Production of an antibody against guinea pig MIF. I. Specificity of the anti-MIF antibody.

An antibody was produced in rabbits against partially purified MIF which was released from the specifically stimulated lymphocytes of tuberculin-hypersensitive guinea pigs. The MIF used as an antigen was fractionated by polyacrylamide gel electrophoresis. The antibody thus prepared was then examined for its specificity for several lymphokines by affinity column chromatography. It was observed that the antibody column adsorbed MIF, but not the other three lymphokines, MCF, NCF, and SRF, indicating a keen specificity of the antibody against MIF.     

Studies on the control of antibody synthesis. IX. Effect of boosting on antibody affinity.

The effect of boosting on antibody affinity was studied in a haptenic system. Generally, boosting results in the prompt synthesis of high affinity anti-hapten antibody. However, repeated boosting frequently leads to a decrease in the amount and affinity of the serum antibody. Repeated boosting with hapten on a carrier different from that used for priming selectively stimulates synthesis of the highest affinity anti-hapten antibody and does not result in a decrease in affinity. Priming with soluble antigen without adjuvants results in the synthesis of low affinity antibody. After such priming, boosting stimulates low affinity antibody synthesis and repeated boosting leads to a moderate increase in antibody affinity.     

Use of the unlabeled antibody immunohistochemical technique for the detection of human antibody.

Two methods have been developed which permit use of the unlabeled antibody immunohistochemical technique for detection of human antibody, without the need for immunization of humans with peroxidase. Human antibody to herpes simplex virus (HSV) reacted with human cell cultures infected with HSV was the experimental system. In the first method an attempt was made to employ rabbit peroxidase-antiperoxidase (PAP) soluble complexes in connectin with human antibody. This was done by sequential addition to the HSV-infected cells of (a) human anti-HSV, (b) rabbit antihuman globulin, (c) guinea pig antirabbit globulin (the bridging reagent) and (d) rabbit PAP. Strong specific staining of HSV-infected cells was obtained; however, difficulties were encountered with nonspecific reactions on uninfected cells. In the second method PAP soluble complexes prepared with baboon antiperoxidase were bridged to the human anti-HSV antibody by rabbit antihuman globulin. Because of the phylogenetic relatedness of human and baboon globulins this resulted in firm binding which gave strong specific staining of HSV-infected cells without significant reaction in uninfected cells.     

Studies on the control of antibody synthesis. XVI. Effect of immunodepression on antibody affinity

The relationship between depression in the magnitude of the immune response and a decrease in affinity of the antibody produced was examined in three different models of immuno-depression (B-cell clonal deletion tolerance, specific suppressor T-cell activity, and anti-genie competition). In B-cell clonal deletion tolerance and in antigen-specific, suppressor T-cell-mediated immunodepression, a small decrease in magnitude (50% or less) is associated with a marked decrease in high-affinity, plaque-forming cells. In contrast, with nonspecific immunodepression, due to antigenic competition, a depression in affinity is only seen when there is a marked (85%) reduction in the magnitude of the response. The results are consistent with the view that when the mechanism of immunodepression involves interaction of antigen with antigen-specific B cells there is a disproportionate loss of high-affinity, antibody-producing cells relative to the decrease in magnitude. In contrast, with nonspecific immuno-depression, where the decrease in affinity is presumably due to inefficient expansion of high-affinity clones, an effect on affinity is only observed in association with a marked depression in the magnitude of the response.     

Reaginic antibody formation in the mouse. VII. Depression of the ongoing IgE antibody formation by suppressor T cells.

The ongoing IgE antibody formation against ovalbumin (OA) in high responder mice was depressed by i.v. injections of either native or urea-denatured ovalbumin (UD-OA). Adoptive transfer experiments to determine the helper function of spleen cells from the treated animals showed that helper function for both IgE and IgG antibody responses diminished after treatment. Evidence was obtained that treatment suppressed the expansion of IgE-G memory cells. When the same treatment with OA or UD-OA was given to OA-primed mice before the appearance of IgE antibody in their serum, OA-specific splenic suppressor T cells were demonstrable. Thus, the transfer of splenic T cells from treated mice into normal mice suppressed the primary IgE and IgG antibody responses of the recipeints to DNP-OA. It was also found that the transfer of the splenic T cells from UD-OA-treated mice into OA-primed mice depressed ongoing IgE antibody formation in the recipients. The results suggested strongly that the decrease of helper function and the depression of ongoing IgE antibody formation by repeated injections of UD-OA was caused by generation of antigen (OA)-specific suppressor T cells.     

Studies on the control of antibody synthesis. XIV. Role of T cells in regulating antibody affinity.

The effect of limiting the number of helper T cells on the affinity of the primary antibody response to a T-dependent antigen (DNP-BGG) was evaluated in a cell transfer system. Lethally irradiated, thymectomized mice were reconstituted with either bone marrow or anti-brain θ antiserum plus complement-treated spleen as the source of B cells. In addition, they received various numbers of thymus cells as a source of helper T cells. The animals were immunized with DNP-BGG 1 day after cell transfer and their splenic anti-DNP PFC response was assayed for magnitude and affinity 3 weeks later. A marked restriction in helper T-cell activity resulted in a primary response which was of low magnitude, which lacked indirect PFC, and which had a very low affinity and restricted heterogeneity. When sufficient thymus cells were given to permit a switch to indirect plaque formation, a highly heterogeneous, high-affinity primary response was elicited. Further increase in the number of thymic cells resulted in a progressive increase in the magnitude of the primary response but had no effect on affinity. Thus, a reduction of 50% in the magnitude of the response as a consequence of limiting the number of T-helper cells had no effect on the affinity of the PFC. The results are consistent with the interpretation that the effect of restriction in T-cell help on antibody affinity is not due to a direct effect on precursors of high-affinity PFC but is secondary to inefficient selection for high-affinity cells when the degree of cell proliferation is markedly reduced.     

Characterization of the antibody response against the type II collagen induced by anti-idiotypic antibody.

We reported that rabbit anti-idiotypic antibody (Ab2) against mAb, termed 1-5 (Ab1) and reactive with human type II collagen (CII) induced antibody response to CII in DBA/1J mice susceptible to collagen-induced arthritis. In the present study, we further characterized the anti-CII antibody response elicited by Ab2 with respect to epitope specificity, putative genetic background, and IgG subclass. Most of anti-CII antibodies (polyclonal Ab3) derived from Ab2-immunized mice were of the IgG1 subclass. We purified polyclonal Ab3, using a CII-coupled immunoadsorbent column and we developed monoclonal Ab3 from Ab2-immunized mice. Both purified polyclonal Ab3 and two monoclonal Ab3s specifically reacted with a selected epitope on CII, recognized by Ab1. The anti-CII antibody response stimulated by Ab2 was observed in DBA/1J (H-2q, Igh-1c) and DBA/2 (H-2q, Igh-1c) mice, but not in the BALB/c (H-2d, Igh-1a) and C57BL/6 (H-2b, Igh-1b) strains, thereby suggesting that the anti-CII antibody response elicited by Ab2 is controlled by the Igh gene.