Provided is a radioactive microsphere including a glass sphere core. The glass sphere core includes a first seed, a second seed and a diffusion region extending inwardly from an outer surface of the glass sphere core, with the second seed distributed in the diffusion region. The first seed and the second seed become radioactive after being activated by neutrons to produce radiations including β-rays or γ-rays, or simultaneously β-rays and γ-rays. A preparation method of a radioactive microsphere is also provided.
