In vivo identification of glycolipid antigen-specific T cells using fluorescent CD1d tetramers

The CD1 family of major histocompatibility complex (MHC)-like molecules spe cializes in presenting lipid and glycolipid antigens to alpha/beta T lympho cytes, but little is known about the size of the CD1-restricted T cell popu lation or the frequency of T lymphocytes specific for a given glycolipid an tigen. Here, we report the generation and use of mouse CD1d1-glycolipid tet ramers to visualize CD1d-restricted T cells. In contrast with previous BIAc ore-based estimates of very short half-lives for CD1d-glycolipid complexes, we found that the dissociation rate of several different CD1d-glycolipid c omplexes was very slow. Fluorescent tetramers of mouse CD1d1 complexed with alpha-galactosylceramide (alpha GalCer), the antigen recognized by mouse V alpha 14-J alpha 281/V beta 8 and human V alpha 24-J alpha Q/V beta 11 nat ural killer T (NKT) cell T cell receptors (TCRs), allowed us for the first time to accurately describe, based on TCR specificity, the entire populatio n of NKT cells in vivo and to identify a previously unrecognized population of NK1.1-negative "NKT" cells, which expressed a different pattern of inte grins. In contrast, natural killer (NK) cells failed to bind the tetramers either empty or loaded with alpha GalCer, suggesting the absence of a CD1d- specific, antigen-nonspecific NK receptor. Mouse CD1d1-alpha GalCer tetrame rs also stained human NKT cells, indicating that they will be useful for pr obing a range of mouse and human conditions such as insulin-dependent diabe tes mellitus, tumor rejection, and infectious diseases where NKT cells play an important role.