Zebrafish has been used as a powerful model system in biological and biomedical studies studying development and diseases. Comparative, functional, and developmental studies on zebrafish morphology require precise visualization of 3D morphological structures. Few methods that can visualize whole-volume of zebrafish tissues are available because optical bio-imaging methods are limited by pigmentation and hard tissues. To overcome these limitations, the 3D microstructures of a hypercholesterolemic zebrafish model are visualized using synchrotron X-ray micro-computed tomography (SR-μCT). The model spatial resolution ranged from sub- to several microns. The microstructures of various zebrafish organs are observed by combining high-contrast staining (osmium tetroxide and uranyl acetate) and embedding a protocol to enhance the image contrast of soft tissues. Furthermore, blood vessels are identified using a barium sulfate injection technique. The internal organs and cells, such as liver, intestine, oocytes, and adipocytes, of a hypercholesterolemic zebrafish are compared with those of normal organs and cells. The SR-μCT is useful for understanding the pathogenesis of circulatory vascular diseases by detecting the modifications in the 3D morphological structures of the whole body of the zebrafish. This bio-imaging technique can be readily used to study other disease models.