Generation and characterisation of CD1d tetramer produced by a lentiviral expression system.
The alpha-galactosylceramide (alphaGalCer)-loaded CD1d tetramer remains the most powerful tool in identifying natural killer T (NKT) cells, a subpopulation of T cells that express an unusual semi-invariant T cell antigen receptor, and mediate a variety of proinflammatory and immunoregulatory functions. The difficulty of generating large amounts of the alphaGalCer-CD1d tetramer has limited its availability and consequently hampered the study of NKT cells. In this report, we used a lentiviral system to generate stable cell lines producing beta2m-CD1d single chain protein in large quantities and in a relatively short period of time. When the protein was loaded with alphaGalCer and tetramerised with fluorescence-labelled streptavidin, its ability to efficiently bind to NKT cells was confirmed both by phenotype analysis and functional study. The CD1d tetramer generated from these stable cell lines should facilitate a wide range of studies on the biology and clinical applications of CD1d-restricted NKT cells.
Li
Demin
D
Medical Research Council Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Headington, Oxford, UK.
Chen
Nan
N
McMichael
Andrew J
AJ
Screaton
Gavin R
GR
Xu
Xiao-Ning
XN
D018949:Antigens, CD1 / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D055610:Antigens, CD1d; D019921:Biotinylation; D017252:CD3 Complex / Q000032:analysis; D015704:CD4 Antigens / Q000032:analysis; D002460:Cell Line; D002469:Cell Separation; D003001:Cloning, Molecular; D003470:Culture Media / Q000378:metabolism; D005434:Flow Cytometry; D005699:Galactosylceramides / Q000378:metabolism*; D006801:Humans; D016130:Immunophenotyping; D007694:Killer Cells, Natural / Q000276:immunology*; D016086:Lentivirus / Q000235:genetics* / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000276:immunology*; D010641:Phenotype; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D001613:beta 2-Microglobulin / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification
D018949:Antigens, CD1; D055610:Antigens, CD1d; C526877:CD1D protein, human; D017252:CD3 Complex; D015704:CD4 Antigens; D003470:Culture Media; D005699:Galactosylceramides; D016693:Receptors, Antigen, T-Cell, alpha-beta; D011993:Recombinant Fusion Proteins; C493154:alpha-galactosylceramide; D001613:beta 2-Microglobulin