Natural killer T (NKT) cells, a subset of T lymphocytes recognise lipid based antigens presented by the MHC class I type molecule CD1d. Two different Types of NKT cells exist in mice and humans and are thought to be phenotypically and functionally distinct. Type-1 NKT cells express a semi-invariant TCR capable of recognising the glycosphingolipid antigen α-galactosyl ceramide (α-GalCer). These cells have been shown to induce potent anti-tumour responses in mice. Much less is known about Type-2 NKT cells that lack the “invariant” TCR-α chain, express more diverse TCR-α/β chains and do not recognise α-GalCer. Previous studies suggest that Type-2 NKT cells have opposing functions to Type-1 NKT cells. For example, Type-2 NKT cells suppress the anti-tumour response mediated by Type-1 NKT cells. However, in hepatitis B virus infections in mice, Type-1 and Type-2 NKT cells act synergistically to clear the infection, albeit through the recognition of distinct antigens. In addition, Type-2 NKT cells seem to play a role in various immune diseases in humans and mice, such as colitis, parasitic infections, obesity and type-1 diabetes. Collectively, these data suggest, that humans and mice have a diverse population of CD1d-restricted Type-2 NKT cells, with unique TCR specificities and functional characteristics that allow them to respond to a broad range of lipid antigens including tumour associated antigens. These antigens include glycolipids such as sulfatide, phospholipids such as phosphatidyl-glycerol and phosphatidyl-ethanolamine, and non-lipid antigens such as PPBF (phenyl 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonate). By producing a panel of Type-2 NKT cell lines, we reveal diversity in antigen specificity and TCR usage by these cells. Furthermore, we used CD1d tetramers loaded with specific lipid antigens to identify Type-2 NKT cells from blood and tissue samples. These technologies enable us to explore Type-2 NKT cells and to determine their role in health and disease.