OBJECTIVE This study was performed to assess the imaging characteristics and pharmacokinetics of 1,3-Bis-[7-(3-amino-2,4,6-triiodophenyl)-heptanoyl]-2-oleoyl glycerol (DHOG, Fenestra LC), a hepatobiliary contrast agent for microCT. METHODS We investigated the abdomen of 18 female C3H mice in a MicroCAT II microCT scanner before contrast agent injection and at multiple time points up to 48 hours after intravenous injection of DHOG (1 g I/kg body weight). The contrast agent effect was determined quantitatively and dynamically by measuring pre- and postcontrast Hounsfield units (HU) of the liver, aorta, spleen, and kidneys. Based on additional phantom measurements, the reproducibility of lesion detection was estimated for different lesion sizes. RESULTS DHOG caused a marked early postcontrast enhancement of blood in the aorta and a very high enhancement of the spleen, both slowly declined after 90 minutes. The liver parenchyma showed a slow contrast agent accumulation and clearly increased HU data between 3 and 7 hours after injection. No significant renal parenchymal enhancement or excretion was noticed. At early time points after administration, DHOG exhibits characteristics of a macromolecular contrast agent by demonstrating a blood pool effect. At later time points, DHOG provides a prolonged, marked liver enhancement on microCT images due to its specific liver uptake. For a lesion size of 1 mm diameter, the variability in between two scans was 27.7 HU (P < .05) and the variability for different planes of one scan was 19.8 HU (P < .05). CONCLUSIONS DHOG yields a very good visualization of the liver and delineation of the surrounding structures with a long plateau. It is a very suitable contrast agent for liver imaging in mice for microCT imaging. The presented protocol provides a high reproducibility for lesion detection with a relatively low radiation dose.