An automated fluorescence image acquisition and analysis system has been develop to collect blood flow data by imaging the 3-dimensional locations of individual fluorescent microspheres in tissue. Applications to blood flow measurements in rat hearts and mouse lungs will be presented. A motor-driven microtome serial sections frozen tissue blocks in steps of 40 µm. Fluorescence images (2kx2k pixels) of the exposed face of the tissue block are acquired using a CCD camera with filtered excitation and emission. Filter wheels permit the use of up to five different colors of microspheres. Approximately 500 images of each of two microspheres colors were analyzed per rat heart yielding 3-dimensional locations of all microspheres (~10,000 each color) lodged in the organ. Regional flow measurements were calculated from microsphere location for tissue volume elements of several sizes and shapes. This instrument is automated, eliminates much of the labor, time an consumable supplies required for traditional indirect measurements, while providing comparable precision.
Research supported by grant HL50898-02 from the National Institutes of Health.
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