AbstractHigh-nuclear lanthanide clusters have shown great potential for the administration of high-dose mononuclear gadolinium chelates in magnetic resonance imaging (MRI). The development of high-nuclear lanthanide clusters with excellent solubility and high stability in water or solution has been challenging and is very important for expanding the performance of MRI. We usedN-methylbenzimidazole-2-methanol (HL) and LnCl3·6H2O to synthesize two spherical lanthanide clusters, Ln32(Ln = Ho, Ho32; and Ln = Gd, Gd32), which are highly stable in solution. The 24 ligands L−are all distributed on the periphery of Ln32and tightly wrap the cluster core, ensuring that the cluster is stable. Notably, Ho32can remain highly stable when bombarded with different ion source energies in HRESI-MS or immersed in an aqueous solution of different pH values for 24 h. The possible formation mechanism of Ho32was proposed to be Ho(III), (L)−and H2O → Ho3(L)3/Ho3(L)4→ Ho4(L)4/Ho4(L)5→ Ho6(L)6/Ho6(L)7→ Ho16(L)19→ Ho28(L)15→ Ho32(L)24/Ho32(L)21/Ho32(L)23. To the best of our knowledge, this is the first study of the assembly mechanism of spherical high-nuclear lanthanide clusters. Spherical cluster Gd32, a form of highly aggregated Gd(III), exhibits a high longitudinal relaxation rate (1 T,r1= 265.87 mM−1·s−1). More notably, compared with the clinically used commercial material Gd-DTPA, Gd32has a clearer and higher-contrastT1-weighted MRI effect in mice bearing 4T1 tumors. This is the first time that high-nuclear lanthanide clusters with high water stability have been utilized for MRI. High-nuclear Gd clusters containing highly aggregated Gd(III) at the molecular level have higher imaging contrast than traditional Gd chelates; thus, using large doses of traditional gadolinium contrast agents can be avoided.