The number of circulating tumor cell (CTC) in the peripheral blood of cancer patients can be a valuable biomarker for cancer diagnosis and treatment monitoring. In this study, we implemented a custom-design video-rate confocal microscopy system in capable of direct visualization of fast flowing CTC at great saphenous vein (GSV) of a live animal model in vivo. Continuous acquisition of video-rate images at GSV revealed the highly dynamic time-dependent changes in the number of intravenously injected circulating tumor cells. By extracting a calibration factor through the hemocytometric analysis of intravenously injected long-circulating red blood cells, we established a novel quantitation method for CTC in whole body blood in vivo.