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, Daniel L. Farkas

Research output: Contribution to journalArticle

19 Scopus citations

Abstract

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.

Original languageEnglish (US)
Pages (from-to)356-364
Number of pages9
JournalBiomedical Optics Express
Volume2
Issue number2
DOIs
StatePublished - Feb 1 2011

ASJC Scopus subject areas

  • Atomic and Molecular Physics, and Optics
  • Biotechnology

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    Hwang, J. Y., Wachsmann-Hogiu, S., Krishnan Ramanujan, V., Nowatzyk, A. G., Koronyo, Y., Medina-Kauwe, L. K., Gross, Z., Gray, H. B., & Farkas, D. L. (2011). Multimodal wide-field two-photon excitation imaging: Characterization of the technique for in vivo applications. Biomedical Optics Express, 2(2), 356-364. https://doi.org/10.1364/BOE.2.000356