Abstract

Fungal keratitis is an infection of the cornea by fungal pathogens. Diagnosis methods based on optical microscopy could be beneficial over the conventional microbiology method by allowing rapid and non-invasive examination. Reflectance confocal microscopy (RCM) and two-photon second harmonic generation microscopy (TPSHGM) have been applied to pre-clinical or clinical studies of fungal keratitis. In this report, RCM and TPSHGM were characterized and compared in the imaging of a fungal keratitis rabbit model ex vivo. Fungal infection was induced by using two strains of fungi: aspergillus fumigatus and candida albicans. The infected corneas were imaged in fresh condition by both modalities sequentially and their images were analyzed. Both RCM and TPSHGM could detect both fungal strains within the cornea based on morphology: aspergillus fumigatus had distinctive filamentous structures, and candida albicans had round structures superficially and elongated structures in the corneal stroma. These imaging results were confirmed by histology. Comparison between RCM and TPSHGM showed several characteristics. Although RCM and TPSHGM images had good correlation each other, their images were slightly different due to difference in contrast mechanism. RCM had relatively low image contrast with the infected turbid corneas due to high background signal. TPSHGM visualized cells and collagen in the cornea clearly compared to RCM, but used higher laser power to compensate low autofluorescence. Since these two modalities provide complementary information, combination of RCM and TPSHGM would be useful for fungal keratitis detection by compensating their weaknesses each other.

© 2016 Optical Society of America

Full Article  |  PDF Article
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References

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    [Crossref] [PubMed]

2015 (1)

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

2014 (2)

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

2013 (1)

2011 (3)

P. Steven, F. Bock, G. Hüttmann, and C. Cursiefen, “Intravital two-photon microscopy of immune cell dynamics in corneal lymphatic vessels,” PLoS One 6(10), e26253 (2011).
[Crossref] [PubMed]

E. A. Gibson, O. Masihzadeh, T. C. Lei, D. A. Ammar, and M. Y. Kahook, “Multiphoton microscopy for ophthalmic imaging,” J. Ophthalmol. 2011, 870879 (2011).
[Crossref] [PubMed]

E. C. Ledbetter, N. L. Irby, and S. G. Kim, “In vivo confocal microscopy of equine fungal keratitis,” Vet. Ophthalmol. 14(1), 1–9 (2011).
[Crossref] [PubMed]

2010 (2)

R. L. Niederer and C. N. McGhee, “Clinical in vivo confocal microscopy of the human cornea in health and disease,” Prog. Retin. Eye Res. 29(1), 30–58 (2010).
[Crossref] [PubMed]

D. Mantopoulos, A. Cruzat, and P. Hamrah, “In vivo imaging of corneal inflammation: new tools for clinical practice and research,” Semin. Ophthalmol. 25(5-6), 178–185 (2010).
[Crossref] [PubMed]

2009 (2)

U. Jurkunas, I. Behlau, and K. Colby, “Fungal keratitis: changing pathogens and risk factors,” Cornea 28(6), 638–643 (2009).
[Crossref] [PubMed]

P. Steven, M. Müller, N. Koop, C. Rose, and G. Hüttmann, “Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies,” J. Biomed. Opt. 14(6), 064040 (2009).
[Crossref] [PubMed]

2008 (1)

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. Tech. 71(2), 83–85 (2008).
[Crossref] [PubMed]

2007 (4)

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

P. A. Thomas and P. Geraldine, “Infectious keratitis,” Curr. Opin. Infect. Dis. 20(2), 129–141 (2007).
[Crossref] [PubMed]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

2006 (1)

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

2003 (2)

W. R. Zipfel, R. M. Williams, and W. W. Webb, “Nonlinear magic: multiphoton microscopy in the biosciences,” Nat. Biotechnol. 21(11), 1369–1377 (2003).
[Crossref] [PubMed]

A. M. Avunduk, R. W. Beuerman, E. D. Varnell, and H. E. Kaufman, “Confocal microscopy of Aspergillus fumigatus keratitis,” Br. J. Ophthalmol. 87(4), 409–410 (2003).
[Crossref] [PubMed]

2001 (1)

J. P. Whitcher, M. Srinivasan, and M. P. Upadhyay, “Corneal blindness: a global perspective,” Bull. World Health Organ. 79(3), 214–221 (2001).
[PubMed]

2000 (1)

K. König, “Multiphoton microscopy in life sciences,” J. Microsc. 200(2), 83–104 (2000).
[Crossref] [PubMed]

1990 (1)

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248(4951), 73–76 (1990).
[Crossref] [PubMed]

Ammar, D. A.

E. A. Gibson, O. Masihzadeh, T. C. Lei, D. A. Ammar, and M. Y. Kahook, “Multiphoton microscopy for ophthalmic imaging,” J. Ophthalmol. 2011, 870879 (2011).
[Crossref] [PubMed]

Avunduk, A. M.

A. M. Avunduk, R. W. Beuerman, E. D. Varnell, and H. E. Kaufman, “Confocal microscopy of Aspergillus fumigatus keratitis,” Br. J. Ophthalmol. 87(4), 409–410 (2003).
[Crossref] [PubMed]

Azumi, H.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Baudouin, C.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Behlau, I.

U. Jurkunas, I. Behlau, and K. Colby, “Fungal keratitis: changing pathogens and risk factors,” Cornea 28(6), 638–643 (2009).
[Crossref] [PubMed]

Beuerman, R. W.

A. M. Avunduk, R. W. Beuerman, E. D. Varnell, and H. E. Kaufman, “Confocal microscopy of Aspergillus fumigatus keratitis,” Br. J. Ophthalmol. 87(4), 409–410 (2003).
[Crossref] [PubMed]

Bock, F.

P. Steven, F. Bock, G. Hüttmann, and C. Cursiefen, “Intravital two-photon microscopy of immune cell dynamics in corneal lymphatic vessels,” PLoS One 6(10), e26253 (2011).
[Crossref] [PubMed]

Borderie, V.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Bourcier, T.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Brasnu, E.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Brown, D. J.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Chen, W. L.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. Tech. 71(2), 83–85 (2008).
[Crossref] [PubMed]

Chen, Y. F.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

Chikama, T.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Colby, K.

U. Jurkunas, I. Behlau, and K. Colby, “Fungal keratitis: changing pathogens and risk factors,” Cornea 28(6), 638–643 (2009).
[Crossref] [PubMed]

Cruzat, A.

D. Mantopoulos, A. Cruzat, and P. Hamrah, “In vivo imaging of corneal inflammation: new tools for clinical practice and research,” Semin. Ophthalmol. 25(5-6), 178–185 (2010).
[Crossref] [PubMed]

Cursiefen, C.

P. Steven, F. Bock, G. Hüttmann, and C. Cursiefen, “Intravital two-photon microscopy of immune cell dynamics in corneal lymphatic vessels,” PLoS One 6(10), e26253 (2011).
[Crossref] [PubMed]

Degorge, S.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Denk, W.

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248(4951), 73–76 (1990).
[Crossref] [PubMed]

Dogru, M.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Dong, C. Y.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. Tech. 71(2), 83–85 (2008).
[Crossref] [PubMed]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Dupas, B.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Efron, N.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Geng, Y.

Geraldine, P.

P. A. Thomas and P. Geraldine, “Infectious keratitis,” Curr. Opin. Infect. Dis. 20(2), 129–141 (2007).
[Crossref] [PubMed]

Gho, Y. S.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Gibson, E. A.

E. A. Gibson, O. Masihzadeh, T. C. Lei, D. A. Ammar, and M. Y. Kahook, “Multiphoton microscopy for ophthalmic imaging,” J. Ophthalmol. 2011, 870879 (2011).
[Crossref] [PubMed]

Hamrah, P.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

D. Mantopoulos, A. Cruzat, and P. Hamrah, “In vivo imaging of corneal inflammation: new tools for clinical practice and research,” Semin. Ophthalmol. 25(5-6), 178–185 (2010).
[Crossref] [PubMed]

Hsiao, C. H.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

Huang, S. C.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

Hunter, J. J.

Hüttmann, G.

P. Steven, F. Bock, G. Hüttmann, and C. Cursiefen, “Intravital two-photon microscopy of immune cell dynamics in corneal lymphatic vessels,” PLoS One 6(10), e26253 (2011).
[Crossref] [PubMed]

P. Steven, M. Müller, N. Koop, C. Rose, and G. Hüttmann, “Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies,” J. Biomed. Opt. 14(6), 064040 (2009).
[Crossref] [PubMed]

Irby, N. L.

E. C. Ledbetter, N. L. Irby, and S. G. Kim, “In vivo confocal microscopy of equine fungal keratitis,” Vet. Ophthalmol. 14(1), 1–9 (2011).
[Crossref] [PubMed]

Jee, S. H.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Jester, J. V.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Jurkunas, U.

U. Jurkunas, I. Behlau, and K. Colby, “Fungal keratitis: changing pathogens and risk factors,” Cornea 28(6), 638–643 (2009).
[Crossref] [PubMed]

Kahook, M. Y.

E. A. Gibson, O. Masihzadeh, T. C. Lei, D. A. Ammar, and M. Y. Kahook, “Multiphoton microscopy for ophthalmic imaging,” J. Ophthalmol. 2011, 870879 (2011).
[Crossref] [PubMed]

Kaufman, H. E.

A. M. Avunduk, R. W. Beuerman, E. D. Varnell, and H. E. Kaufman, “Confocal microscopy of Aspergillus fumigatus keratitis,” Br. J. Ophthalmol. 87(4), 409–410 (2003).
[Crossref] [PubMed]

Kawamoto, K.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Kenney, M. C.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Kim, K. H.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Kim, M. J.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Kim, S. G.

E. C. Ledbetter, N. L. Irby, and S. G. Kim, “In vivo confocal microscopy of equine fungal keratitis,” Vet. Ophthalmol. 14(1), 1–9 (2011).
[Crossref] [PubMed]

Kimura, K.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Kojima, T.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
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König, K.

K. König, “Multiphoton microscopy in life sciences,” J. Microsc. 200(2), 83–104 (2000).
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Koop, N.

P. Steven, M. Müller, N. Koop, C. Rose, and G. Hüttmann, “Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies,” J. Biomed. Opt. 14(6), 064040 (2009).
[Crossref] [PubMed]

Laroche, L.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Ledbetter, E. C.

E. C. Ledbetter, N. L. Irby, and S. G. Kim, “In vivo confocal microscopy of equine fungal keratitis,” Vet. Ophthalmol. 14(1), 1–9 (2011).
[Crossref] [PubMed]

Lee, J. H.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Lee, S.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Lei, T. C.

E. A. Gibson, O. Masihzadeh, T. C. Lei, D. A. Ammar, and M. Y. Kahook, “Multiphoton microscopy for ophthalmic imaging,” J. Ophthalmol. 2011, 870879 (2011).
[Crossref] [PubMed]

Lin, H. C.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

Lin, H. H.

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Lin, S. J.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Lin, W. C.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Lo, W.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. Tech. 71(2), 83–85 (2008).
[Crossref] [PubMed]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Ma, D. H.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

Mantopoulos, D.

D. Mantopoulos, A. Cruzat, and P. Hamrah, “In vivo imaging of corneal inflammation: new tools for clinical practice and research,” Semin. Ophthalmol. 25(5-6), 178–185 (2010).
[Crossref] [PubMed]

Masihzadeh, O.

E. A. Gibson, O. Masihzadeh, T. C. Lei, D. A. Ammar, and M. Y. Kahook, “Multiphoton microscopy for ophthalmic imaging,” J. Ophthalmol. 2011, 870879 (2011).
[Crossref] [PubMed]

McGhee, C. N.

R. L. Niederer and C. N. McGhee, “Clinical in vivo confocal microscopy of the human cornea in health and disease,” Prog. Retin. Eye Res. 29(1), 30–58 (2010).
[Crossref] [PubMed]

Merigan, W. H.

Morishige, N.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Morita, Y.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Müller, M.

P. Steven, M. Müller, N. Koop, C. Rose, and G. Hüttmann, “Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies,” J. Biomed. Opt. 14(6), 064040 (2009).
[Crossref] [PubMed]

Niederer, R. L.

R. L. Niederer and C. N. McGhee, “Clinical in vivo confocal microscopy of the human cornea in health and disease,” Prog. Retin. Eye Res. 29(1), 30–58 (2010).
[Crossref] [PubMed]

Nishida, T.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Ohta, H.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Palczewska, G.

Palczewski, K.

Patel, S. V.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Peng, J. L.

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Pflugfelder, S. C.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Rodallec, T.

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

Rose, C.

P. Steven, M. Müller, N. Koop, C. Rose, and G. Hüttmann, “Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies,” J. Biomed. Opt. 14(6), 064040 (2009).
[Crossref] [PubMed]

Sharma, R.

Shin-Gyou-Uchi, R.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

So, P. T.

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Song, I. S.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Sonoda, K. H.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Srinivasan, M.

J. P. Whitcher, M. Srinivasan, and M. P. Upadhyay, “Corneal blindness: a global perspective,” Bull. World Health Organ. 79(3), 214–221 (2001).
[PubMed]

Steven, P.

P. Steven, F. Bock, G. Hüttmann, and C. Cursiefen, “Intravital two-photon microscopy of immune cell dynamics in corneal lymphatic vessels,” PLoS One 6(10), e26253 (2011).
[Crossref] [PubMed]

P. Steven, M. Müller, N. Koop, C. Rose, and G. Hüttmann, “Comparison of Cornea Module and DermaInspect for noninvasive imaging of ocular surface pathologies,” J. Biomed. Opt. 14(6), 064040 (2009).
[Crossref] [PubMed]

Strickler, J. H.

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248(4951), 73–76 (1990).
[Crossref] [PubMed]

Sun, Y.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. Tech. 71(2), 83–85 (2008).
[Crossref] [PubMed]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Takahara, A.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Tan, H. Y.

W. L. Chen, Y. Sun, W. Lo, H. Y. Tan, and C. Y. Dong, “Combination of multiphoton and reflective confocal imaging of cornea,” Microsc. Res. Tech. 71(2), 83–85 (2008).
[Crossref] [PubMed]

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Tchah, H.

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Teng, S. W.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

S. W. Teng, H. Y. Tan, J. L. Peng, H. H. Lin, K. H. Kim, W. Lo, Y. Sun, W. C. Lin, S. J. Lin, S. H. Jee, P. T. So, and C. Y. Dong, “Multiphoton autofluorescence and second-harmonic generation imaging of the ex vivo porcine eye,” Invest. Ophthalmol. Vis. Sci. 47(3), 1216–1224 (2006).
[Crossref] [PubMed]

Thomas, P. A.

P. A. Thomas and P. Geraldine, “Infectious keratitis,” Curr. Opin. Infect. Dis. 20(2), 129–141 (2007).
[Crossref] [PubMed]

Upadhyay, M. P.

J. P. Whitcher, M. Srinivasan, and M. P. Upadhyay, “Corneal blindness: a global perspective,” Bull. World Health Organ. 79(3), 214–221 (2001).
[PubMed]

Varnell, E. D.

A. M. Avunduk, R. W. Beuerman, E. D. Varnell, and H. E. Kaufman, “Confocal microscopy of Aspergillus fumigatus keratitis,” Br. J. Ophthalmol. 87(4), 409–410 (2003).
[Crossref] [PubMed]

Villani, E.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Wahlert, A. J.

N. Morishige, A. J. Wahlert, M. C. Kenney, D. J. Brown, K. Kawamoto, T. Chikama, T. Nishida, and J. V. Jester, “Second-harmonic imaging microscopy of normal human and keratoconus cornea,” Invest. Ophthalmol. Vis. Sci. 48(3), 1087–1094 (2007).
[Crossref] [PubMed]

Webb, W. W.

W. R. Zipfel, R. M. Williams, and W. W. Webb, “Nonlinear magic: multiphoton microscopy in the biosciences,” Nat. Biotechnol. 21(11), 1369–1377 (2003).
[Crossref] [PubMed]

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248(4951), 73–76 (1990).
[Crossref] [PubMed]

Whitcher, J. P.

J. P. Whitcher, M. Srinivasan, and M. P. Upadhyay, “Corneal blindness: a global perspective,” Bull. World Health Organ. 79(3), 214–221 (2001).
[PubMed]

Williams, D. R.

Williams, R. M.

W. R. Zipfel, R. M. Williams, and W. W. Webb, “Nonlinear magic: multiphoton microscopy in the biosciences,” Nat. Biotechnol. 21(11), 1369–1377 (2003).
[Crossref] [PubMed]

Wu, R. J.

H. Y. Tan, Y. Sun, W. Lo, S. W. Teng, R. J. Wu, S. H. Jee, W. C. Lin, C. H. Hsiao, H. C. Lin, Y. F. Chen, D. H. Ma, S. C. Huang, S. J. Lin, and C. Y. Dong, “Multiphoton fluorescence and second harmonic generation microscopy for imaging infectious keratitis,” J. Biomed. Opt. 12(2), 024013 (2007).
[Crossref] [PubMed]

Yamada, N.

N. Morishige, R. Shin-Gyou-Uchi, H. Azumi, H. Ohta, Y. Morita, N. Yamada, K. Kimura, A. Takahara, and K. H. Sonoda, “Quantitative analysis of collagen lamellae in the normal and keratoconic human cornea by second harmonic generation imaging microscopy,” Invest. Ophthalmol. Vis. Sci. 55(12), 8377–8385 (2014).
[Crossref] [PubMed]

Yin, L.

Zhivov, A.

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Zipfel, W. R.

W. R. Zipfel, R. M. Williams, and W. W. Webb, “Nonlinear magic: multiphoton microscopy in the biosciences,” Nat. Biotechnol. 21(11), 1369–1377 (2003).
[Crossref] [PubMed]

Biomed. Opt. Express (1)

Br. J. Ophthalmol. (2)

E. Brasnu, T. Bourcier, B. Dupas, S. Degorge, T. Rodallec, L. Laroche, V. Borderie, and C. Baudouin, “In vivo confocal microscopy in fungal keratitis,” Br. J. Ophthalmol. 91(5), 588–591 (2007).
[Crossref] [PubMed]

A. M. Avunduk, R. W. Beuerman, E. D. Varnell, and H. E. Kaufman, “Confocal microscopy of Aspergillus fumigatus keratitis,” Br. J. Ophthalmol. 87(4), 409–410 (2003).
[Crossref] [PubMed]

Bull. World Health Organ. (1)

J. P. Whitcher, M. Srinivasan, and M. P. Upadhyay, “Corneal blindness: a global perspective,” Bull. World Health Organ. 79(3), 214–221 (2001).
[PubMed]

Cornea (1)

U. Jurkunas, I. Behlau, and K. Colby, “Fungal keratitis: changing pathogens and risk factors,” Cornea 28(6), 638–643 (2009).
[Crossref] [PubMed]

Curr. Eye Res. (1)

E. Villani, C. Baudouin, N. Efron, P. Hamrah, T. Kojima, S. V. Patel, S. C. Pflugfelder, A. Zhivov, and M. Dogru, “In vivo confocal microscopy of the ocular surface: from bench to bedside,” Curr. Eye Res. 39(3), 213–231 (2014).
[Crossref] [PubMed]

Curr. Opin. Infect. Dis. (1)

P. A. Thomas and P. Geraldine, “Infectious keratitis,” Curr. Opin. Infect. Dis. 20(2), 129–141 (2007).
[Crossref] [PubMed]

Exp. Eye Res. (1)

J. H. Lee, S. Lee, Y. S. Gho, I. S. Song, H. Tchah, M. J. Kim, and K. H. Kim, “Comparison of confocal microscopy and two-photon microscopy in mouse cornea in vivo,” Exp. Eye Res. 132, 101–108 (2015).
[Crossref] [PubMed]

Invest. Ophthalmol. Vis. Sci. (3)

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Supplementary Material (10)

NameDescription
» Visualization 1: MOV (552 KB)      TPM of aspergillus fumigatus in aqueous media
» Visualization 2: MOV (499 KB)      TPM of candida albicans in aqueous media
» Visualization 3: MOV (1780 KB)      RCM of aspergillus infected-rabbit cornea number 1
» Visualization 4: MOV (1908 KB)      TPSHGM of aspergillus infected-rabbit cornea number 1
» Visualization 5: MOV (1085 KB)      RCM of aspergillus infected-rabbit cornea number 2
» Visualization 6: MOV (1285 KB)      TPSHGM of aspergillus infected-rabbit cornea number 2
» Visualization 7: MOV (1380 KB)      RCM of candida infected-rabbit cornea number 1
» Visualization 8: MOV (1385 KB)      TPSHGM of candida infected-rabbit cornea number 1
» Visualization 9: MOV (731 KB)      RCM of candida infected-rabbit cornea number 2
» Visualization 10: MOV (1317 KB)      TPSHGM of candida infected-rabbit cornea number 2

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Figures (7)

Fig. 1
Fig. 1 Leica SP5 scanning microscope system and acrylic eye holder.
Fig. 2
Fig. 2 TPM images of fungal cells in aqueous media in the x-y plane. (a): aspergillus fumigatus (Visualization 1) and (b): zoomed image of white box of (a). (c): candida ablicans (Visualization 2) and (d): zoomed image of white box of (c). These images are maximum projection images (MPI) from 3D TPM data. Scale bar: 50 µm
Fig. 3
Fig. 3 RCM and TPSHGM images of the aspergillus fumigatus-infected rabbit cornea #1 at several depths. (a-c): RCM images at 60, 120, and 250 µm deep from the surface, respectively (Visualization 3), (d-f) and (g-i): TPSHGM AF and SHG images at the same depths as RCM images (Visualization 4, green: autofluorescence, blue: SHG). Yellow dashed circle: cluster of aspergillus fumigatus, yellow arrow head: single aspergillus fumigatus cells. Scale bar: 100 µm
Fig. 4
Fig. 4 3D RCM and TPSHGM images of the aspergillus fumigatus-infected rabbit cornea # 2 in the x-y plane at several depths. (a-c): RCM images (Visualization 5) at 16, 56, and 80 µm deep from the surface, (d-f) and (g-i): corresponding TPSHGM AF and SHG images (Visualization 6, green: autofluorescence, blue: SHG). Yellow arrow head: aspergiilus fumigatus cells. Scale bar: 100 µm
Fig. 5
Fig. 5 3D RCM and TPSHGM images of candida albicans-infected rabbit cornea # 1 in the x-y plane at three depths. (a-c): RCM images at surface, 25, and 60 µm deep from the surface (Visualization 7), (d-f) and (g-i): corresponding TPSHGM AF and SHG images to the RCM images (Visualization 8, green: autofluorescence, blue: SHG). Scale bar: 100 µm
Fig. 6
Fig. 6 3D RCM and TPSHGM images of candida albicans-infected rabbit cornea # 2 in the x-y plane at three different depths. (a-c): RCM images at surface, 60, and 100 µm deep from the surface (Visualization 9), (d-f) and (g-i): corresponding TPSHGM AF and SHG images as the RCM images (Visualization 10, green: AF, blue: SHG). Yellow arrow head: candida albicans cells, white arrow head: inflammatory cells, yellow dashed circle: neuron. Scale bar: 100 µm
Fig. 7
Fig. 7 Histology images of aspergillus fumigatus and candida albicans-infected rabbit corneas. (a) and (b): aspergillus and candida-infected corneas. Scale bar: 100 µm

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