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With the use of fluorescent cells labeled with green fluorescent protein (GFP) in the nucleus and red
fluorescent protein (RFP) in the cytoplasm and a highly sensitive small animal imaging system with both
macro-optics and micro-optics, we have developed subcellular real-time imaging of cancer cell trafficking in
live mice. Dual-color cancer cells were injected by a vascular route in an abdominal skin flap in nude mice.
The mice were imaged with an Olympus OV100 small animal imaging system with a sensitive CCD camera
and four objective lenses, parcentered and parfocal, enabling imaging from macrocellular to subcellular. We
observed the nuclear and cytoplasmic behavior of cancer cells in real time in blood vessels as they moved by
various means or adhered to the vessel surface in the abdominal skin flap. During extravasation, real-time
dual-color imaging showed that cytoplasmic processes of the cancer cells exited the vessels first, with nuclei
following along the cytoplasmic projections. Both cytoplasm and nuclei underwent deformation during
extravasation. Different cancer cell lines seemed to strongly vary in their ability to extravasate. We have also
developed real-time imaging of cancer cell trafficking in lymphatic vessels. Cancer cells labeled with GFP
and/or RFP were injected into the inguinal lymph node of nude mice. The labeled cancer cells trafficked
through lymphatic vessels where they were imaged via a skin flap in real-time at the cellular level until they
entered the axillary lymph node. The bright dual-color fluorescence of the cancer cells and the real-time
microscopic imaging capability of the Olympus OV100 enabled imaging the trafficking cancer cells in both
blood vessels and lymphatics. With the dual-color cancer cells and the highly sensitive imaging system
described here, the subcellular dynamics of cancer metastasis can now be observed in live mice in real time.
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