Production and characterization of antibody probes directed at constant regions of the alpha and beta subunit of the human T cell receptor.
Fabbi M., Acuto O., Bensussan A., Poole CB., Reinherz EL.
To generate antibodies directed at constant regions of the human T cell receptor, purified alpha and beta subunits of a human T cell antigen/major histocompatibility complex receptor from the REX tumor (Ti-REX) were isolated by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and utilized to immunize rabbits. H36 (anti-alpha subunit) and H38 (anti-beta subunit) antisera were strongly reactive with the denatured subunits and also immunoprecipitated the Ti heterodimer from 125I surface-labeled lysates of REX, inducer, suppressor and cytotoxic T cell clones, peripheral T lymphocytes and thymocytes. Moreover, immunodepletion experiments showed that such antisera recognized antigenic determinant(s) shared by all Ti molecules expressed in the thymus. Several observations were made with these anticonstant region antibodies. First, peptide map analysis showed that the T cell receptor molecules recognized by the anti-clonotype and the anti-constant region heteroantisera on a given T cell clone are identical, thus supporting the view that the T cell receptor undergoes allelic exclusion. Second, since the individual antisera were weakly cross-reactive with the other denatured subunit, such subunits probably share conserved sequences. Third, the absence of antisera reactivity with intact cells implies that most of these constant region epitopes must be obscured by associated molecules, perhaps including one or more of the 20-25-kDa T3 subunits. Fourth, the extensive difference in two-dimensional peptide maps of Ti alpha subunits from clones of differing specificities makes it likely that the subunit contributes in a major way to antigen/major histocompatibility complex binding.