title: Isolation and functional use of human NKT cells.
abstract: This unit details methods for the isolation, in vitro expansion, and functional characterization of human iNKT cells. The term iNKT derives from the fact that a large fraction of murine NKT cells recognize the MHC class I-like CD1d protein, are CD4+ or CD4-CD8- (double negative), and use an identical "invariant" TCRalpha chain, which is generated by precise Valpha14 and Jalpha281 (now renamed Jalpha18) rearrangements with either no N-region diversity or subsequent trimming to nearly identical amino-acid sequence (hence, 'iNKT'). Basic Protocol 1 and Alternate Protocol 1 use multi-color FACS analysis to identify and quantitate rare iNKT cells from human samples. Basic Protocol 2 describes iNKT cell purification. Alternate Protocol 2 describes a method for high-speed FACS sorting of iNKT cells. Alternate Protocol 3 employs a cell sorting approach to isolate iNKT cell clones. A Support Protocol for secondary stimulation and rapid expansion of iNKT cells is also included. Basic Protocol 3 explains functional analysis of iNKT.
authors.affiliation: Cancer Biology Program, Hematology-Oncology Division, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, USA.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Exley
authors.forename: Brian
authors.initials: B
authors.lastname: Wilson
authors.forename: Steven P
authors.initials: SP
authors.lastname: Balk
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism*; D018929:Cell Culture Techniques / Q000379:methods; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods*; D002999:Clone Cells; D016207:Cytokines / Q000378:metabolism; D003602:Cytotoxicity, Immunologic; D005434:Flow Cytometry; D006801:Humans; D051379:Mice; D055611:Natural Killer T-Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism*
chemical_list: D015703:Antigens, CD; D016207:Cytokines