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Deep Tissue Fluorescent Imaging in Scattering Specimens Using Confocal Microscopy

Published online by Cambridge University Press:  24 June 2011

Sherry G. Clendenon*
Affiliation:
Department of Medicine, Division of Nephrology, Indiana University Medical Center, Indianapolis, IN 46202, USA
Pamela A. Young
Affiliation:
Department of Medicine, Division of Nephrology, Indiana University Medical Center, Indianapolis, IN 46202, USA
Michael Ferkowicz
Affiliation:
Herman B. Wells Center for Pediatric Research and Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Carrie Phillips
Affiliation:
Division of Nephrology and Department of Pathology, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Kenneth W. Dunn
Affiliation:
Department of Medicine, Division of Nephrology, Indiana University Medical Center, Indianapolis, IN 46202, USA
*
Corresponding author. E-mail: [email protected]
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Abstract

In scattering specimens, multiphoton excitation and nondescanned detection improve imaging depth by a factor of 2 or more over confocal microscopy; however, imaging depth is still limited by scattering. We applied the concept of clearing to deep tissue imaging of highly scattering specimens. Clearing is a remarkably effective approach to improving image quality at depth using either confocal or multiphoton microscopy. Tissue clearing appears to eliminate the need for multiphoton excitation for deep tissue imaging.

Type
Technology and Software Development Light and Confocal Microscopy
Copyright
Copyright © Microscopy Society of America 2011

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References

REFERENCES

Amann, K., Nichols, C., Tornig, J., Schwarz, U., Zeier, M., Mall, G. & Ritz, E. (1996). Effect of ramipril, nifedipine, and moxonidine on glomerular morphology and podocyte structure in experimental renal failure. Nephrol Dial Transplant 11, 10031011.Google Scholar
Appleton, P.L., Quyn, A.J., Swift, S. & Nathke, I. (2009). Preparation of wholemount mouse intestine for high-resolution three-dimensional imaging using two-photon microscopy. J Microsc 234, 196204.Google Scholar
Centonze, V.E. & White, J.G. (1998). Multiphoton excitation provides optical sections from deeper within scattering specimens than confocal imaging. Biophys J 75, 20152024.Google Scholar
Clendenon, J.L., Phillips, C.L., Sandoval, R.M., Fang, S. & Dunn, K.W. (2002). Voxx: A PC-based, near real-time volume rendering system for biological microscopy. Am J Physiol Cell Physiol 282, C213C218.Google Scholar
Dickie, R., Bachoo, R.M., Rupnick, M.A., Dallabrida, S.M., DeLoid, G.M., Lai, J., DePinho, R.A. & Rogers, R.A. (2006). Three-dimensional visualization of microvessel architecture of whole-mount tissue by confocal microscopy. Microvasc Res 72, 2026.CrossRefGoogle ScholarPubMed
Ferkowicz, M.J. & Yoder, M.C. (2011). Whole embryo imaging of hematopoietic cell emergence and migration. In Stem Cell Migration, Filippi, M.-D. & Geiger, H. (Eds.), pp. 143155. New York: Humana Press.Google Scholar
Gerritsen, H.C. & De Grauw, C.J. (1999). Imaging of optically thick specimen using two-photon excitation microscopy. Micros Res Tech 47, 206209.Google Scholar
Gustafsson, M.G., Agard, D.A. & Sedat, J.W. (1999). I5M: 3D widefield light microscopy with better than 100 nm axial resolution. J Microsc 195, 1016.Google Scholar
Hell, S., Reiner, G., Cremer, C. & Stelzer, H.K. (1993). Aberrations in confocal fluorescence microscopy induced by mismatches in refractive index. J Microsc 169, 391405.Google Scholar
Miller, C.E., Thompson, R.P., Bigelow, M.R., Gittinger, G., Trusk, T.C. & Sedmera, D. (2005). Confocal imaging of the embryonic heart: How deep? Microsc Microanal 11, 216223.Google Scholar
Pawley, J.B. (2002). Limitations on optical sectioning in live-cell confocal microscopy. Scanning 24, 241246.Google Scholar
Theer, P. & Denk, W. (2006). On the fundamental imaging-depth limit in two-photon microscopy. J Opt Soc Am A 23, 31393149.Google Scholar
Tuchin, V.V. (2005a). Optical Clearing of Tissues and Blood. Bellingham, WA: SPIE Publications.Google Scholar
Tuchin, V.V. (2005b). Optical clearing of tissues and blood using the immersion method. J Phys D 38, 24972518.Google Scholar
Tuchin, V.V., Altshuler, G.B., Gavrilova, A.A., Pravdin, A.B., Tabatadze, D., Childs, J. & Yaroslavsky, I.V. (2006). Optical clearing of skin using flash lamp-induced enhancement of epidermal permeability. Lasers Surg Med 38, 824836.Google Scholar