T cell activation by preformed, long-lived Ia-peptide complexes. Quantitative aspects.
Planar membrane-bound complexes between a fluorescent peptide, FITC-OVA(323-339), and the class II MHC Ag, I-Ad, were analyzed by fluorescence microscopy and in biological assays to determine the optimum distance between peptide-Ia complexes required for maximum activation of IL-2 production by the Th cell hybridoma DO-11.10. Optimum responses were obtained when the average distance between peptide-Ia complexes was of the order of 200 A. This implies that T cell activation by Ag-MHC requires cross-linking of the TCR via closely packed Ag-MHC complexes. The same dose response curve to the preformed complexes was obtained whether one used a fixed concentration of Ia and varied the peptide concentration or a fixed concentration of peptide and varied the Ia concentration. In both cases there was a linear relationship between the number of peptide-Ia complexes and the response of the T cells. The association between Ia and peptide in vitro is an inefficient process, requiring prolonged incubation and a large excess of peptide over Ia. Once formed, however, the complex is extremely stable with no detectable dissociation at neutral pH after days at 4 degrees C. With several different preparations of Ia it was found that only about 10 to 20% of the purified Ia is capable of forming the long-lived complex with peptide.
D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D016650:Fluorescein-5-isothiocyanate; D005452:Fluoresceins; D000949:Histocompatibility Antigens Class II / Q000032:analysis* / Q000276:immunology; D007376:Interleukin-2 / Q000096:biosynthesis; D008213:Lymphocyte Activation; D046911:Macromolecular Substances; D051379:Mice; D010047:Ovalbumin / Q000276:immunology / Q000378:metabolism; D010446:Peptide Fragments / Q000276:immunology / Q000378:metabolism*; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D013861:Thiocyanates
D005452:Fluoresceins; D000949:Histocompatibility Antigens Class II; D007376:Interleukin-2; D046911:Macromolecular Substances; D010446:Peptide Fragments; D013861:Thiocyanates; D010047:Ovalbumin; D016650:Fluorescein-5-isothiocyanate