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Biomedical Optics Express

Biomedical Optics Express

  • Editor: Joseph A. Izatt
  • Vol. 2, Iss. 2 — Feb. 1, 2011
  • pp: 356–364

Multimodal wide-field two-photon excitation imaging: characterization of the technique for in vivo applications

Jae Youn Hwang, Sebastian Wachsmann-Hogiu, V Krishnan Ramanujan, Andreas G. Nowatzyk, Yosef Koronyo, Lali K. Medina-Kauwe, Zeev Gross, Harry B. Gray, and Daniel L. Farkas  »View Author Affiliations

Biomedical Optics Express, Vol. 2, Issue 2, pp. 356-364 (2011)

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We report fast, non-scanning, wide-field two-photon fluorescence excitation with spectral and lifetime detection for in vivo biomedical applications. We determined the optical characteristics of the technique, developed a Gaussian flat-field correction method to reduce artifacts resulting from non-uniform excitation such that contrast is enhanced, and showed that it can be used for ex vivo and in vivo cellular-level imaging. Two applications were demonstrated: (i) ex vivo measurements of beta-amyloid plaques in retinas of transgenic mice, and (ii) in vivo imaging of sulfonated gallium(III) corroles injected into tumors. We demonstrate that wide-field two photon fluorescence excitation with flat-field correction provides more penetration depth as well as better contrast and axial resolution than the corresponding one-photon wide field excitation for the same dye. Importantly, when this technique is used together with spectral and fluorescence lifetime detection modules, it offers improved discrimination between fluorescence from molecules of interest and autofluorescence, with higher sensitivity and specificity for in vivo applications.

© 2011 OSA

OCIS Codes
(110.4234) Imaging systems : Multispectral and hyperspectral imaging
(180.4315) Microscopy : Nonlinear microscopy

ToC Category:

Original Manuscript: September 22, 2010
Revised Manuscript: December 10, 2010
Manuscript Accepted: December 20, 2010
Published: January 13, 2011

Jae Youn Hwang, Sebastian Wachsmann-Hogiu, V Krishnan Ramanujan, Andreas G. Nowatzyk, Yosef Koronyo, Lali K. Medina-Kauwe, Zeev Gross, Harry B. Gray, and Daniel L. Farkas, "Multimodal wide-field two-photon excitation imaging: characterization of the technique for in vivo applications," Biomed. Opt. Express 2, 356-364 (2011)

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