Specific lymphocyte-target cell conjugate formation between tumor-specific helper T-cell hybridomas and IA-bearing RCS tumors and IE-bearing allogeneic cells. I. Role of Ia and both L3T4 and LFA-1 antigens in recognition/binding.
The studies reported here describe the feasibility of using single cell techniques with nonadherent target cells for the formation of T helper lymphocyte-target cell conjugates in an Ia recognition system. We have taken advantage of four tumor-specific T cell hybridomas lines, two of which respond only to IA-bearing RCS tumor cells of SJL/J (H-2s) origin, and the other two that respond to both RCS and IA- or IE-bearing allogeneic cells of H-2k,d haplotypes. The conjugate frequency between the T cell hybridomas and target cells was scored microscopically and was facilitated by labeling the lymphocyte with fluorescein. The frequency of conjugate formation ranged from 20 to 40% above background. Conjugate formation was antigen specific and correlated well with the hybridoma specificity determined by IL 2 responses after antigenic stimulation. The cross-reactive hybridomas formed conjugates with RCS and LPS blasts derived from CBA or DBA/2 origin, but not with cells of syngeneic or other allogeneic strains. Conjugate formation with RCS was inhibited greater than 50% with mAb directed against IAs determinants on the RCS tumor cells, and conjugate formation with allogeneic cells was blocked only with mAb directed to either IA/IEk or IA/IEd specificities directed against the alpha or beta polypeptide chain. Blocking of conjugate formation was also achieved by various mAb directed against surface membrane molecules associated with the T cell hybridomas. LFA-1 mAb inhibited significantly the formation of conjugates. However, L3T4 mAb blocked only partially the conjugates. Other antibodies directed against Lyt-1 or Thy-1.2 antigens were without blocking effect. The poor blocking observed with L3T4 mAb did not correlate with the almost complete blocking observed in the IL 2 response by the same hybridomas. These studies of the syngeneic anti-RCS tumor response directed against IA-bearing RCS showed that the conjugate assay permits mapping of tumor-associated Ia epitopes. In addition, the results of these studies demonstrate the feasibility of conjugate formation in determining the antigenic specificity of the T helper system. This assay system can be used to establish the minimal frequency of antigen-reactive cells and can divide the T helper response into multiple steps (i.e., recognition/binding, activation, proliferation, and lymphokine release) and determine the surface membrane molecules involved in recognition.
D000818:Animals; D000945:Antigens, Differentiation, T-Lymphocyte; D000951:Antigens, Neoplasm / Q000276:immunology; D000954:Antigens, Surface / Q000276:immunology*; D002448:Cell Adhesion; D000949:Histocompatibility Antigens Class II / Q000276:immunology*; D006825:Hybridomas / Q000276:immunology; D007376:Interleukin-2 / Q000276:immunology; D016169:Lymphocyte Function-Associated Antigen-1; D016403:Lymphoma, Large B-Cell, Diffuse / Q000276:immunology*; D051379:Mice; D008815:Mice, Inbred Strains; D006377:T-Lymphocytes, Helper-Inducer / Q000276:immunology*
D000945:Antigens, Differentiation, T-Lymphocyte; D000951:Antigens, Neoplasm; D000954:Antigens, Surface; D000949:Histocompatibility Antigens Class II; D007376:Interleukin-2; D016169:Lymphocyte Function-Associated Antigen-1