Edmond A. Goidl

Production of auto-anti-idiotypic antibody during the normal immune response: IX. Characteristics of the auto-anti-idiotype antibody and its production

Using hapten-reversible inhibition of plaque formation as an assay for auto-anti-idiotype antibody (anti-Id) and as a means for following idiotype (Id) expression, we have obtained evidence that following immunization with trinitrophenyl (TNP) conjugates (a) there are differences in Id expression in the anti-TNP antibody response to different TNP conjugates although there is some overlap; (b) different strains, although showing some differences in Id expression, tend to produce cross-reactive Ids, thus no obvious allotype linked inheritance of Id expression is observed in this heterogeneous immune response; (c) the auto-anti-Id produced following immunization with TNP-Brucella abortus or TNP-Ficoll tends to be of the IgG2a and IgG2b isotypes.

Determination of antibody avidity at the cellular level by the plaque inhibition technique: effect of valence of the inhibitor.

Inhibition of plaque formation by multivalent and univalent ligands was compared as an assay of avidity of antibody produced by PFC. Multivalent ligands are much more effective as inhibitors and their use tends to impart an appearance of lack of heterogeneity and high avidity to the PFC populations being studied. It is thus probably generally advisable to employ univalent ligands in such studies.

Changes in affinity of 19 and 7S antibodies at the cellular level in responses to hapten conjugates of varying T dependency

Abstract The increase in affinity and heterogeneity of antibody with respect to time after immunization to the 2,4,6-trinitrophenyl (TNP) determinant was studied using TNP-brucella (BA) and TNP-type III pneumococcal polysaccharide (SIII). Experimental evidence is presented in support of the maturation of 19S antibody affinity. The difficulties which have been encountered in some previous investigations in detecting such a maturation appear to be the tendency of the cells to switch from IgM to IgG synthesis early after the peak of the primary response. Data are presented indicating that this switch occurs in a non-antibody-secreting memory cell population prior to, or more likely very shortly after, boosting. We also present evidence that the use of an antigen that does not induce a massive switch from IgM to IgG antibody synthesis offers a way of unmasking maturation of the 19S response. Thus, with the T-independent antigen TNP-SIII, a definite increase in heterogeneity could be detected in the 19S response upon secondary boosting. A greater increase in heterogeneity was noted in nude mice and was possibly due to the absence of suppressor T cells.

Production of auto-anti-idiotypic antibody during the normal immune response. XII. An enzyme-linked immunosorbent assay for auto-anti-idiotype antibody☆

An enzyme-linked immunosorbent assay (ELISA) to detect anti-idiotype (Id) antibody is described. Using this assay auto-anti-Id was detected in the serum of aged mice immunized with 2,4,6-trinitrophenylated-Ficoll (TNP-F). Hapten eluates from anti-TNP-F immune spleen cells also contained readily detectable auto-anti-Id.

Age-Associated Changes in Antibody-Forming Cells (B Cells)

Conclusion Despite considerable experimental effort, it is not possible to state unequivocally that with aging there occur some intrinsic changes in B cells. We have some indication that the number of pre-B cells may be affected in the bone marrow of the aged. If these changes extend to a decrease of the actual number of pluripotent stem cells, then this would indeed contribute to a lack of self-renewing capacity and affect the total peripheral B cell population. In those instances when actual precursor cell frequencies have been enumerated, even though for the majority of antigen systems studied B cell precursor frequency decreases in the aged, the responding B cells showed no individual functional decline (i.e., amount of antibody synthesized). Evidence has been obtained that changes in regulatory mechanisms are responsible, in part, for the decline in the immune response of the aged. This is particularly obvious when VH gene usage in antibody is compared between the total available repertoire (i.e., following B cell polyclonal activation) and that seen after antigen stimulation. The T cell compartment plays an important role in the expression of the antibody repertoire in both the young and the aged (77). Thymic involution, decrease in T cell function, and changes in T cell receptor repertoire may lead to altered T cell regulation as the animal senesces. We have presented evidence that in aging there is a decrease in the magnitude of antibody responses to both thymic-independent and TD antigens. There is also a decline in the affinity of antibody produced to TD antigen. An age-related change in antibody repertoire has been documented at both the serologic and molecular levels. Changes in regulatory mechanisms involving T cells and the immune network have also been described. Along with all of these changes, an intrinsic alteration in B cells may also occur in senescence. Both intrinsic and regulatory changes may contribute to the immune response as seen in the aged.

Studies of immune responses in mice prone to autoimmune disorders: I. Heterogeneity of the affinities of antihapten antibodies produced by NZB, NZW, and related strains of mice ☆

Mice of the NZB and NZW strains and their F1 hybrid produce antihapten plaque-forming cell (PFC) responses to T-dependent antigens (trinitrophenylated bovine gamma globulin and dansylated keyhole limpet hemocyanin) which are of unusually restricted heterogeneity of affinity, are relatively lacking in low-affinity PFC, and are of relatively high average affinity. Since some low-affinity PFC are present in NZB mice early after immunization, the results suggest a particularly marked down-regulation of low-affinity antibody production by these strains. The non-autoimmune-prone F1 hybrid (NZB X CBA) produces a typical heterogeneous response containing a high proportion of low-affinity PFC. Thus, the tendency to down-regulate low-affinity PFC is not inherited as a simple Mendelian dominant trait. The response of NZB mice to T-independent antigens does not show the same restricted heterogeneity of affinity. In fact, late after injections of trinitrophenylated Ficoll, NZB mice tend to have more heterogeneous responses than nonautoimmune-prone BALB/c mice in which a marked down-regulation of high-affinity antibody-producing PFC is seen. The possible relationship between these unusual features of the immune response of NZB and some related strains and their tendency to develop autoimmune disease is discussed.

Studies on the control of antibody synthesis. XV. Effect of nonspecific immunodepression on antibody affinity.

Abstract The effect on antibody affinity of marked nonspecific immunodepression early in the primary response was studied using three different immunosuppressive agents: 6-mercaptopurine, cyclophosphamide, and heterologous antilymphocyte antiserum. Marked nonspecific immunosuppression, even if limited to an early period in the primary response, resulted in a profound decrease in high-affinity plaque-forming cells (PFC) late in the primary response. Boosting led to the rapid appearance of high-affinity PFC, but the average affinity of the secondary PFC population was low in animals immunosuppressed early in the primary response. It would appear that selection for high-affinity B memory cells took place during the primary response despite the absence of detectable high-affinity PFC in the immunodepressed mice. Finally, the data suggest that in the absence of high-affinity antibody production, there may be an increased production of low-affinity antibodies, presumably as a consequence of a lack of antibody-mediated immunoregulation.

Studies of immune responses in mice prone to autoimmune disorders. II. Decreased down-regulation by auto-anti-idiotype antibody in autoimmune-prone mice

Three lines of evidence are presented which suggest that autoimmune-prone mice are deficient in the production of auto-anti-idiotype antibody during their immune response to trinitrophenylated Ficoll (TNP-F). NZB, MRL lpr/lpr and older BXSB male mice have no hapten-augmentable plaque-forming cells (PFC). Hapten-augmentable PFC have been previously shown to be cells whose secretion of antibody has been inhibited by the binding of auto-anti-idiotype antibody to cell surface idiotype. Sera from TNP-F immunized NZB mice lack PFC inhibiting activity (anti-idiotype antibody). Spleen cells from TNP-F immune NZB mice fail to transfer anti-idiotype antibody-mediated suppression to naive mice as do spleen cells from immune non-autoimmune-prone mice. Taken together these data suggest that autoimmune-prone mice are deficient in auto-anti-idiotype antibody-mediated downward regulation of their immune responses. It was further shown that the immune response of NZB mice to TNP-F shows a slower decline in splenic PFC and a greater heterogeneity of PFC affinity than do the responses of non-autoimmune-prone strains. Since athymic (nude) mice, which were previously shown to be defective in the production of auto-anti-idiotype antibody, also show a slower decline in splenic PFC and an increased heterogeneity of PFC affinity, it is suggested that these peculiarities of the immune responses of autoimmune-prone and athymic mice are also the consequences of the lack of auto-anti-idiotype antibody-mediated down-regulation.

Production of auto-anti-idiotypic antibody during the normal immune response. VIII. Effect of auto-anti-idiotypic antibody on contact sensitivity.

An eluate prepared by brief incubation of spleen cells from 2,4,6-trinitrophenyl (TNP) lysyl-Ficoll immunized mice with TNP-epsilon-amino-n-caproic acid causes a specific inhibition of the induction of contact sensitivity by 2,4,6-trinitrochlorobenzene skin painting. The active factor in the eluate binds to an anti-mouse immunoglobulin (Ig) immunoadsorbent column but not to a TNP immunoadsorbent column and is therefore Ig but not anti-TNP antibody. The active factor does bind to an immunoadsorbent prepared from anti-TNP antibody, suggesting that the factor has anti-idiotype specificity. Evidence based upon hapten-reversible inhibition of plaque formation and an enzyme-linked immunosorbent assay (ELISA) indicates that the eluates contain auto-anti-idiotype antibody specific for anti-TNP antibody. It is suggested that auto-anti-idiotype antibody spontaneously produced during the immune response to a T-independent antigen can specifically downregulate contact sensitization to the same epitope.

AUTOANTI‐IDIOTYPE ANTIBODY PRODUCTION FOLLOWING ANTIGEN INJECTION AND IMMUNE REGULATION *

During the course of studies on antibody affinity at the plaque-forming cell (PFC) level, it was observed that, at times, more PFC were detected in the presence of low concentrations of hapten than in the absence of hapten.’ This observation was clearly contrary to the usual view of this system, according to which adding hapten to the PFC assay medium should inhibit plaque formation or have no effect, depending upon the affinity of the antibody secreted and the concentration of hapten. We hypothesized that the increase in the number of PFC might be due to the “displacement” by hapten of autoanti-idiotype antibody (anti-id) from cell surface idiotype. According to this hypothesis, the binding of autoanti-id on the cell surface inhibits the secretion of antibody. This inhibition is reversible and, when the anti-id dissociates, secretion of antibody is reinitiated. Hapten and anti-id, in effect, compete for the same cell surface idiotype. Therefore, once dissociated in the presence of hapten, anti-id cannot cause further inhibition of secretion. Under certain experimental conditions, the number of hapten-augmentable PFC is very great.l For example, when normal nonirradiated AKR mice received spleen cells from syngeneic mice that had received 10 pg 2,4,6-trinitrophenylated ficoll (TNP-F) seven days before their use as cell donors, the number of PFC observed in the presence of hapten was two or three times that seen in the absence of hapten. We took advantage of this situation to test the hypothesis that hapten-augmentable PFC are cells whose secretion of antibody had been inhibited by the binding of autoanti-id to cell surface antibody molecules. If the hypothesis were true, we would expect that incubation with hapten would elute material with properties of anti-id from immune cell populations