title: Natural killer cell proliferation is dependent on human serum and markedly increased utilizing an enriched supplemented basal medium.
abstract: We studied the effects of basal medium and human serum on the ex vivo expansion of CD56+/CD3- natural killer cells (NK). We demonstrated that sorted NK cultured for 18 days with 10% human AB serum without accessory cells in a 2:1 DMEM/F12 basal medium expanded significantly greater (6.4 +/- 0.9-fold, n = 14) than when cultured in standard RPMI 1640 basal medium (3.7 +/- 0.67-fold, n = 16; p = 0.019). Supplementation of the DMEM/F12 mixture with 2-mercaptoethanol (2ME), ethanolamine, L-ascorbic acid, and sodium selenite significantly augmented NK proliferation (16.3 +/- 2.5-fold, n = 11; p < 0.001) compared with DMEM/F12 without supplements. NK growth kinetics demonstrated that both the growth rate and the duration of exponential growth were increased by medium supplements. Addition of 2ME-containing supplements to cultures of NK with accessory cells augments NK proliferation, recruits NK progenitors, and has a serum-sparing effect. For the large-scale expansion of NK, we demonstrate a greater than 30-fold increase (n = 7) in the number of activated natural killer cells (ANK) derived from CD5/CD8-depleted PBMNC when cultured for 35 days in supplemented DMEM/F12 versus RPMI 1640 basal medium (197.6 +/- 95.0-fold versus 6.3 +/- 2.1-fold, respectively). Serum-free media (AIM-V and X-VIVO 10) were unable to support NK proliferation. These data suggest that utilizing 2:1 DMEM/F12 + 2ME-containing supplements instead of standard RPMI 1640 as a basal medium can increase the proliferation of cytotoxic NK while reducing the culture interval and the amount of serum needed.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Medicine, University of Minnesota, Minneapolis 55455, USA.
authors.forename: B A
authors.initials: BA
authors.lastname: Pierson
authors.forename: P B
authors.initials: PB
authors.lastname: McGlave
authors.forename: W S
authors.initials: WS
authors.lastname: Hu
authors.forename: J S
authors.initials: JS
authors.lastname: Miller
mesh_terms: D000328:Adult; D001790:Blood Physiological Phenomena; D018929:Cell Culture Techniques / Q000379:methods*; D002455:Cell Division / Q000187:drug effects; D003470:Culture Media; D006801:Humans; D007694:Killer Cells, Natural / Q000166:cytology / Q000187:drug effects*; D007700:Kinetics; D008875:Middle Aged
chemical_list: D003470:Culture Media