Abstract
This protocol outlines methods to measure vascular parameters, including angiogenesis (e.g., vascular density, length, diameter) and hemodynamics (e.g., erythrocyte velocity), in tumors and normal vascular networks in mice. Fluorescein-isothiocyanate (FITC)-dextran is injected into the tail vein of mice to visualize microvessels within 150 μm (using single-photon microscopy) or ~600 μm (using multiphoton laser-scanning microscopy [MPLSM]) of a tumor/window interface. Randomly selected areas (three to six locations/tumor or animal) are investigated using long-working-distance objectives with appropriate magnification. During each observation period, FITC-fluorescence images are recorded for 60 sec, and the videotapes are analyzed off-line for single-photon microscopy; or a three-dimensional (3D) image stack of the vessel network is generated, and vessel properties are measured for MPLSM. If desired, red blood cell (RBC) flux can be measured on a vessel-by-vessel basis using fluorescent tracer RBCs.
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