H-2-restricted antigen binding by a hybridoma clone that produces antigen-specific helper factor

Abstract

Somatic cell hybrids were prepared by fusing the AKR mouse lymphoma BW-5147 with splenic T cells from mice immunized with 4-hydroxy-3-nitrophenylacetic acid (NP) conjugated to chicken serum globulin (CG). From 500 fusion lines 11 were selected on the basis of binding radioiodinated NP-CG. The autoradiographic binding assay was based on previous findings which showed that Lyt-1+ T cells need a lymphokine, lymphocyte-activating factor (LAF), for optimal antigen binding and that they bind preferentially a self-Ia-associated antigen complex, IAC, which is released by adherent cells upon incubation with antigen. Six of the 11 antigen-binding positive lines were tested for helper activity and specific helper factor production in vitro. All of them were found to be positive. One clone was characterized in more detail. It secretes a CG-specific helper factor that contains immunoglobulin heavy chain variable region and I-A determinants. The hybridoma cells bind Ia-containing CG complexes specifically. For binding they need to be treated with LAF, and the binding is restricted to syngenicity in H-2 between the adherent cells used to produce IAC and the antigen-binding hybridoma cells. Regular CG does not bind significantly and does not compete even at high excess with the binding of CG-IAC. These data are interpreted to suggest that the antigen is bound by cells of a clone functional helper T-cell hybridoma line in conjunction with products controlled by H-2I and that the receptor of these cells may have considerably higher affinity for Ia-associated than for regular antigen.