The invention provides a ferric ion detection reagent, and a preparation method and application thereof. The ferric ion detection reagent provided by the invention is characterized by high crystalline degree, phase stability, spherical nano-particle, average grain size of 418nm, uniform distribution and regular appearance. Fe<3+> has higher fluorescence quenching to Tb2O2SO4:Er, Yb as high as 74.23%, so that the ferric ion detection reagent can be used for detecting Fe<3+>. Besides, the ferric ion detection reagent has higher biocompatibility, is easily eaten by cells, and has less toxic and side effects.