Combined non-linear imaging techniques were used to deeply image human ex-vivo fresh biopsies of bladder as well as to discriminate between healthy bladder mucosa and carcinoma in situ. Morphological examination by two-photon excited fluorescence and second-harmonic generation has shown a good agreement with corresponding common routine histology performed on the same samples. Tumor cells appeared slightly different in shape and with a smaller cellular-to-nuclear dimension ratio with respect to corresponding normal cells. Further differences between the two tissue types were found in both spectral emission and fluorescence lifetime distribution by performing temporal- and spectral- resolved analysis of fluorescence. This method may represent a promising tool to be used in a multi-photon endoscope, in a confocal endoscope or in a spectroscopic probe for in-vivo optical diagnosis of bladder cancer.

© 2010 OSA

1. ACS, (2006), http://www.cancer.org/downloads/STT/CAFF2006PWSecured.pdf , Cancer Facts & Figs. (2006).
2. J. C. Kah, W. K. Lau, P. H. Tan, C. J. Sheppard, and M. Olivo, “Endoscopic image analysis of photosensitizer fluorescence as a promising noninvasive approach for pathological grading of bladder cancer in situ,” J. Biomed. Opt. 13(5), 054022 (2008). [CrossRef] [PubMed]
3. E. L. Larsen, L. L. Randeberg, O. A. Gederaas, C. J. Arum, A. Hjelde, C. M. Zhao, D. Chen, H. E. Krokan, and L. O. Svaasand, “Monitoring of hexyl 5-aminolevulinate-induced photodynamic therapy in rat bladder cancer by optical spectroscopy,” J. Biomed. Opt. 13(4), 044031 (2008). [CrossRef] [PubMed]
4. S. Berrahmoune, N. Fotinos, L. Bezdetnaya, N. Lange, J. C. Guedenet, F. Guillemin, and M. A. D’Hallewin, “Analysis of differential PDT effect in rat bladder tumor models according to concentrations of intravesical hexyl-aminolevulinate,” Photochem. Photobiol. Sci. 7(9), 1018–1024 (2008). [CrossRef] [PubMed]
5. D. Jocham, F. Witjes, S. Wagner, B. Zeylemaker, J. van Moorselaar, M. O. Grimm, R. Muschter, G. Popken, F. König, R. Knüchel, and K. H. Kurth, “Improved detection and treatment of bladder cancer using hexaminolevulinate imaging: a prospective, phase III multicenter study,” J. Urol. 174(3), 862–866, discussion 866 (2005). [CrossRef] [PubMed]
6. N. Ramanujam, “Fluorescence spectroscopy of neoplastic and non-neoplastic tissues,” Neoplasia 2(1/2), 89–117 (2000). [CrossRef] [PubMed]
7. M. Anidjar, O. Cussenot, S. Avrillier, D. Ettori, P. Teillac, and A. Le Duc, “The role of laser-induced autofluorescence spectroscopy in bladder tumor detection. Dependence on the excitation wavelength,” Ann. N. Y. Acad. Sci. 838(1 ADVANCES IN O), 130–141 (1998). [CrossRef] [PubMed]
8. F. Koenig, F. J. McGovern, A. F. Althausen, T. F. Deutsch, and K. T. Schomacker, “Laser induced autofluorescence diagnosis of bladder cancer,” J. Urol. 156(5), 1597–1601 (1996). [CrossRef] [PubMed]
9. D. Zaak, H. Stepp, R. Baumgartner, P. Schneede, R. Waidelich, D. Frimberger, A. Hartmann, R. Künchel, A. Hofstetter, and A. Hohla, “Ultraviolet-excited (308 nm) autofluorescence for bladder cancer detection,” Urology 60(6), 1029–1033 (2002). [CrossRef] [PubMed]
10. W. W. Chin, P. S. P. Thong, R. Bhuvaneswari, K. C. Soo, P. W. S. Heng, and M. Olivo, “In-vivo optical detection of cancer using chlorin e6-polyvinylpyrrolidone induced fluorescence imaging and spectroscopy,” BMC Med. Imaging 9(1), 1–8 (2009). [CrossRef] [PubMed]
11. Z. Yuan, Z. Wang, R. Pan, J. Liu, H. Cohen, and Y. Pan, “High-resolution imaging diagnosis and staging of bladder cancer: comparison between optical coherence tomography and high-frequency ultrasound,” J. Biomed. Opt. 13(5), 054007 (2008). [CrossRef] [PubMed]
12. Z. G. Wang, D. B. Durand, M. Schoenberg, and Y. T. Pan, “Fluorescence guided optical coherence tomography for the diagnosis of early bladder cancer in a rat model,” J. Urol. 174(6), 2376–2381 (2005). [CrossRef] [PubMed]
13. B. Hermes, F. Spöler, A. Naami, J. Bornemann, M. Först, J. Grosse, G. Jakse, and R. Knüchel, “Visualization of the basement membrane zone of the bladder by optical coherence tomography: feasibility of noninvasive evaluation of tumor invasion,” Urology 72(3), 677–681 (2008). [CrossRef] [PubMed]
14. F. Koenig, J. Knittel, L. Schnieder, M. George, M. Lein, and D. Schnorr, “Confocal laser scanning microscopy of urinary bladder after intravesical instillation of a fluorescent dye,” Urology 62(1), 158–161 (2003). [CrossRef] [PubMed]
15. G. A. Sonn, S. N. Jones, T. V. Tarin, C. B. Du, K. E. Mach, K. C. Jensen, and J. C. Liao, “Optical biopsy of human bladder neoplasia with in vivo confocal laser endomicroscopy,” J. Urol. 182(4), 1299–1305 (2009). [CrossRef] [PubMed]
16. M. E. Llewellyn, R. P. J. Barretto, S. L. Delp, and M. J. Schnitzer, “Minimally invasive high-speed imaging of sarcomere contractile dynamics in mice and humans,” Nature 454(7205), 784–788 (2008). [PubMed]
17. B. R. Masters, P. T. C. So, and E. Gratton, “Optical biopsy of in vivo human skin: multi-photon excitation microscopy,” Lasers Med. Sci. 13(3), 196–203 (1998). [CrossRef]
18. P. T. C. So, H. Kim, and I. E. Kochevar, “Two-Photon deep tissue ex vivo imaging of mouse dermal and subcutaneous structures,” Opt. Express 3(9), 339–350 (1998). [CrossRef] [PubMed]
19. J. C. Malone, A. F. Hood, T. Conley, J. Nürnberger, L. A. Baldridge, J. L. Clendenon, K. W. Dunn, and C. L. Phillips, “Three-dimensional imaging of human skin and mucosa by two-photon laser scanning microscopy,” J. Cutan. Pathol. 29(8), 453–458 (2002). [CrossRef] [PubMed]
20. K. König and I. Riemann, “High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution,” J. Biomed. Opt. 8(3), 432–439 (2003). [CrossRef] [PubMed]
21. K. Steenkeste, S. Lécart, A. Deniset, P. Pernot, P. Eschwège, S. Ferlicot, S. Lévêque-Fort, R. Briandet, and M. P. Fontaine-Aupart, “Ex vivo fluorescence imaging of normal and malignant urothelial cells to enhance early diagnosis,” Photochem. Photobiol. 83(5), 1157–1166 (2007). [CrossRef] [PubMed]
22. S. M. Zhuo, J. X. Chen, T. Luo, X. S. Jiang, and S. S. Xie, “Multiphoton microscopy of unstained bladder mucosa based on two-photon excited autofluorescence and second-harmonic generation,” Laser Phys. Lett. 6(1), 80–83 (2009). [CrossRef]
23. R. Yadav, S. Mukherjee, M. Hermen, G. Tan, F. R. Maxfield, W. W. Webb, and A. K. Tewari, “Multiphoton microscopy of prostate and periprostatic neural tissue: a promising imaging technique for improving nerve-sparing prostatectomy,” J. Endourol. 23(5), 861–867 (2009). [CrossRef] [PubMed]
24. W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248(4951), 73–76 (1990). [CrossRef] [PubMed]
25. 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]
26. 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]
27. 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]
28. 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]
29. E. Brown, T. McKee, E. diTomaso, A. Pluen, B. Seed, Y. Boucher, and R. K. Jain, “Dynamic imaging of collagen and its modulation in tumors in vivo using second-harmonic generation,” Nat. Med. 9(6), 796–801 (2003). [CrossRef] [PubMed]
30. P. P. Provenzano, K. W. Eliceiri, J. M. Campbell, D. R. Inman, J. G. White, and P. J. Keely, “Collagen reorganization at the tumor-stromal interface facilitates local invasion,” BMC Med. 4(1), 38 (2006). [CrossRef] [PubMed]
31. R. Cicchi, S. Sestini, V. De Giorgi, D. Massi, T. Lotti, and F. S. Pavone, “Non-linear laser imaging of skin lesions,” J. Biophoton. 1(1), 62–73 (2008). [CrossRef]
32. L. H. Laiho, S. Pelet, T. M. Hancewicz, P. D. Kaplan, and P. T. C. So, “Two-photon 3-D mapping of ex vivo human skin endogenous fluorescence species based on fluorescence emission spectra,” J. Biomed. Opt. 10(2), 024016 (2005). [CrossRef] [PubMed]
33. P. J. Tadrous, “Methods for imaging the structure and function of living tissues and cells: 2. Fluorescence lifetime imaging,” J. Pathol. 191(3), 229–234 (2000). [CrossRef] [PubMed]
34. P. J. Tadrous, J. Siegel, P. M. W. French, S. Shousha, N. Lalani, and G. W. H. Stamp, “Fluorescence lifetime imaging of unstained tissues: early results in human breast cancer,” J. Pathol. 199(3), 309–317 (2003). [CrossRef] [PubMed]
35. Y. Chen and A. Periasamy, “Characterization of two-photon excitation fluorescence lifetime imaging microscopy for protein localization,” Microsc. Res. Tech. 63(1), 72–80 (2004). [CrossRef]
36. S. Y. Breusegem, M. Levi, and N. P. Barry, “Fluorescence correlation spectroscopy and fluorescence lifetime imaging microscopy,” Nephron, Exp. Nephrol. 103(2), e41–e49 (2006). [CrossRef]
37. R. Cicchi, D. Massi, S. Sestini, P. Carli, V. De Giorgi, T. Lotti, and F. S. Pavone, “Multidimensional non-linear laser imaging of Basal Cell Carcinoma,” Opt. Express 15(16), 10135–10148 (2007). [CrossRef] [PubMed]
38. R. Cicchi, L. Sacconi, A. Jasaitis, R. P. O’Connor, D. Massi, S. Sestini, V. De Giorgi, T. Lotti, and F. S. Pavone, “Multidimensional custom-made non-linear microscope: from ex-vivo to in-vivo imaging,” Appl. Phys. B 92(3), 359–365 (2008). [CrossRef]
39. S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30(17), 2275–2277 (2005). [CrossRef] [PubMed]
40. J. Paoli, M. Smedh, A. M. Wennberg, and M. B. Ericson, “Multiphoton laser scanning microscopy on non-melanoma skin cancer: morphologic features for future non-invasive diagnostics,” J. Invest. Dermatol. 128(5), 1248–1255 (2008). [CrossRef]
41. O. Warburg, “The metabolism of tumors,” Constabel, London (1930).
42. M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eickhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, “In vivo multiphoton fluorescence lifetime imaging of protein-bound and free nicotinamide adenine dinucleotide in normal and precancerous epithelia,” J. Biomed. Opt. 12(2), 024014 (2007). [CrossRef] [PubMed]
43. D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, “Metabolic mapping of MCF10A human breast cells via multiphoton fluorescence lifetime imaging of the coenzyme NADH,” Cancer Res. 65(19), 8766–8773 (2005). [CrossRef] [PubMed]
44. M. C. Skala, K. M. Riching, A. Gendron-Fitzpatrick, J. Eickhoff, K. W. Eliceiri, J. G. White, and N. Ramanujam, “In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia,” Proc. Natl. Acad. Sci. U.S.A. 104(49), 19494–19499 (2007). [CrossRef] [PubMed]

Ann. N. Y. Acad. Sci.

• M. Anidjar, O. Cussenot, S. Avrillier, D. Ettori, P. Teillac, and A. Le Duc, “The role of laser-induced autofluorescence spectroscopy in bladder tumor detection. Dependence on the excitation wavelength,” Ann. N. Y. Acad. Sci. 838(1 ADVANCES IN O), 130–141 (1998). [CrossRef] [PubMed]

Appl. Phys. B

• R. Cicchi, L. Sacconi, A. Jasaitis, R. P. O’Connor, D. Massi, S. Sestini, V. De Giorgi, T. Lotti, and F. S. Pavone, “Multidimensional custom-made non-linear microscope: from ex-vivo to in-vivo imaging,” Appl. Phys. B 92(3), 359–365 (2008). [CrossRef]

BMC Med.

• P. P. Provenzano, K. W. Eliceiri, J. M. Campbell, D. R. Inman, J. G. White, and P. J. Keely, “Collagen reorganization at the tumor-stromal interface facilitates local invasion,” BMC Med. 4(1), 38 (2006). [CrossRef] [PubMed]

BMC Med. Imaging

• W. W. Chin, P. S. P. Thong, R. Bhuvaneswari, K. C. Soo, P. W. S. Heng, and M. Olivo, “In-vivo optical detection of cancer using chlorin e6-polyvinylpyrrolidone induced fluorescence imaging and spectroscopy,” BMC Med. Imaging 9(1), 1–8 (2009). [CrossRef] [PubMed]

Cancer Res.

• D. K. Bird, L. Yan, K. M. Vrotsos, K. W. Eliceiri, E. M. Vaughan, P. J. Keely, J. G. White, and N. Ramanujam, “Metabolic mapping of MCF10A human breast cells via multiphoton fluorescence lifetime imaging of the coenzyme NADH,” Cancer Res. 65(19), 8766–8773 (2005). [CrossRef] [PubMed]

J. Biomed. Opt.

• M. C. Skala, K. M. Riching, D. K. Bird, A. Gendron-Fitzpatrick, J. Eickhoff, K. W. Eliceiri, P. J. Keely, and N. Ramanujam, “In vivo multiphoton fluorescence lifetime imaging of protein-bound and free nicotinamide adenine dinucleotide in normal and precancerous epithelia,” J. Biomed. Opt. 12(2), 024014 (2007). [CrossRef] [PubMed]
• Z. Yuan, Z. Wang, R. Pan, J. Liu, H. Cohen, and Y. Pan, “High-resolution imaging diagnosis and staging of bladder cancer: comparison between optical coherence tomography and high-frequency ultrasound,” J. Biomed. Opt. 13(5), 054007 (2008). [CrossRef] [PubMed]
• J. C. Kah, W. K. Lau, P. H. Tan, C. J. Sheppard, and M. Olivo, “Endoscopic image analysis of photosensitizer fluorescence as a promising noninvasive approach for pathological grading of bladder cancer in situ,” J. Biomed. Opt. 13(5), 054022 (2008). [CrossRef] [PubMed]
• E. L. Larsen, L. L. Randeberg, O. A. Gederaas, C. J. Arum, A. Hjelde, C. M. Zhao, D. Chen, H. E. Krokan, and L. O. Svaasand, “Monitoring of hexyl 5-aminolevulinate-induced photodynamic therapy in rat bladder cancer by optical spectroscopy,” J. Biomed. Opt. 13(4), 044031 (2008). [CrossRef] [PubMed]
• L. H. Laiho, S. Pelet, T. M. Hancewicz, P. D. Kaplan, and P. T. C. So, “Two-photon 3-D mapping of ex vivo human skin endogenous fluorescence species based on fluorescence emission spectra,” J. Biomed. Opt. 10(2), 024016 (2005). [CrossRef] [PubMed]
• K. König and I. Riemann, “High-resolution multiphoton tomography of human skin with subcellular spatial resolution and picosecond time resolution,” J. Biomed. Opt. 8(3), 432–439 (2003). [CrossRef] [PubMed]

J. Biophoton.

• R. Cicchi, S. Sestini, V. De Giorgi, D. Massi, T. Lotti, and F. S. Pavone, “Non-linear laser imaging of skin lesions,” J. Biophoton. 1(1), 62–73 (2008). [CrossRef]

J. Cutan. Pathol.

• J. C. Malone, A. F. Hood, T. Conley, J. Nürnberger, L. A. Baldridge, J. L. Clendenon, K. W. Dunn, and C. L. Phillips, “Three-dimensional imaging of human skin and mucosa by two-photon laser scanning microscopy,” J. Cutan. Pathol. 29(8), 453–458 (2002). [CrossRef] [PubMed]

J. Endourol.

• R. Yadav, S. Mukherjee, M. Hermen, G. Tan, F. R. Maxfield, W. W. Webb, and A. K. Tewari, “Multiphoton microscopy of prostate and periprostatic neural tissue: a promising imaging technique for improving nerve-sparing prostatectomy,” J. Endourol. 23(5), 861–867 (2009). [CrossRef] [PubMed]

J. Invest. Dermatol.

• J. Paoli, M. Smedh, A. M. Wennberg, and M. B. Ericson, “Multiphoton laser scanning microscopy on non-melanoma skin cancer: morphologic features for future non-invasive diagnostics,” J. Invest. Dermatol. 128(5), 1248–1255 (2008). [CrossRef]

J. Pathol.

• P. J. Tadrous, “Methods for imaging the structure and function of living tissues and cells: 2. Fluorescence lifetime imaging,” J. Pathol. 191(3), 229–234 (2000). [CrossRef] [PubMed]
• P. J. Tadrous, J. Siegel, P. M. W. French, S. Shousha, N. Lalani, and G. W. H. Stamp, “Fluorescence lifetime imaging of unstained tissues: early results in human breast cancer,” J. Pathol. 199(3), 309–317 (2003). [CrossRef] [PubMed]

J. Urol.

• F. Koenig, F. J. McGovern, A. F. Althausen, T. F. Deutsch, and K. T. Schomacker, “Laser induced autofluorescence diagnosis of bladder cancer,” J. Urol. 156(5), 1597–1601 (1996). [CrossRef] [PubMed]
• G. A. Sonn, S. N. Jones, T. V. Tarin, C. B. Du, K. E. Mach, K. C. Jensen, and J. C. Liao, “Optical biopsy of human bladder neoplasia with in vivo confocal laser endomicroscopy,” J. Urol. 182(4), 1299–1305 (2009). [CrossRef] [PubMed]
• D. Jocham, F. Witjes, S. Wagner, B. Zeylemaker, J. van Moorselaar, M. O. Grimm, R. Muschter, G. Popken, F. König, R. Knüchel, and K. H. Kurth, “Improved detection and treatment of bladder cancer using hexaminolevulinate imaging: a prospective, phase III multicenter study,” J. Urol. 174(3), 862–866, discussion 866 (2005). [CrossRef] [PubMed]
• Z. G. Wang, D. B. Durand, M. Schoenberg, and Y. T. Pan, “Fluorescence guided optical coherence tomography for the diagnosis of early bladder cancer in a rat model,” J. Urol. 174(6), 2376–2381 (2005). [CrossRef] [PubMed]

Laser Phys. Lett.

• S. M. Zhuo, J. X. Chen, T. Luo, X. S. Jiang, and S. S. Xie, “Multiphoton microscopy of unstained bladder mucosa based on two-photon excited autofluorescence and second-harmonic generation,” Laser Phys. Lett. 6(1), 80–83 (2009). [CrossRef]

Lasers Med. Sci.

• B. R. Masters, P. T. C. So, and E. Gratton, “Optical biopsy of in vivo human skin: multi-photon excitation microscopy,” Lasers Med. Sci. 13(3), 196–203 (1998). [CrossRef]

Microsc. Res. Tech.

• Y. Chen and A. Periasamy, “Characterization of two-photon excitation fluorescence lifetime imaging microscopy for protein localization,” Microsc. Res. Tech. 63(1), 72–80 (2004). [CrossRef]

Nat. Biotechnol.

• 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]
• 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]

Nat. Med.

• E. Brown, T. McKee, E. diTomaso, A. Pluen, B. Seed, Y. Boucher, and R. K. Jain, “Dynamic imaging of collagen and its modulation in tumors in vivo using second-harmonic generation,” Nat. Med. 9(6), 796–801 (2003). [CrossRef] [PubMed]

Nature

• M. E. Llewellyn, R. P. J. Barretto, S. L. Delp, and M. J. Schnitzer, “Minimally invasive high-speed imaging of sarcomere contractile dynamics in mice and humans,” Nature 454(7205), 784–788 (2008). [PubMed]

Neoplasia

• N. Ramanujam, “Fluorescence spectroscopy of neoplastic and non-neoplastic tissues,” Neoplasia 2(1/2), 89–117 (2000). [CrossRef] [PubMed]

Nephron, Exp. Nephrol.

• S. Y. Breusegem, M. Levi, and N. P. Barry, “Fluorescence correlation spectroscopy and fluorescence lifetime imaging microscopy,” Nephron, Exp. Nephrol. 103(2), e41–e49 (2006). [CrossRef]

Opt. Express

• R. Cicchi, D. Massi, S. Sestini, P. Carli, V. De Giorgi, T. Lotti, and F. S. Pavone, “Multidimensional non-linear laser imaging of Basal Cell Carcinoma,” Opt. Express 15(16), 10135–10148 (2007). [CrossRef] [PubMed]
• P. T. C. So, H. Kim, and I. E. Kochevar, “Two-Photon deep tissue ex vivo imaging of mouse dermal and subcutaneous structures,” Opt. Express 3(9), 339–350 (1998). [CrossRef] [PubMed]

Opt. Lett.

• S. J. Lin, R. J. Wu, H. Y. Tan, W. Lo, W. C. Lin, T. H. Young, C. J. Hsu, J. S. Chen, S. H. Jee, and C. Y. Dong, “Evaluating cutaneous photoaging by use of multiphoton fluorescence and second-harmonic generation microscopy,” Opt. Lett. 30(17), 2275–2277 (2005). [CrossRef] [PubMed]

Photochem. Photobiol.

• K. Steenkeste, S. Lécart, A. Deniset, P. Pernot, P. Eschwège, S. Ferlicot, S. Lévêque-Fort, R. Briandet, and M. P. Fontaine-Aupart, “Ex vivo fluorescence imaging of normal and malignant urothelial cells to enhance early diagnosis,” Photochem. Photobiol. 83(5), 1157–1166 (2007). [CrossRef] [PubMed]

Photochem. Photobiol. Sci.

• S. Berrahmoune, N. Fotinos, L. Bezdetnaya, N. Lange, J. C. Guedenet, F. Guillemin, and M. A. D’Hallewin, “Analysis of differential PDT effect in rat bladder tumor models according to concentrations of intravesical hexyl-aminolevulinate,” Photochem. Photobiol. Sci. 7(9), 1018–1024 (2008). [CrossRef] [PubMed]

Proc. Natl. Acad. Sci. U.S.A.

• 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]
• 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]
• M. C. Skala, K. M. Riching, A. Gendron-Fitzpatrick, J. Eickhoff, K. W. Eliceiri, J. G. White, and N. Ramanujam, “In vivo multiphoton microscopy of NADH and FAD redox states, fluorescence lifetimes, and cellular morphology in precancerous epithelia,” Proc. Natl. Acad. Sci. U.S.A. 104(49), 19494–19499 (2007). [CrossRef] [PubMed]

Science

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

Urology

• B. Hermes, F. Spöler, A. Naami, J. Bornemann, M. Först, J. Grosse, G. Jakse, and R. Knüchel, “Visualization of the basement membrane zone of the bladder by optical coherence tomography: feasibility of noninvasive evaluation of tumor invasion,” Urology 72(3), 677–681 (2008). [CrossRef] [PubMed]
• F. Koenig, J. Knittel, L. Schnieder, M. George, M. Lein, and D. Schnorr, “Confocal laser scanning microscopy of urinary bladder after intravesical instillation of a fluorescent dye,” Urology 62(1), 158–161 (2003). [CrossRef] [PubMed]
• D. Zaak, H. Stepp, R. Baumgartner, P. Schneede, R. Waidelich, D. Frimberger, A. Hartmann, R. Künchel, A. Hofstetter, and A. Hohla, “Ultraviolet-excited (308 nm) autofluorescence for bladder cancer detection,” Urology 60(6), 1029–1033 (2002). [CrossRef] [PubMed]

Other

• ACS, (2006), http://www.cancer.org/downloads/STT/CAFF2006PWSecured.pdf , Cancer Facts & Figs. (2006).
• O. Warburg, “The metabolism of tumors,” Constabel, London (1930).

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