Method of culturing nk cells and kits containing medium additions

Abstract

A kit for use in adding to a cultural medium for culturing natural killer (NK) cells includes a B unit including a basic solution including IL-2, L-glutamine dissolved in a cell culture medium, a C1 unit including of a cytokine 1 solution including IL-12 and IL-18 dissolved in the basic solution, a C2 unit including of a cytokine 2 solution including IL-15 dissolved in the basic solution, an A1 unit including of an antibody 1 solution including an anti-CD16 antibody and an anti-CD56 antibody dissolved in the basic solution, an A2 unit including of an antibody 2 solution including the antibody 1 solution and the basic solution in a volume ratio of 1:6-10, and a D unit including of an antibody-cytokine mixed solution including an anti-CD3 antibody dissolved in the cytokine 1 solution.

What is claimed is: 
     
         1 . A kit for use in adding to a cultural medium for culturing natural killer (NK) cells, the kit comprising:
 a B unit comprising a basic solution comprising IL-2, L-glutamine dissolved in a cell culture medium;   a C1 unit comprising of a cytokine 1 solution comprising IL-12 and IL-18 dissolved in the basic solution;   a C2 unit comprising of a cytokine 2 solution comprising IL-15 dissolved in the basic solution;   an A1 unit comprising of an antibody 1 solution comprising an anti-CD16 antibody and an anti-CD56 antibody dissolved in the basic solution;   an A2 unit comprising of an antibody 2 solution comprising the antibody 1 solution and the basic solution in a volume ratio of 1:6-10; and   a D unit comprising of an antibody-cytokine mixed solution comprising an anti-CD3 antibody dissolved in the cytokine 1 solution.   
     
     
         2 . The kit of  claim 1 , wherein the A1 unit is added to the lymphocyte culture medium prior to the D unit. 
     
     
         3 . The kit of  claim 1 , wherein the anti-CD16 antibody, the anti-CD56 antibody and the basic solution of the A1 unit are separately packaged and mixed during the medium addition step. 
     
     
         4 . The kit of  claim 1 , wherein the anti-CD16 antibody, the anti-CD56 antibody and the basic solution of the A2 unit are separately packaged and mixed during the medium addition step. 
     
     
         5 . The kit of  claim 1 , wherein anti-CD3 antibody and the cytokine 1 solution of the D unit are separately packaged and mixed during the medium addition step. 
     
     
         6 . The kit of  claim 1 , wherein the units are configured to be added to a lymphocyte culture medium in the order of the A1 unit, the D unit, the C1 unit, the A2 unit, the C2 unit, the A2 unit and the B unit. 
     
     
         7 . A culture medium for culturing natural killer (NK) cell, the culture medium comprising the kit of  claim 1 . 
     
     
         8 . The culture medium of  claim 7 , wherein the culture medium is obtained by culturing NK cells for 8 days or less, and the number of NK cells contained in the culture medium is 5×10 7  or more. 
     
     
         9 . The culture medium of  claim 7 , wherein the number of NK cells contained in the culture medium is 2×10 9  or more, and the ratio of NK cells is 80% or more. 
     
     
         10 . The culture medium of  claim 7 , wherein the number of NK cells contained in the culture medium is 2×10 9  or more, and the ratio of NK cells is 80% or more.