The invention relates to the field of
stem cell biology, and discloses a preparation method of natural killer cells differentiated from human pluripotent stem cells. The preparation method comprises the steps of S1, forming an
embryoid body; S2, performing directional permanent hematopoiesis differentiation on the
embryoid body to obtain hematopoietic precursor cells; and S3, differentiating the hematopoietic precursor cells to obtain natural killer cells. The invention also discloses application of the
natural killer cell prepared by the method as a pharmaceutical composition for preventing and / or treating tumors by cells. According to the method, efficient and stable differentiation of the NK cells is achieved, EB adherent differentiation is achieved in the induced differentiation period, TGFB inhibitor treatment is prolonged, 10% or above of CD34+CD45+permanent hematopoietic HPCs can be obtained through rapid induction. The method has the advantages of being efficient, stable, low in cost and suitable for large-scale
cell preparation production; and the method is easy and convenient to operate, the NK cells obtained through differentiation have typical NK
cell surface marking molecules, the anti-tumor function of the NK cells is superior to that of the NK cells derived from
umbilical cord blood, and great scientific research and clinical application potentials are achieved.