Research Papers

Two-color, two-photon, and excited-state absorption microscopy

[+] Author Affiliations
Dan Fu

Princeton University, Department of Chemistry, Princeton, New Jersey 08544

Tong Ye, Thomas E. Matthews

Duke University, Department of Chemistry, Durham, North Carolina 27708

Gunay Yurtsever

Duke University, Department of Biomedical Engineering, Durham, North Carolina 27708

Warren S. Warren

Duke University, Department of Chemistry and Radiology, Durham, North Carolina 27708

J. Biomed. Opt. 12(5), 054004 (September 17, 2007). doi:10.1117/1.2780173
History: Received November 08, 2006; Revised March 18, 2007; Accepted May 12, 2007; Published September 17, 2007
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We develop a new approach in imaging nonfluorescent species with two-color two-photon and excited state absorption microscopy. If one of two synchronized mode-locked pulse trains at different colors is intensity modulated, the modulation transfers to the other pulse train when nonlinear absorption takes places in the medium. We can easily measure 106 absorption changes caused by either two-photon absorption or excited-state absorption with a RF lock-in amplifier. Sepia melanin is studied in detail as a model system. Spectroscopy studies on the instantaneous two-photon absorption (TPA) and the relatively long-lived excited-state absorption (ESA) of melanin are carried out in solution, and imaging capability is demonstrated in B16 cells. It is found that sepia melanin exhibits two distinct excited states with different lifetimes (one at 3ps, one lasting hundreds of nanoseconds) when pumped at 775nm. Its characteristic TPA/ESA enables us to image its distribution in cell samples with high resolution comparable to two-photon fluorescence microscopy (TPFM). This new technique could potentially provide valuable information in diagnosing melanoma.

Figures in this Article
© 2007 Society of Photo-Optical Instrumentation Engineers

Topics

Absorption ; Photons

Citation

Dan Fu ; Tong Ye ; Thomas E. Matthews ; Gunay Yurtsever and Warren S. Warren
"Two-color, two-photon, and excited-state absorption microscopy", J. Biomed. Opt. 12(5), 054004 (September 17, 2007). ; http://dx.doi.org/10.1117/1.2780173


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