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A. Jayagopal, P.K. Russ, F.R. Haselton; Quantum Dot–Based in vivo Retinal Imaging of Inflammation . Invest. Ophthalmol. Vis. Sci. 2006;47(13):3238.
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The principal cellular and molecular mediators involved in inflammation are numerous, and their specific roles have not been completely elucidated. In vivo imaging strategies that are currently used to probe inflammatory activity for the development of diagnostic techniques and the discovery of potential therapeutic targets are often limited by low signal to noise ratios, invasiveness, low spatial and temporal resolution, and an inability to image multiple cellular or molecular targets simultaneously. High–intensity quantum dot nanocrystals may be suitable candidates for monitoring real–time cell trafficking and molecular expression in vivo. We explored the utility of a fluorescence microscopy–based in vivo imaging technique for the real–time imaging of leukocyte subtypes and cell adhesion molecules in a rat model of diabetes.
The retina was used to non–invasively probe inflammatory activity in the circulation, using quantum dot–antibody conjugates directed toward the endothelial cell surface proteins PECAM–1, VCAM–1, and ICAM–1, and specific leukocyte cell surface markers. Bioconjugates were selected for minimal spectral overlap to enable multispectral imaging applications. Nanoparticles and antibodies were engineered using polymeric and biological masking strategies to reduce clearance and immunogenicity. Quantum dot probes were systemically injected by tail vein catheterization, and retinal imaging of streptozotocin–treated diabetic rats and untreated controls was performed using a high–speed digital acquisition system in conjunction with inverted fluorescence microscopy.
We report elevated fluorescence emission intensities due to endothelial cell–targeted quantum dot conjugates in the retinal circulation of STZ–treated rats relative to untreated controls. Immunofluorescence analysis is negative for nonspecific binding to endothelial linings and leukocyte surfaces and correlate with in vivo observations. Multispectral fluorescence imaging was readily achieved with this technique in both major vessels and the microcirculation, enabling the real–time visualization of molecular expression and cellular trafficking.
Quantum dots can be used in conjunction with in vivo fluorescence microscopy to characterize complex leukocyte–endothelial cell interactions with high signal to noise ratios. Such multi–pronged approaches which examine both the cells and molecules involved in disease throughout its development are likely to have a major impact on the development of detailed, early diagnostic techniques, as well as the identification of promising therapeutic targets.
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