Abstract

A portable confocal system with the excitations at 355nm and 457nm was instrumented to investigate the depth-resolved fluorescence of cervical tissue. The study focused on extracting biochemical and morphological information carried in the depth-resolved signals measured from the normal squamous epithelial tissue and squamous intraepithelial lesions. Strong keratin fluorescence with the spectral characteristics similar to collagen were observed from the topmost keratinizing layer of all tissue samples. It was found that NADH and FAD fluorescence measured from the underlying non-keratinizing epithelial layer were strongly correlated to the tissue pathology. This study demonstrates that the depth-resolved fluorescence spectroscopy can potentially provide more accurate diagnostic information for determining tissue pathology.

© 2005 Optical Society of America

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References

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Cancer

M. G. Muller, T. A. Valdez, I. Georgakoudi, V. Backman, C. Fuentes, S. Kabani, N. Laver, Z. Wang, C. W. Boone, R R. Dasari, S. M. Shapshay, M. S. Feld, �??Spectroscopic detection and evaluation of morphologic and biochemical changes in early human oral carcinoma,�?? Cancer 97, 1681-169 (2003)
[CrossRef] [PubMed]

Hum. Pathol.

C. Carrilho, M. Alberto, L. Buane and L. David, �??Keratins 8, 10, 13, and 17 are useful markers in the diagnosis of human cervix carcinomas,�?? Hum. Pathol. 35, 546-551(2004)
[CrossRef] [PubMed]

IEEE Trans. Biomed. Eng.

S. K. Chang, M. Follen, A. Malpica, U. Utzinger, G. Staerkel, D. Cox, E. N. Atkinson, C. MacAulay and R. Richards-Kortum, �??Optimal excitation wavelengths for discrimination of cervical neoplasia,�?? IEEE Trans. Biomed. Eng. 49, 1102-1110 (2002)
[CrossRef] [PubMed]

IEEE. J. Sel. Top. Quantum Electron.

T. Collier, D. Arifler, A. Malpica, M. Follen and R. Richards-Kortum, �??Determination of Epithelial tissue scattering coefficient using confocal microscopy,�?? IEEE. J. Sel. Top. Quantum Electron. 9, 307-313(2003)
[CrossRef]

J. Biol. Chem.

B Chance, B. Schoener, R. Oshino, F. Itshak, and Y. Nakase, �??Oxidation-reduction ratio studies of mitochondria in freeze-trapped samples,�?? J. Biol. Chem. 254, 4764-4771 (1979)
[PubMed]

Lasers Surg. Med.

G. M. Palmer, C. L. Marshek, K. M. Vrotsos and N. Ramanujam, "Optimal methods for fluorescence and diffuse reflectance measurements of tissue biopsy samples," Lasers Surg. Med. 30, 191-200 (2002)
[CrossRef] [PubMed]

Opt. Express

Photochem. Photobiol.

R. Drezek, C. Brookner, I. Pavlova, I. Boiko, A. Malpica, R. Lotan, M. Follen and R. Richards-Kortum, �??Autofluoresecence microscopy of fresh cervical-tissue sections reveals alterations in tissue biochemistry with dysplasia,�?? Photochem. Photobiol. 73, 636-641 (2001)
[CrossRef] [PubMed]

I. Pavlova, K. Sokolov K, R. Drezek, A. Malpica, M. Follen and R. Richards-Kortum, "Microanatomical and biochemical origins of normal and precancerous cervical autofluorescence using laser-scanning fluorescence confocal microscopy," Photochem. Photobiol. 77, 550-555 (2003)
[CrossRef] [PubMed]

Proc. Natl. Acad. Sci. USA.

N. Ramanujam, M. F. Mitchell, A. Mahadevan, S. Warren, S. Thomsen, E. Silva and R. Richards-Kortum, �??In vivo diagnosis of cervical intraepithelial neoplasia using 377-nm-excited laser-induced fluorescence,�?? Proc. Natl. Acad. Sci. USA. 91, 10193-10197 (1994)
[CrossRef] [PubMed]

Supplementary Material (2)

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Figures (4)

Fig. 1.
Fig. 1.

Representative depth-resolved autofluorescence signals of ectocervical tissue and corresponding histology. a (209kB), b: spectra recorded from a sample with highly keratinizing epithelium; c (250kB), d: spectra recorded from a sample with lightly keratinizing epithelium; e, f: corresponding H&E- and Mason-stained sections of the sample with highly keratinizing epithelium; g, h: corresponding H&E- and Mason-stained sections of the sample with lightly keratinizing epithelium; Scale bar in the histology e-h: 100µm.

Fig. 2.
Fig. 2.

Depth-resolved fluorescence spectra and corresponding histology of endocervical tissue samples. a, b: spectra and histology of a sample with normal submucosa; c, d: spectra and histology of a sample with highly vascularized submucosa. Scale bar in histology: 100 µm.

Fig. 3.
Fig. 3.

The averaged fluorescence intensities of the groups of normal tissue, HPV infection and CIN; a: 355nm excitation; b: 457nm excitation. Error bars represent standard deviations.

Fig. 4.
Fig. 4.

(a) Scatter plot for the UV/Blue ratio of each measurement site as a function of depth; (b) p-value of student’s t-test of normal and HPV vs. CIN groups as a function of depth.

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