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Direct labeling and visualization of blood vessels with lipophilic carbocyanine dye DiI.

Nature protocols (2008-10-11)
Yiwen Li, Ying Song, Lian Zhao, Gabriel Gaidosh, Alan M Laties, Rong Wen
ABSTRACT

We describe a protocol to rapidly and reliably visualize blood vessels in experimental animals. Blood vessels are directly labeled by cardiac perfusion using a specially formulated aqueous solution containing 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI), a lipophilic carbocyanine dye, which incorporates into endothelial cell membranes upon contact. By lateral diffusion, DiI also stains membrane structures, including angiogenic sprouts and pseudopodial processes that are not in direct contact. Tissues can be immediately examined by conventional and confocal fluorescence microscopy. High-quality serial optical sections using confocal microscopy are obtainable from thick tissue sections, especially at low magnification, for three-dimensional reconstruction. It takes less than 1 h to stain the vasculature in a whole animal. Compared with alternative techniques to visualize blood vessels, including space-occupying materials such as India ink or fluorescent dye-conjugated dextran, the corrosion casting technique, endothelial cell-specific markers and lectins, the present method simplifies the visualization of blood vessels and data analysis.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Glycerol, BioXtra, ≥99% (GC)
Sigma-Aldrich
Sodium phosphate dibasic, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium phosphate monobasic, BioXtra, ≥99.0%
Sigma-Aldrich
Sodium chloride, BioXtra, ≥99.5% (AT)
Sigma-Aldrich
1,1′-Dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate, BioReagent, suitable for fluorescence, ≥98.0% (TLC)