The invention discloses a preparation method of CIK (cytokine-induced killer) blocked by PD-1 for cancer therapy and belongs to the field of medical treatment. The preparation method includes the coating step, the activating step, the cleaning step, the blocking step and the filtering step. In the coating step, an anti-human CD3 monoclonal antibody is coated in a cell culture flask; in the activating step, the CD3 monoclonal antibody, IFN gamma and IL-2 are utilized to activate and culture a lymphocyte; in the cleaning step, the lymphocyte which is activated and cultured by three times is cleaned; in the blocking step, the cleaned lymphocyte is suspended by normal saline to obtain suspended cell solution, the PD-1 monoclonal antibody is added into the suspended cell solution, and then suspending incubation is carried out; in the filtering step, the cell suspension obtained by incubation of the PD-1 monoclonal antibody is filtered and then is refilled with normal saline to obtain feedback cell suspension. The CIK blocked by PD-1 has the advantages of low consumption of the PD-1 monoclonal antibody, good blocking effect of PD-1 molecules, high killing activity and the like, and also has the advantages of operational simplicity, low preparation cost and convenience in clinical application.