Synthesis of magnetic particles with superparamagnetism, high magnetization, excellent biocompatibility and small particle size is highly desirable for biomedical applications. Herein, we report that sub-100 nm biocompatible Fe₃O₄ nanocrystal clusters can be prepared via a solvothermal method by using water as a size-control agent and sodium citrate as a stabilizer. The resultant Fe₃O₄ particles have uniform spherical shape, superparamagnetic properties, high magnetization values, excellent colloidal stability and good biocompatibility. Notably, the particle size can be tuned over a range of 70–180 nm by changing the content of H₂O in the system. Moreover, the T₂-weighted magnetic resonance imaging (MRI) experiments show that 85 nm Fe₃O₄ particles have an ultra-high r₂ relaxivity of 175.4 mM⁻¹ s⁻¹, suggesting a great potential as MRI probes. In addition, the labeling efficiency of mesenchymal stem cells with as-obtained Fe₃O₄ particles is much higher than that of commercial superparamagnetic iron oxide nanoparticles, which further indicates their promising in biomedicine.