The invention relates to the technical field of microculture, in particular to a culture medium for culturing fat stem cells and a preparation method thereof. The culture medium comprises a basic culture medium and additives, and the additives include peptone, sodium chloride, glucose, lactose, phosphorus source, pig gall salt, mannitol, sodium pyruvate, amino acid, vitamin, antibiotic, metal salt, plant polysaccharide and distilled water. The culture medium contains the plant polysaccharide, thereby being capable of inhibiting growth of other cells except for fat stem cells and promoting proliferation of the fat stem cells and conducive to increasing concentration of the fat stem cells in the culture medium and shortening culture time; through synergism among ingredients, sufficient nutrition and good environment needed for cell growth and proliferation can be provided, cell growth can be promoted, albumin synthesis can be promoted, and cell activity can be maintained for a long time.