Jerne (1) has proposed that the immune system functions as an interacting network of inducible elements that normally exist in a steady state. In this network, the immune response is regulated by idiotype-anti-idiotype interactions. Antigen administration stimulates some elements of the network, and the subsequent alterations to restore the perturbed steady-state condition represent the cellular and molecular reactions that occur during the immune response.In many experimental systems that examine idiotype-anti-idiotype interactions, the anti-idiotypic antibodies have been induced in species other than the one in which the idiotype was produced (2-6). Induction of isologous anti-idiotypic antibodies has also been achieved (4-12). Spontaneous, autologous anti-idiotypic-antibody and (or) anti-receptor-antibody production during the course of an immune response has been described for haptens (13-16), sheep erythrocytes (SRBC) 1 (i 5, 16), and alloantigens (16)(17)(18)(19). In addition, the decrease in binding affinity of anti-tobacco mosaic virus antibodies has been attributed to the appearance, during the course of the immune response, of lymphocytes bearing auto-anti-idiotypic receptors (20).Although cyclic changes in serum-antibody levels and affinity have been observed during the immune response to many antigens, they have generally been attributed to variations in the degree of masking, by serum antibody, of antigenic determinants on persisting antigen (21,22). Nevertheless, a rapid decrease in affinity of individual plaque-forming cells (PFC) (23) or serum antibody (20, 24) cannot be convincingly explained in this manner. Urbain (25) has suggested that an auto-anti-idiotypic

Hapten-augmentable plaque-forming cells PFC) are cells whose secretion of antibody is specifically inibited by surface-bound auto-anti-iodotype antibody th can be displaced by hapten. This study showed that the percentage of hapten-augmentable PFC present in mice during the primary response to trinitrohenat FicolI(NP-F) increases with age.The data suggest gat there is a relative increase in the autoanti-idiotypic antibody response with age and therefore a greater down-regulation of antibody production. The effect of age on idiotype expression was also studied. Hapten-reversible inhibition of plaque formation was used as an assay for antiidiotype antibody and idiotype bearing antibody-secreting cells.Sera from aged (21-to 22onth-otd)C57BL/6 mice immunized with TNP-F significantly inhibited plaque formation, in a hapten-reversible manner, by spleen cells from 81% of TNP-F-immunized aged mice. However, these sera inhibited plaque formation by cells from only 50% of similarly immunized young adult (6-to 8-week-old) mice and 20% of immature (3-to 4-week-old) syngeneic mice. Similarly, sera from TNP-F-immunized young adult or immature mice inhibited formation of plaques by cells from immunized donors of the same age as the mice from whom the serum was obtained, but only rarely inhibited plaque formation by cells from mice of other age groups. The data thus suggest that the repertoire of TNP-specific idiotypes that are produced in response to TNP-F varies with age in syngeneic mice.We have shown that, in the presence of low concentrations of hapten, there frequently is an increase in the number of plaque-forming cells (PFC) detected (1, 2) in a Jerne PFC assay (3). These hapten-augmentable PFC represent potential antibody-producing cells whose secretion appears to have been inhibited by the binding of auto-anti-idiotype antibody to cell-surface antibody molecules. Hapten competes with antiidiotype antibody for available cell-surface antibody molecules and, in this manner, displaces bound anti-idiotype antibody and reverses its inhibitory effect on antibody secretion. We have also shown that serum anti-idiotype antibody can be assayed by its ability to cause a hapten-reversible inhibition of plaque formation (2). By using these assays, we have found that the immune response to both thymic-dependent and thymic-independent antigens is down-regulated by the production of auto-anti-idiotype antibodies (unpublished results). From these studies, we have concluded that auto-anti-idiotype antibody serves as a normal mechanism for immune regulation in a manner compatible with predictions based on Jerne's network theory (4). Several other groups have also shown that autoanti-idiotype antibodies are produced during a normal immune response (5-7).The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.6788

In the accompanying paper (1), we have shown that after day 4 of the immune response of AKR/J mice to 2,4,6-trinitrophenyl-lys-Ficoll (TNP-F) 1, the addition of free hapten to a plaque-forming-cell (PFC) assay increased the number of observed splenic anti-trinitrophenol (TNP) PFC. Immune spleen cells, taken 7 d after immunization, transferred this property of the immune response to normal recipients; spleen cells from such recipients, assayed 4 d after cell transfer and TNP-F injection, manifested an exaggerated form of this phenomenon. It was hypothesized that the increase in PFC was the result of the displacement, by hapten, of auto-anti-idiotypic antibodies that were synthesized during the course of the normal immune response. If this hypothesis were correct, it should be possible to obtain auto-anti-idiotypic antibodies by hapten elution from appropriate immune spleen cells. Results in the accompanying paper (1) also suggested that the putative auto-anti-idiotypic-antibody response was involved in the downward regulation of the immune response of AKR/ J mice to TNP-F. One might, therefore, expect to find auto-anti-idiotypic antibody in the serum of AKR/J mice immediately after the abrupt decrease in the number of detectable splenic PFC: i.e., 7 d after antigen injection.In the present paper, evidence is presented to support these hypotheses in that hapten-reversible inhibition of PFC in vitro can be demonstrated with hapten eluates from immune cells and with immune serum. The factor responsible for inhibition has immunoglobulin-like determinants, lacks anti-TNP-antibody activity, and is absorbable by an AKR/J anti-TNP-antibody immunoadsorbent. Hapten-reversible inhibition of PFC represents a simple in vitro assay for anti-idiotypic antibody. In this

It has generally been felt that tolerance can be induced more readily in neonatal than in adult animals. While this is clearly true in allograft tolerance induction (1), it has been less obvious with other antigens. Thus, Siskind et al.(2) and Howard and Hale (3) found no difference between neonatal and adult mice in regard to tolerance induction with polysaccharide antigens and Dresser (4) found that the dose of aggregate-free bovine gamma globulin (BGG) 1 required to induce tolerance in mice was the same in neonatal and adult animals. It has been unclear as to whether immature lymphocytes are or are not more sensitive to tolerance induction than are mature lymphoid cells. Recently, in vitro studies (5-7) have shown that tolerance could be induced in vitro in B lymphocytes from immature donors at antigen concentrations far lower than that required for tolerance induction in mature B lymphocytes.In the studies reported here a cell transfer system was employed so that the susceptibility of B cells to tolerance induction could be assayed in the absence of T cells and in a constant, adult, in vivo environment. It was found that with two hapten-carrier conjugates neonatal cells were indeed more susceptible to tolerance induction, both in vivo and in vitro, than were adult B cells. However, with BGG as antigen no difference was detected between B cells from 17-day fetal mice, neonatal mice, and adult mice with regard to the ease of tolerance induction either in vivo or in vitro. The results thus suggest that the relative ease of tolerance induction in immature B cells may be limited to moderately polyvalent antigens such as hapten-carrier conjugates.

Immune responses to trinitrophenylated hemocyanin (TNP-KLH), Ficoll (TNP-Ficoll), and Brucella abortus (TNP-BA) were examined in BALB/c mice bearing subcutaneous transplants of TEPC-1017 and TEPC-1033, the two known IgD-secreting BALB/c plasmacytomas. Both primary and secondary 19S and 7S splenic plaque-forming cell (PFC) responses in spleen to intravenously injected TNP-KLH were enhanced three to fivefold. Primary responses to TNP-Ficoll were 1.5-2 times higher than in control mice (particularly the 7S PFC response). Primary responses to TNP-BA were enhanced by TEPC-1017 but suppressed by TEPC-1033, while secondary responses to TNP-BA were enhanced three to sevenfold by both tumors. Intraperitoneal injections of ascites fluid from mice bearing TEPC-1017 or TEPC-1033, or of IgD isolated from such ascites fluid, caused a similar enhancement of the primary response to TNP-KLH, as did the tumor itself, particularly when injected approximately 1 wk before antigen injection. IgD-containing ascites fluid had no effect on the response of athymic (nu/nu) BALB/c mice to TNP-KLH. These findings suggest the existence of an IgD-responsive immunoregulatory T cell.