material and methods

Fig. 1. System setup of harmonics optical biopsy.
Fig. 2. Measured lateral resolution of in vivo epi-THG microscopy vs. depth in hamster oral cavity.

3. Results and discussion

3.1 Optical virtual biopsy of hamster oral cavity using epi-THG microscopy

To demonstrate the feasibility for future clinical diagnosis on oral diseases, Fig. 3 shows examples of the horizontally sectioned epi-SHG (green) and epi-THG (red) images taken from the hamster buccal pouch oral mucosa from surface deep into the sub-mucosa layer. From Fig. 3, the general histological structures in both mucosa and sub-mucosa layers can be identified through the THG modality due to its sensitivity to local optical inhomogeneities [1

1. D. Yelin and Y. Silberberg, “Laser scanning third-harmonic-generation microscopy in biology,” Opt. Express 5, 169–175 (1999. [CrossRef] [PubMed]

,11

11. G. Peleg, A. Lewis, M. Linial, and L. M. Loew “Nonlinear optical measurement of membrane potential around single molecules at selected celluar sites,” Proc. Natl. Acad. Sci. 96, 6700–6704 (1999). [CrossRef] [PubMed]

,12

12. J. A. Squier, M. Muller, G. J. Brakenhoff, and K. R. Wilson, “Third harmonic generation microscopy,” Opt. Express 3, 315–324 (1998). [CrossRef] [PubMed]

]. For example, the morphology of stratum corneum (Fig. 3(a), SB), stratum granulosa (Fig. 3(a), SG), stratum basale layer (Fig. 3(b), SB), and red blood cells (Fig. 3(b), RBC) in the capillary can all be easily picked up through the epi-THG microscopy. SHG, on the other hand, mainly reflects the distribution of connective tissues in the sub-mucosa [2

2. S.-W. Chu, I.-S. Chen, T.-M. Liu, C.-K. Sun, S.-P. Lee, B.-L. Lin, P.-C. Cheng, M.-X. Kuo, D.-J. Lin, and H.-L. Liu, “Nonlinear bio-photonic crystal effects revealed with multi-modal nonlinear microscopy,” J. Microsc. 208, Part 3, 190–200 (2002). [CrossRef] [PubMed]

, 13

13. I-H. Chen, S.-W. Chu, C.-K. Sun, P.-C. Cheng, and B.-L. Lin, “Wavelength dependent cell damages in biological multi-photon confocal microscopy: A micro-spectroscopic comparison between femtosecond Ti:sapphire and Cr:forsterite laser sources,” Opt. Quantum. Electron. 34, 1251–1266 (2002). [CrossRef]

, and 14

14. 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. 81, 493–508 (2002). [CrossRef]

] layer [represented in green color in a. 3(b)] such as lamina propria due to strong SHG generation in collagen fibers. It can be found that the epi-SHG signals are concentrated in the sub-mucosa and can provide information of the structure and distribution of collagen fibrils that may assist clinical diagnosis of connective tissue diseases.

Figures 3 (c) and 3(d) show more detailed microstructures of stratum granulosa and stratum basale provided by the epi-THG microscopy. Clear cell-cell junctions [indicated by yellow arrows in Fig. 3 (d)] between basal cells can be observed. Figure 4 is an example movie showing the blood flow in the capillary imaged by the epi-THG microscopy. The movement of RBCs in the capillary can be clearly resolved. Therefore, epi-THG microscopy can not only provide structure contours and histological information in both mucosa and sub-mucosa but also can monitor the capillary network and the blood flows. This observation indicates that epi-THG microscopy could be an ideal tool to observe angiogenesis, which is not only one of the early signs of caners but also many other important human diseases.

Fig. 3. In vivo horizontally sectioned epi-SHG (green) and epi-THG (red) images of hamster mucosa and sub-mucosa taken at different depths. From (a) and (b), we can identify the morphology of stratum corneum (SC), stratum granulosa (SG), stratum basale (SB), and red blood cells (RBC) in the oral cavity by using epi-THG microscopy. Besides, epi-SHG microscopy can provide the distribution and density information of collagen fibrils in the sub-mucosa. At higher magnification, (c) and (d) show more detailed morphologies of stratum granulosa and stratum basale. Clear cell-cell junctions (indicated by yellow arrows) can be picked up by epi-THG microscopy. Scale bar: 40µm.
Fig. 4. (1.67 MB) An example movie of the in vivo horizontally sectioned epi-THG microscopy showing the blood flow in the capillary in the sub-mucosa layer of the hamster oral cavity. Image size: 80µm×80µm

Figure 5 is an example movie of a sequential set of horizontally sectioned images taken by epi-THG and epi-SHG microscopes. The movie is composed of 30 horizontal sections and the depth difference between two adjacent images is 2.8-µm. Different profiles between two adjacent images can be found. Therefore, this optical biopsy technique enables direct acquisition of high resolution 3-D structural images of the oral cavity and provides an ideal new platform for clinical optical virtual biopsy in the future without using fluorescence and confocal pinhole. The current scanning rate of the system is limited by the speed of the adopted Fluoview300 galvo-mirrors, and real-time epi-THG microscopy can be achieved by using high speed scanners such as polygon mirror scanner [15

15. K. H. Kim, C. Buehler, and P. T. C. So, “High speed, two-photon scanning microscope,” Appl. Opt. 38, 6004–6009 (1999). [CrossRef]

] to meet the requirements of clinical diagnosis in the future.

Fig. 5. (1.85 MB) This movie shows a stack of depth-resolved in vivo horizontal sections in the hamster oral cavity. Using epi-SHG and epi-THG microscopy, a series of horizontal sections from mucosa to sub-mucosa is demonstrated. This movie is composed of 30 horizontal images. The optical depth difference between adjacent images is 2.8 µm. Image size: 240µm×240µm

3.2 Acetic acid as contrast agent of epi-THG microscopy

Fig. 6. Vertical sections taken in the live hamster oral cavity by epi-THG microscopy (a) before and (b) after adding 6% acetic acid solution on mucosa. After adding 6% acetic acid solution, the enhanced epi-THG intensity of epithelial cells, except stratum corneum (SC), can be observed. Scale bar: 30µm
Fig. 7. In vivo horizontal sections taken in the mucosa layer of the live hamster oral cavity at different depths by epi-THG microscopy are presented after adding 6% acetic acid solution. The nuclear morphology of squamous cells can be clearly observed by epi-THG microscopy (indicated by green arrow). The yellow arrow indicates the cell membrane. Scale bar: 10µm

4. Conclusion

The first in vivo virtual biopsy based on epi-THG microscopy is demonstrated using Syrian hamster oral mucosa as a model system. This optical virtual biopsy technique does not require invasive fluorescence staining and the harmonic generation processes leave no energy deposition to the examined tissues. Without complex physical biopsy procedures, it can provide high spatial resolution images of oral mucosa and sub-mucosa in all three dimensions. The demonstrated intra-vital epi-THG microscopy provide high resolution observation of blood flow in the capillary and should be a promising tool to image angiogenesis, which is an important feature of many human diseases including malignancies. The system setup of epi-THG microscopy can be easily integrated with other nonlinear optical microscopy such as SHG and multi-photon fluorescence microscopy by using the same laser system to provide better integrated molecular and structural information for future clinical diagnosis. By applying 6% acetic acid solution on the oral mucosa, enhanced epi-THG contrast on the borders of nuclei can be observed due to the alterations of their linear and nonlinear THG susceptibilities. Our study also supports the fact that direct-backward THG is one of the dominant signals we collected. With a virtual-transition-based technology without using fluorescence, the optical epi-THG biopsy we demonstrated shows a promising future for noninvasive in vivo diseases examinations. Combined with the acetic acid that serves as the THG contrast agent for the nuclei, our developed functional epi-THG is especially ideal for the differential diagnosis of diseases associated with nuclear morphology.

Acknowledgment

The authors gratefully acknowledge financial support from the National Health Research Institute (NHRI-EX95-9201EI) of Taiwan, National Science Council of Taiwan (NSC 94-2314-B-006-067, NSC 94-2120-M-006-004) and the National Taiwan University Center for Genomic Medicine.

References and links

1.

D. Yelin and Y. Silberberg, “Laser scanning third-harmonic-generation microscopy in biology,” Opt. Express 5, 169–175 (1999. [CrossRef] [PubMed]

2.

S.-W. Chu, I.-S. Chen, T.-M. Liu, C.-K. Sun, S.-P. Lee, B.-L. Lin, P.-C. Cheng, M.-X. Kuo, D.-J. Lin, and H.-L. Liu, “Nonlinear bio-photonic crystal effects revealed with multi-modal nonlinear microscopy,” J. Microsc. 208, Part 3, 190–200 (2002). [CrossRef] [PubMed]

3.

C.-K. Sun, C.-C. Chen, S.-W. Chu, T.-H. Tsai, Y.-C. Chen, and B.-L. Lin, “Multi-harmonic-generation biopsy of skin,” Opt. Lett. 28, 2488–2490 (2003). [CrossRef] [PubMed]

4.

S.-P. Tai, T.-H. Tsai, W.-J. Lee, D.-B. Shieh, Y.-H. Liao, H.-Y. Huang, K. Y.-J. Zhang, H.-L. Liu, and C.-K. Sun, “Optical biopsy of fixed human skin with backward-collected optical harmonics signals,” Opt. Express 13, 8231–8242 (2005). [CrossRef] [PubMed]

5.

T.-H Tsai, S.-P. Tai, W.-J. Lee, H.-Y. Huang, Y.-H. Liao, and C.-K. Sun, “Optical signal degradation study in fixed human skin using confocal microscopy and higher-harmonic optical microscopy,” Opt. Express 14, 749–758 (2006). [CrossRef] [PubMed]

6.

V. Barzda, C. Greenhalgh, J. Aus der Au, S. Elmore, J. H. van Beek, and J. Squier, “Visualization of mitochondria in cardiomyocytes by simultaneous harmonic generation andfluorescence microscopy,” Opt. Express 13, 8263–8276 (2005). [CrossRef] [PubMed]

7.

D. Debarre, W. Supatto, A.-M. Pena, A. Fabre, T. Tordjmann, L. Combettes, M.-C. Schanne-Klein, and E. Beaurepaire, “Imaging lipid bodies in cells and tissues using third-harmonic generation microscopy,” Nat. Methods 3, 47–53 (2006). [CrossRef]

8.

C.-K. Sun, S.-W. Chu, S.-Y. Chen, T.-H. Tsai, T.-M. Liu, C.-Y. Lin, and H.-J. Tsai, “Higher harmonic generation microscopy for developmental biology,” J. Struct. Biol. 147, 19–30 (2004). [CrossRef] [PubMed]

9.

L. Canioni, S. Rivet, L. Sarger, R. Barille, P. Vacher, and P. Voisin, “Imaging of Ca2+ intracellular dynamics with a third-harmonic generation microscope,” Opt. Lett. 26, 515–517 (2001).). [CrossRef]

10.

T.-M. Liu, S.-W Chiu, C.-K Sun, B.-L. Lin, P. C. Cheng, and I. Johnson, “Multi-photon scanning microscopy using a femtosecond Cr:forsterite laser,” Scanning 23, 249–254 (2001). [CrossRef] [PubMed]

11.

G. Peleg, A. Lewis, M. Linial, and L. M. Loew “Nonlinear optical measurement of membrane potential around single molecules at selected celluar sites,” Proc. Natl. Acad. Sci. 96, 6700–6704 (1999). [CrossRef] [PubMed]

12.

J. A. Squier, M. Muller, G. J. Brakenhoff, and K. R. Wilson, “Third harmonic generation microscopy,” Opt. Express 3, 315–324 (1998). [CrossRef] [PubMed]

13.

I-H. Chen, S.-W. Chu, C.-K. Sun, P.-C. Cheng, and B.-L. Lin, “Wavelength dependent cell damages in biological multi-photon confocal microscopy: A micro-spectroscopic comparison between femtosecond Ti:sapphire and Cr:forsterite laser sources,” Opt. Quantum. Electron. 34, 1251–1266 (2002). [CrossRef]

14.

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. 81, 493–508 (2002). [CrossRef]

15.

K. H. Kim, C. Buehler, and P. T. C. So, “High speed, two-photon scanning microscope,” Appl. Opt. 38, 6004–6009 (1999). [CrossRef]

16.

J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, “An epi-detected coherent anti-stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity,” J. Phys. Chem. B 105, 1277–1280 (2001). [CrossRef]

17.

A. Volkmer, J. X. Cheng, and X. S. Xie, “Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy,” Phys. Rev. Lett. 87, 023901 (2001). [CrossRef]

18.

J. X. Cheng, A. Volkmer, and X. S. Xie, “Theoretical and experimental characterization of coherent anti-Stokes Raman scattering microscopy,” J. Opt. Soc. Am. B 19, 1363–1375 (2002). [CrossRef]

19.

A. Volkmer, “Vibrational imaging and microspectroscopies based on coherent anti-Stoke Raman scattering microscopy,” J. Phys. D: Appl. Phys. 38, R59–R81 (2005). [CrossRef]

20.

J. X. Cheng and X. S. Xie, “Green’s function formulation for third-harmonic generation microscopy,” J. Opt. Soc. Am. B 19, 1604–1610 (2002). [CrossRef]

21.

S.-Y. Chen, S.-P. Tai, T.-H. Tsai, and C.-K. Sun, “Direct backward-emitted third-harmonic generation and its application to clinical microscopy”, in Technical Digest of Conference on Laser and Electro-optics/Quantum Electronics and Laser Science Conference (CLEO/QELS 2005), Baltimore, MD, USA, paper QMI3 (2005).

22.

D. W. Fawcett, An Atlas of Fine Structure, The cell: Its organelles and inclusions, (Saunders, Philadelphia1996).

23.

L. T. Perelman and V. Backman, “Light scattering spectroscopy of epithelial tissue: Principles and applications,” Handbook of Optical Biomedical Diagnostics, (SPIE, Bellingham, WA2002) Chap. 12, 675–724.

24.

L. Burke, D. Antonioli, and B. Durcatman, Colposcopy Text and Atlas, (Appleton and Lange., 1991).

25.

T. Collier, P. Shen, B. de Pradier, K. -B. Sung, R. Richards-Kortum, M. Follen, and A. Malpica, “Near real time confocal microscopy of amelanotic tissue: Dynamics of aceto-whitening enable nuclear segmentation,” Opt. Express 6, 40–48 (2000). [CrossRef] [PubMed]

26.

R. A. Drezek, T. Collier, C. K. Brookner, A. Malpica, R. Lotan, R. R. Richards-Kortum, and M. Follen, “Laser scanning confocal microscopy of cervical tissue before and after application of acetic acid,” Am. J. Obstet. Gynecol. 182, 1135–1139 (2000). [CrossRef] [PubMed]

27.

M. Rajadhyaksha, G. Menaker, T. Flotte, P. J. Dwyer, and S. Gonzalez, “Confocal examinazation of nonmelanoma cancers in thick skin excisions to potentially guide Mohs micrographic surgery without frozen histopathology,” J. Invest. Dermatol. 117, 1137–1143 (2001). [CrossRef] [PubMed]

28.

A. A. Lacy, T. Collier, J. E. Price, S. Dharmawardhane, and R. R. Richards-Kortum, “Near real-time in vivo confocal imaging of mouse mammary tumors,” Front. Biosci. 7, F1–F7 (2002). [CrossRef] [PubMed]

29.

A. F. Zuluaga, R. Drzek, T. Collier, R. Lotan, and M. Follen, “Contrast agents for confocal microscopy: how simple chemicals affect confocal images of normal and cancer cells in suspension,” J. of Biomed. Opt. 7, 398–403 (2002). [CrossRef]

30.

A. N. Yaroslavsky, J. Barbosa, V. Neel, C. DiMarzio, and R. R. Anderson, “Combining multispectral polarized light imaging and confocal microscopy for localization of nonmelanoma skin cancer,” J. of Biomed. Opt. 10, 014011 (2005). [CrossRef]

31.

K. Carlson, I. Pavlova, M. Descour, M. Follen, and R. R. -Kortum, “Confocal microscopy: Imaging cervical precancerous lesions,” Gynecol. Oncol. 99, S84–S88 (2005). [CrossRef] [PubMed]

32.

C.-C. ChangChien, H. Lin, S. W. Leung, C.-Y. Hsu, and C.-L. Cho, “Effect of acetic acid on telomerse acitivity in cervical intraepithelial neoplasia,” Gynecol. Oncol. 71, 99–103 (1998). [CrossRef] [PubMed]

33.

B. W. Pogue, H. B. Kaufman, A. Zelnechuk, W. Harper, G. C. Burke, E. E. Burke, and D. M. Harper, “Analysis of acetic acid-induced whitening of high-grade squamous intraepithelial lesions,” J. of Biomed. Opt. 6, 397–403 (2001). [CrossRef]

34.

R. Lambert, J. F. Rey, and R. Sankaranarayanan, “Magnification and chromoscopy with the acetic cid test,” Endoscopy 35, 437–445, (2003). [CrossRef] [PubMed]

35.

M. Rajadhyaksha, S. Gonzalez, and J. M. Zavislan, “Detectability of contrast agents for confocal reflectance imaging of skin and microcirculation,” J. of Biomed. Opt. 9, 323–331 (2002). [CrossRef]

36.

A. B. MacLean, “Acetowhite epithelium,” Gynecol. Oncol. 95, 691–694 (2004). [CrossRef] [PubMed]

37.

J. F. Rey, H. Inoue, and M. Guelrud, “Magnification endoscopy with acetic acid for Barrett’s esophagus,” Endoscopy 37, 583–586, (2005). [CrossRef] [PubMed]

38.

W.-J. Lee, C.-H. Yu, S-P. Tai, H.-Y. Huang, and C.-K. Sun, “Functional third-harmonic generation microscopy of cell nuclei by using acetic acid as a contrast agent,” in preparation.

OCIS Codes
(180.5810) Microscopy : Scanning microscopy
(180.6900) Microscopy : Three-dimensional microscopy
(190.4160) Nonlinear optics : Multiharmonic generation

ToC Category:
Microscopy

History
Original Manuscript: May 15, 2006
Revised Manuscript: June 6, 2006
Manuscript Accepted: June 8, 2006
Published: June 26, 2006

Virtual Issues
Vol. 1, Iss. 7 Virtual Journal for Biomedical Optics

Citation
Shih-Peng Tai, Wen-Jeng Lee, Dar-Bin Shieh, Ping-Ching Wu, Hsin-Yi Huang, Che-Hang Yu, and Chi-Kuang Sun, "In vivo optical biopsy of hamster oral cavity with epi-third-harmonic-generation microscopy," Opt. Express 14, 6178-6187 (2006)
http://www.opticsinfobase.org/vjbo/abstract.cfm?URI=oe-14-13-6178


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References

  1. D. Yelin and Y. Silberberg, "Laser scanning third-harmonic-generation microscopy in biology," Opt. Express 5, 169-175 (1999. [CrossRef] [PubMed]
  2. S.-W. Chu, I.-S. Chen, T.-M. Liu, C.-K. Sun, S.-P. Lee, B.-L. Lin, P.-C. Cheng, M.-X. Kuo, D.-J. Lin, and H.-L. Liu, "Nonlinear bio-photonic crystal effects revealed with multi-modal nonlinear microscopy," J. Microsc. 208, Part 3, 190-200 (2002). [CrossRef] [PubMed]
  3. C.-K. Sun, C.-C. Chen, S.-W. Chu, T.-H. Tsai, Y.-C. Chen and B.-L. Lin, "Multi-harmonic-generation biopsy of skin," Opt. Lett. 28, 2488-2490 (2003). [CrossRef] [PubMed]
  4. S.-P. Tai, T.-H. Tsai, W.-J. Lee, D.-B. Shieh, Y.-H. Liao, H.-Y. Huang, K. Y.-J. Zhang, H.-L. Liu, and C.-K. Sun, "Optical biopsy of fixed human skin with backward-collected optical harmonics signals," Opt. Express 13, 8231-8242 (2005). [CrossRef] [PubMed]
  5. T.-H Tsai, S.-P. Tai, W.-J. Lee, H.-Y. Huang, Y.-H. Liao and C.-K. Sun, "Optical signal degradation study in fixed human skin using confocal microscopy and higher-harmonic optical microscopy," Opt. Express 14, 749-758 (2006). [CrossRef] [PubMed]
  6. V. Barzda, C. Greenhalgh, J. Aus der Au, S. Elmore, J. H. van Beek, and J. Squier, "Visualization of mitochondria in cardiomyocytes by simultaneous harmonic generation andfluorescence microscopy," Opt. Express 13, 8263-8276 (2005). [CrossRef] [PubMed]
  7. D. Debarre, W. Supatto, A.-M. Pena, A. Fabre, T. Tordjmann, L. Combettes, M.-C. Schanne-Klein, and E. Beaurepaire, "Imaging lipid bodies in cells and tissues using third-harmonic generation microscopy," Nat. Methods 3, 47-53 (2006). [CrossRef]
  8. C.-K. Sun, S.-W. Chu, S.-Y. Chen, T.-H. Tsai, T.-M. Liu, C.-Y. Lin, and H.-J. Tsai, "Higher harmonic generation microscopy for developmental biology," J. Struct. Biol. 147, 19-30 (2004). [CrossRef] [PubMed]
  9. L. Canioni, S. Rivet, L. Sarger, R. Barille, P. Vacher, and P. Voisin, "Imaging of Ca2+ intracellular dynamics with a third-harmonic generation microscope," Opt. Lett. 26, 515-517 (2001).). [CrossRef]
  10. T.-M. Liu, S.-W Chiu, C.-K Sun, B.-L. Lin, P. C. Cheng, and I. Johnson, "Multi-photon scanning microscopy using a femtosecond Cr:forsterite laser," Scanning 23, 249-254 (2001). [CrossRef] [PubMed]
  11. G. Peleg, A. Lewis, M. Linial, and L. M. Loew "Nonlinear optical measurement of membrane potential around single molecules at selected celluar sites," Proc. Natl. Acad. Sci. 96, 6700-6704 (1999). [CrossRef] [PubMed]
  12. J. A. Squier, M. Muller, G. J. Brakenhoff, and K. R. Wilson, "Third harmonic generation microscopy," Opt. Express 3, 315-324 (1998). [CrossRef] [PubMed]
  13. I-H. Chen, S.-W. Chu, C.-K. Sun, P.-C. Cheng, and B.-L. Lin, "Wavelength dependent cell damages in biological multi-photon confocal microscopy: A micro-spectroscopic comparison between femtosecond Ti:sapphire and Cr:forsterite laser sources," Opt. Quantum. Electron. 34, 1251-1266 (2002). [CrossRef]
  14. 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. 81, 493-508 (2002). [CrossRef]
  15. K. H. Kim, C. Buehler, and P. T. C. So, "High speed, two-photon scanning microscope," Appl. Opt. 38, 6004-6009 (1999). [CrossRef]
  16. J. X. Cheng, A. Volkmer, L. D. Book, and X. S. Xie, "An epi-detected coherent anti-stokes Raman scattering (E-CARS) microscope with high spectral resolution and high sensitivity," J. Phys. Chem. B 105, 1277-1280 (2001). [CrossRef]
  17. A. Volkmer, J. X. Cheng, and X. S. Xie, "Vibrational imaging with high sensitivity via epidetected coherent anti-Stokes Raman scattering microscopy," Phys. Rev. Lett. 87, 023901 (2001). [CrossRef]
  18. J. X. Cheng, A. Volkmer, and X. S. Xie, "Theoretical and experimental characterization of coherent anti-Stokes Raman scattering microscopy," J. Opt. Soc. Am. B 19, 1363-1375 (2002). [CrossRef]
  19. A. Volkmer, "Vibrational imaging and microspectroscopies based on coherent anti-Stoke Raman scattering microscopy," J. Phys. D: Appl. Phys. 38, R59-R81 (2005). [CrossRef]
  20. J. X. Cheng, and X. S. Xie, "Green's function formulation for third-harmonic generation microscopy," J. Opt. Soc. Am. B 19, 1604-1610 (2002). [CrossRef]
  21. S.-Y. Chen, S.-P. Tai, T.-H. Tsai and C.-K. Sun, "Direct backward-emitted third-harmonic generation and its application to clinical microscopy", in Technical Digest of Conference on Laser and Electro-optics/Quantum Electronics and Laser Science Conference (CLEO/QELS 2005), Baltimore, MD, USA, paper QMI3 (2005).
  22. D. W. Fawcett, An Atlas of Fine Structure, The cell: Its organelles and inclusions, (Saunders, Philadelphia 1996).
  23. L. T. Perelman, and V. Backman, "Light scattering spectroscopy of epithelial tissue: Principles and applications," Handbook of Optical Biomedical Diagnostics, (SPIE, Bellingham, WA 2002) Chap. 12, 675-724.
  24. L. Burke, D. Antonioli, B. Durcatman, Colposcopy Text and Atlas, (Appleton and Lange., 1991).
  25. T. Collier, P. Shen, B. de Pradier, K. -B. Sung, R. Richards-Kortum, M. Follen, and A. Malpica, "Near real time confocal microscopy of amelanotic tissue: Dynamics of aceto-whitening enable nuclear segmentation," Opt. Express 6, 40-48 (2000). [CrossRef] [PubMed]
  26. R. A. Drezek, T. Collier, C. K. Brookner, A. Malpica, R. Lotan, R. R. Richards-Kortum, and M. Follen, "Laser scanning confocal microscopy of cervical tissue before and after application of acetic acid," Am. J. Obstet. Gynecol. 182, 1135-1139 (2000). [CrossRef] [PubMed]
  27. M. Rajadhyaksha, G. Menaker, T. Flotte, P. J. Dwyer, and S. Gonzalez, "Confocal examinazation of nonmelanoma cancers in thick skin excisions to potentially guide Mohs micrographic surgery without frozen histopathology," J. Invest. Dermatol. 117,1137-1143 (2001). [CrossRef] [PubMed]
  28. A. A. Lacy, T. Collier, J. E. Price, S. Dharmawardhane and R. R. Richards-Kortum, "Near real-time in vivo confocal imaging of mouse mammary tumors," Front. Biosci. 7, F1-F7 (2002). [CrossRef] [PubMed]
  29. A. F. Zuluaga, R. Drzek, T. Collier, R. Lotan, and M. Follen, "Contrast agents for confocal microscopy: how simple chemicals affect confocal images of normal and cancer cells in suspension," J. of Biomed. Opt. 7, 398-403 (2002). [CrossRef]
  30. A. N. Yaroslavsky, J. Barbosa, V. Neel, C. DiMarzio, and R. R. Anderson, "Combining multispectral polarized light imaging and confocal microscopy for localization of nonmelanoma skin cancer," J. of Biomed. Opt. 10, 014011 (2005). [CrossRef]
  31. K. Carlson, I. Pavlova, M. Descour. M. Follen, and R. R.-Kortum, "Confocal microscopy: Imaging cervical precancerous lesions, " Gynecol. Oncol. 99, S84-S88 (2005). [CrossRef] [PubMed]
  32. C.-C. ChangChien, H. Lin, S. W. Leung, C.-Y. Hsu, and C.-L. Cho, "Effect of acetic acid on telomerse acitivity in cervical intraepithelial neoplasia," Gynecol. Oncol. 71, 99-103 (1998). [CrossRef] [PubMed]
  33. B. W. Pogue, H. B. Kaufman, A. Zelnechuk, W. Harper, G. C. Burke, E. E. Burke, and D. M. Harper, "Analysis of acetic acid-induced whitening of high-grade squamous intraepithelial lesions," J. Biomed. Opt. 6, 397-403 (2001). [CrossRef]
  34. R. Lambert, J. F. Rey, and R. Sankaranarayanan, "Magnification and chromoscopy with the acetic cid test," Endoscopy 35, 437-445, (2003). [CrossRef] [PubMed]
  35. M. Rajadhyaksha, S. Gonzalez, J. M. Zavislan, "Detectability of contrast agents for confocal reflectance imaging of skin and microcirculation," J. of Biomed. Opt. 9, 323-331 (2002). [CrossRef]
  36. A. B. MacLean, "Acetowhite epithelium," Gynecol. Oncol. 95, 691-694 (2004). [CrossRef] [PubMed]
  37. J. F. Rey, H. Inoue, and M. Guelrud, "Magnification endoscopy with acetic acid for Barrett’s esophagus," Endoscopy 37, 583-586, (2005). [CrossRef] [PubMed]
  38. W.-J. Lee, C.-H. Yu, S-P. Tai, H.-Y. Huang, and C.-K. Sun, "Functional third-harmonic generation microscopy of cell nuclei by using acetic acid as a contrast agent," in preparation.

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