A novel in vitro culture method for human primary nk cell

Abstract

This invention discloses a novel method for culturing natural killer cells in vitro without added feeder cells. The method maintains and expands natural killer cells isolated from umbilical cord blood of normal healthy volunteers using a culture medium composition. This culture medium composition comprises culture medium, heat-inactivated fetal bovine serum, antibiotics (penicillin-streptomycin), and two cytokines, IL-2 and IL-15. This culture medium composition provides a safe and functional material and method for culturing natural killer cells.

Claims

1. A culture medium composition comprising a basal medium, 5-20% heat-inactivated fetal bovine serum, 1-10% antibiotics, and 0.001-1% cytokines, the cytokines being IL-2 and IL-15.

2. The culture medium composition as claimed in claim 1, wherein the basal culture medium is a stem cell growth medium.

3. The culture medium composition as claimed in claim 1, wherein the antibiotic comprises penicillin and streptomycin.

4. The culture medium composition as claimed in claim 1, wherein the amount of IL-2 present in the culture medium composition is 0.01 to 1 U / mL, and the amount of IL-15 present in the culture medium composition is 0.001 to 0.1 U / mL.

5. The culture medium composition as claimed in claim 1, wherein the heat-inactivated fetal bovine serum is present in the culture medium composition at an amount of 4 to 6% (v / v).

6. The culture medium composition as claimed in claim 1, wherein the antibiotic is present in the culture medium composition at an amount of 0.5 to 1.5% (v / v).

7. A method for culturing natural killer cells, comprising the following steps: Step a: isolating monocytes (MNCs) from human umbilical cord blood; Step b: isolating natural killer cells from the monocytes; Step c: culturing the natural killer cells in a culture medium composition as described in claim 1; Step d: obtaining a cell population in which more than 80% are natural killer cells, the natural killer cells being CD3-CD56+ cells, wherein the amount of natural killer cells in step d is more than 1.5 times that in step b.

8. The culture method as described in claim 7, characterized in that step b is to obtain natural killer cells using a human natural killer cell isolation kit.

9. The method as described in claim 7, wherein no trophoblast cells are added.

10. The culture method as described in claim 7, wherein more than 95% of the cell population obtained in step d are natural killer cells.

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