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A single‐step enrichment method for generating highly purified mouse NK cell populations by negative selection
Author(s) -
Tran Jimmy,
Kung Sam K.P.,
Yuan Ning,
Milton Graeme T,
Wognum Albertus W,
Eaves Allen C,
Thomas Terry E,
Guilbault Benoit G
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.672.23
Subject(s) - nkg2d , immunomagnetic separation , biotinylation , negative selection , cytotoxic t cell , cd49b , biology , lysis , microbiology and biotechnology , cell , natural killer cell , k562 cells , antibody , cd8 , interleukin 21 , immunology , antigen , in vitro , biochemistry , gene , genome
Natural Killer (NK) cells play a critical role in innate immunity and tumor immunology. They are particularly difficult to isolate because they are rare and express several markers found on other cell types. Using column‐free immunomagnetic cell separation technology (EasySep®), we sought to develop a single‐step negative selection method for NK cell isolation, where negative selection enables the isolation of cells with minimal impact on cell function. Unwanted cells were cross‐linked to magnetic particles using a highly optimized cocktail of 10 biotinylated antibodies. This enabled us to achieve NK cell purities of 92.3 ± 2.6% (defined as %CD49b + CD3 neg ; n=6). The isolated cells retained the expression of several markers associated with an NK phenotype, including NKG2D and LY49C/I/F/H. The purified cells could be readily expanded when cultured in the presence of IL‐2 (> 8‐fold expansion after 5 days, n=3). Cr 51 release assays demonstrated the cytotoxic capacity of the isolated cells, as specific lysis was detected at an effector to target ratio of 1:1 and 10:1 for both YAC‐1 cells and Daudi B lymphoma cells, the latter being more resistant to NK‐specific killing as expected (n=3). In conclusion, we present here a novel and unique enrichment method for the isolation of pure NK cells in a single step. Supported by the Manitoba Health Research Council and The Dean of Medicine Strategic Research Fund.