Ultrasound is a convenient non-invasive imaging modality used for the diagnosis or detection of various diseases and assessment of therapeutic effects. However, when imaging internal organs, the ultrasound probe must be handled by an operator. The ability to perform hands-free ultrasound imaging of internal organs is likely to offer an unprecedented advantage in various situations such as internal organ monitoring during exercise tests and prolonged monitoring. Toward this end, we have developed a new method of hands-free monitoring using three-dimensional (3D) ultrasound and used this method in portal vein monitoring, which is important for functional evaluation of hepatic and gastrointestinal systems. In previous studies, we developed a hand-made probe holder and used it to capture images of the portal vein, using image tracking and registration to compare the same position of the portal vein. In this study, we first used an abdomen phantom to assess image tracking qualitatively and quantitatively. After validating the method on the phantom, we monitored the portal vein in three healthy subjects using our 3D ultrasound method. Image tracking and registration of the portal veins in three subjects were successfully performed offline. Finally, respiratory analysis and vein diameter measurement were performed based on the image tracking results. The respiratory analysis quantified the respiration-induced portal vein movements. The vein diameter showed changes that might be induced by respiration and heartbeat. These results indicate that our 3D ultrasound method is a potentially useful tool for hands-free monitoring of internal organs.