A high-efficiency system of natural killer cell cloning

The culture of human natural killer (NK) cell clones has traditionally been a long, laborious process with an efficiency of only 1-2%. Recently, a stem cell growth medium (SCGM) has been described to expand preferentially polyclonal NK cells from peripheral blood. We have tested SCGM in a single cell sorting system and shown a 4-5 fold increase in the number of proliferating NK clones compared to standard RPMI media. The cloning efficiency was further enhanced by the provision of irradiated feeder cells derived from multiple donors combined with the addition of the anti-CD3 antibody, OKT3. The combination of SCGM, single cell sorting and these multiple optimisations enhanced NK cloning efficiency by more than tenfold to greater than 20% for short-term cultures when deriving 105 cells and as high as 10% for longer term cultures when deriving more than 2 × 106 cells. This novel system thus facilitates the generation of NK clones and allows larger scale studies of NK function that were beyond the scope of previous methodology.