Shared idiotypic determinants on B and T lymphocytes reactive against the same antigenic determinants. III. Physical fractionation of specific immunocompetent T lymphocytes by affinity chromatography using anti-idiotypic antibodies.

Abstract

Anti-idiotypic antibodies made against antigen-binding receptors on T lymphocytes with specificity for certain Ag-B locus antigens selectively react with T lymphocytes with potential immune reactivity against the very same Ag-B antigens. This was shown by affinity chromatography of normal Lewis T lymphocytes on anti-Ig columns after contact with the relevant anti-idiotypic antiserum. Here, it could be shown that incubation of the cells with an anti-(Lewis-anti-BN) antiserum caused subsequent selective retention of potential graft-vs.-host (GvH)-reactive cells against BN on the anti-Ig column, whereas Lewis T cells with reactivity against DA or August (Au) (carrying distinct Ag-B antigens in comparison to BN) passed through. The retained cells could be eluted and shown to display highly increased reactivity against BN with virtually no reactivity left against DA or Au antigens. Analogous results were obtained using an anti-(Lewis-anti-DA) antiserum. The anti-idiotypic antibodies can be used in fluorescent antibody tests to directly visualize the idiotype-positive cells. Using the separation design described above we analyzed selectively enriched or deleted T lymphocytes for presence of idiotypic cells as well as specific GvH reactivity. A highly significant positive correlation was found between percentage of a given idiotype in a population of T cells and the relevant GvH potential of the same T cells that can be visualized are indeed the very same T cells that express immune reactivity against the expected antigens. The present data would thus directly demonstrate the existence of a largely nonoverlapping population of immunocompetent T cells capable of reacting against the various Ag-B locus antigens in the rat. Highly purified, functionally intact immunocompetent T lymphocytes with restricted immune reactivity can thus be produced from normal lymphocyte populations for further analysis.

CD4+ T lymphocytes mediate in vivo clearance of plasmid DNA vaccine antigen expression and potentiate CD8+ T-cell immune responses

“…14 More than 95% of macrophages were depleted as determined by a flow cytometric analysis using an anti-CD14 antibody (BD Biosciences) on days 1, 14, and 28 after plasmid DNA inoculation. CD4 ϩ T-cell 15,16 and CD8 ϩ T-cell depletions 17 were performed with either anti-CD4 (GK1.5) or anti-CD8␣ antibodies (53-6.7; BioLegend, San Diego, CA). Isotype rat IgG control (Sigma-Aldrich) was used for control experiments.…”

Induction of specific immune unresponsiveness with purified mixed leukocyte culture-activated T lymphoblasts as autoimmunogen. III. Proof for the existence of autoanti-idiotypic killer T cells and transfer of suppression to normal syngeneic recipients by T or B lymphocytes.

“…Earlier studies have taught us that idiotype-positive T cells can be rendered Ig positive by reaction with auti-idiotypic antibodies. Such Ig-coated T cells can then be selectively removed from other T cells via filtration through anti-Ig bead column and the retained cells can be recovered in a functionally intact form by using mechanical elution (12). In principle, it should be possible to do the very same thing with anti-idiotypic T cells, that is, incubating the present suppressor T cells from C57BL/6 mice immunized with autoblasts of anti-CBA specificity with conventional C57BL/6-anti-CBA alloantiserum followed by fractionation of the cells over a second anti-Ig column (the T cells were prepared by filtration over a first anti-Ig column before incubation with the alloantiserum).…”

“…In exp. 9, a crucial experiment was included by applying principles that were previously fruitful when attempting to fractionate T lymphocytos according to their idiotypic receptors, allowing recovery of T cells in a functionally intact form (12). Here, T cells from the blast-immunized animals were incubated with idiotypic C57BL/6-anti-CBA alloantiserum.…”

T cell receptors with allo-major histocompatibility complex specificity from rat and mouse. Similarity of size, plasmin susceptibility, and localization of antigen-binding region.

“…Macrophages were harvested from the peritoneal cavity of F1 rats, washed once, and irradiated with 2,500 rad. Macrophages obtained from one rat were distributed over five 3013 Falcon tissue culture flasks and cultured in 5 ml EHAA medium, complemented with 10% fetal calf serum (FCS) and 20% TCGF for 24 h. The anti-idiotypic nature of the antiserum pool 3 of specificity anti-(Lewis anti-DA) has been published in detail (32). The Ig fraction of this antiserum was coupled to cyanogen bromide (CNBr)-activated Sepharose 4B following the instructions of the manufacturers (Pharmacia Fine Chemicals).…”