Objective Previous studies have shown that ulcerative colitis (UC) is associated with the presence of lamina propria non-invariant (Type II) NKT cells producing IL-13 and mediating epithelial cell cytotoxicity. Here we sought to define the antigen(s) stimulating the NKT cells and to quantitate these cells in the UC lamina propria. Design Detection of Type II NKT cells in UC lamina propria mononuclear cells (LPMC) with lyso-sulfatide loaded tetramer and quantum dot-based flow cytometry and staining. Culture of UC LPMCs with lyso-sulfatide glycolipid to determine sulfatide induction of epithelial cell cytotoxicity, IL-13 production and IL-13R alpha 2 expression. Blinded quantum dot-based phenotypic analysis to assess UC LPMC expression of IL-13R alpha 2, CD161 and IL-13. Results Approximately 36% of UC LPMC were lysosulfatide tetramer positive, whereas few, if any, control LPMCs were positive. When tested, the positive cells were also CD3 and IL-13R alpha 2 positive. Culture of UC LPMC with lyso-sulfatide glycolipid showed that sulfatide stimulates UC LPMC production of IL-13 and induces UC CD161 LPMC-mediated cytotoxicity of activated epithelial cells; additionally, lyso-sulfatide induces enhanced expression of IL-13R alpha 2. Finally, blinded phenotypic analysis of UC LP MC using multicolour quantum dot-staining technology showed that approximately 60% of the LPMC bear both IL-13R alpha 2 and CD161 and most of these cells also produce IL-13. Conclusions These studies show that UC lamina propria is replete with Type II NKT cells responsive to lyso-sulfatide glycolipid and bearing IL-13R alpha 2. Since lyso-sulfatide is a self-antigen, these data suggest that an autoimmune response is involved in UC pathogenesis.