OBJECTIVE: Tumor imaging via molecular magnetic resonance imaging (MRI) that uses specific superparamagnetic iron oxide particles (SPIOs) has been addressed in the literature several times in the last 20 years. To our knowledge, none of the reported approaches is currently used for routine clinical diagnostic evaluation, nor are any in clinical development. This raises questions as to whether SPIO-enhanced molecular MRI is sensitive and specific enough for use in clinical practice. The aim of our preclinical study was to investigate the minimum requirements for obtaining sensitive molecular MRI for use in tumor evaluations under optimal conditions. The well-vascularized F9 teratocarcinoma tumor model, which exhibits high levels of the highly accessible target CD105 (endoglin), was used to compare the accumulation and visualization of target-specific SPIOs by MRI. MATERIAL AND METHODS: Superparamagnetic iron oxide particles were optimized in the following ways: (a) proton relaxivity was increased for higher imaging sensitivity, (b) a coating material was used for optimal loading density of the αCD105 antibody, and (c) binding activity to the target CD105 was increased. Binding activity and specificity were confirmed in vitro using enzyme-linked immunosorbent assay and in vivo using pharmacokinetic and biodistribution studies of 11 F9 teratoma-bearing mice together with micro-autoradiography. CD105 target expression was determined using immunohistochemistry and quantitative enzyme-linked immunosorbent assay. The transverse relaxation rate R2* was quantified by 3.0-T MRI in the tumors, kidneys, and muscles before and up to 60 minutes after injection in 11 mice. The use of [Fe]-labeled SPIOs for all in vivo experiments allowed for the direct correlation of the imaging results with SPIO accumulation. RESULTS: High-relaxivity αCD105-polyacrylic acid-SPIOs (r2 up to 440 L mmol Fe s) with strong binding activity accumulated specifically in tumors (1.4% injected dose/g) and kidneys (4.1% injected dose/g) in a manner dependent on the target concentration. The accumulation occurred within the first 3 minutes after injection. Visualization of specific SPIOs was accomplished with MRI. In contrast to the successful use of MRI in all examined kidneys (mean ± SEM ΔR2*, 61 ± 11 s), only 6 of 11 tumors (mean ± SEM ΔR2*, 15 ± 7 s) showed a clear signal when compared with the control even though optimal conditions were used. CONCLUSION: The accumulation of CD105-specific SPIOs in F9 mouse teratomas was robust. However, visualization of the specifically accumulated SPIOs by MRI was not reliable because of its limited signal detection sensitivity. We postulate that it will be challenging to improve the imaging properties of targeted SPIOs further. Therefore, molecular MRI by targeted SPIOs is currently not suitable for clinical tumor imaging using routinely applicable sequences and field strength.