Objective:To investigate the similarities and differences in amplification multiples,biological phenotypes and cytotoxicity of human natural killer cell(NK cell)derived from human peripheral blood mononuclear cells activated and expanded by three in vitro culture systems.Methods:Collect peripheral blood donated by healthy donors and enrich mononuclear cells using density gradient centrifugation mediated by lymphocyte separation solution.Using serum free culture medium containing autologous plasma and recombinant human cytokines,IL-2 and IL-15 are added to each system,anti-human CD3 monoclonal antibody is added to system B,and IL-18 is added to system C.Culture and amplify NK cells under the same conditions for 14 days,and compare the differences in cell phenotype,cell viability,and cytotoxicity of NK cells from different culture systems.Results:In three in vitro culture systems,the amplification ratio of NK cells in the system A,system B and system C are 811.43±30.07,2800.79±36.42,and 1047.97±85.21,respectively,which means anti-human CD3 monoclonal antibodies can enhance the amplification of NK cells.There are no statistically significant differences in cell phenotypes,cell cycle,cell viability,and cytotoxicity among the three systems for cultivating NK cells.Conclusion:Using a serum-free culture medium containing autologous plasma,recombinant human IL-2,IL-15 cytokines,and anti-human CD3 monoclonal antibodies,large-scale NK cell culture can be achieved at 37℃and 5%CO2 to obtain high-purity NK cells.
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