title: A high-efficiency system of natural killer cell cloning.
abstract: 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 10(5) cells and as high as 10% for longer term cultures when deriving more than 2 x 10(6) 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.
authors.affiliation: Department of Medical Microbiology, Wales College of Medicine, Cardiff University, UK.
authors.forename: Rebecca J
authors.initials: RJ
authors.lastname: Morris
authors.forename: Lee K
authors.initials: LK
authors.lastname: Chong
authors.forename: Gavin W G
authors.initials: GW
authors.lastname: Wilkinson
authors.forename: Eddie C Y
authors.initials: EC
authors.lastname: Wang
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002999:Clone Cells / Q000737:chemistry / Q000166:cytology / Q000276:immunology; D018920:Coculture Techniques / Q000379:methods; D003470:Culture Media / Q000494:pharmacology; D003601:Cytotoxicity Tests, Immunologic; D005434:Flow Cytometry; D006801:Humans; D007376:Interleukin-2 / Q000494:pharmacology; D020014:K562 Cells; D007694:Killer Cells, Natural / Q000737:chemistry / Q000166:cytology* / Q000276:immunology; D007963:Leukocytes, Mononuclear / Q000166:cytology; D016853:Muromonab-CD3 / Q000494:pharmacology; D051920:NK Cell Lectin-Like Receptor Subfamily D / Q000032:analysis; D011971:Receptors, Immunologic / Q000032:analysis; D055607:Receptors, Natural Killer Cell
chemical_list: D003470:Culture Media; D007376:Interleukin-2; D016853:Muromonab-CD3; D051920:NK Cell Lectin-Like Receptor Subfamily D; D011971:Receptors, Immunologic; D055607:Receptors, Natural Killer Cell