Generation of class I MHC-restricted T-T hybridomas.
In this report we describe a system for the generation of functional, class I MHC-restricted, T-T hybridomas. The BW5147 cell line was transfected with the CD8 gene. BW5147 transfectants were obtained that stably expressed CD8 and this expression was maintained after somatic cell hybridization with activated T lymphocytes. To determine whether the stable expression of CD8 would facilitate the generation of class I MHC-specific T-T hybridomas, the transfected cells were fused with alloreactive T cells and the resultant hybrids were screened for their ability to produce lymphokines in response to antigenic stimulation. Somatic cell hybridizations with BW5147-CD8 transfectants give rise to a much higher frequency of class I MHC-specific T-T hybridomas relative to parallel fusions with BW5147. To determine whether the BW5147-CD8 transfectants would also support the generation of Ag-specific, class I MHC-restricted T-T hybridomas, they were fused with OVA-specific CTL. Several T-T hybrid clones were identified that produced lymphokines after stimulation with a transfected APC that was synthesizing OVA, or with a tryptic digest of OVA in the presence of syngeneic APC. The stimulation by Ag was MHC-restricted and mapped to the Kb molecule. An anti-CD8 mAb inhibited the stimulation of these hybridomas by Ag plus APC, whereas their stimulation by mitogen was unaffected. Cytolytic activity was not detected when several of the OVA-specific or alloreactive hybridomas were tested for their ability to kill target cells bearing the appropriate Ag. These results demonstrate that the BW5147-CD8 transfectants allow the generation of class I MHC-restricted T-T hybridomas. The potential utility of this system is discussed.
Rock
K L
KL
Department of Pathology, Harvard Medical School, Boston, MA 02115.
Rothstein
L
L
Gamble
S
S
D000818:Animals; D000945:Antigens, Differentiation, T-Lymphocyte / Q000032:analysis / Q000502:physiology; D016827:CD8 Antigens; D015395:Histocompatibility Antigens Class I / Q000276:immunology*; D006825:Hybridomas / Q000276:immunology*; D008222:Lymphokines / Q000096:biosynthesis; D051379:Mice; D008815:Mice, Inbred Strains; D010641:Phenotype; D013601:T-Lymphocytes / Q000276:immunology*; D014162:Transfection
D000945:Antigens, Differentiation, T-Lymphocyte; D016827:CD8 Antigens; D015395:Histocompatibility Antigens Class I; D008222:Lymphokines