Monoclonal anti-Id antibodies react with varying proportions of human B lineage cells

Monoclonal antibodies to idiotypic determinants are being used with increasing frequency for analysis and treatment of B cell malignancies. In the present study we have compared the idiotypic specificities of a panel of 39 mouse monoclonal anti-idiotype (anti-Id) antibodies developed against 16 monoclonal human immunoglobulins (Ig). The Id cross-reactivities of these antibodies with Ig products of normal and abnormal B cells were examined by immunofluorescence and immunochemical methods. The reactivity patterns of these anti-Id antibodies with a normal population of plasma cells were highly variable in the immunofluorescence assay. Six were reactive with 2 to 10% of normal plasma cells, 30 with 0.1 to 2% of plasma cells, and three with less than 0.1% of plasma cells from blood, bone marrow, spleen, or tonsils. These reactivity patterns were relatively consistent among samples from 23 Caucasian, black, and Oriental adults. Although the reactivities of most anti-Id antibodies in the panel were not restricted to a particular Ig isotype, several were preferentially reactive with a particular heavy or light chain isotype: one IgM-, two IgA-, two kappa-, and three lambda-restricted antibodies. The immunofluorescence data was confirmed by biosynthetic analysis of Id+ molecules produced by a normal plasma cell population. When the reactivity of this panel of anti-Id antibodies with nonhomologous B cell neoplasms was examined, seven of 30 myelomas or leukemia-derived products and one of nine B cell leukemias or lymphomas without paraproteins were found to be cross-reactive with one or two of the anti-Id antibodies. Although clearly significant, the cross-reactivity between the Id of these paraproteins appeared to be of lower affinity than the reactivity of the homologous Id with their respective anti-Id antibodies. The results reveal a remarkable diversity in the specificities of monoclonal antibodies classified by conventional criteria as anti-Id antibodies, and indicate the potential usefulness of a panel of antibodies for analyzing clonal diversity in normal and abnormal B cell development.     

Strategies for production of monoclonal anti-idiotype antibodies against human B cell lymphomas

Murine monoclonal antibodies (MAB) against the idiotype (Id) of B lymphocyte malignancies are powerful reagents for the study of these diseases, and are potentially useful for treatment. Different strategies for the production of these anti-Id MAB have been compared. Initially, the Id Ig from nonsecreting B cell tumors was "rescued" by human X mouse or human X human hybridization. These somatic cell hybridizations resulted in the secretion of human Ig in 10 and 100% of the fusions, respectively. In a second step, anti-Id MAB were produced by using the "rescued" Id Ig as immunogen. A more streamlined approach is based on a one-step procedure in which the tumor cell suspension is used as immunogen. This method of immunization, coupled with a four-layer ELISA, results in the detection of anti-Id MAB in a frequency of approximately 1% of the total hybrids. By using a pool of 10 different anti-Id MAB, each reactive with the tumor of one patient, we searched for idiotypic relatedness among a panel of 50 additional tumors. No cross-reactions were found, indicating that our current strategy results in the identification of unique idiotypic determinants among human B cell tumors. Idiotypic Ig can be found in the serum of patients with B cell tumors. Among groups of patients, there is a wide spectrum of serum Id levels, ranging from less than 0.01 microgram/ml to greater than 500 micrograms/ml.     

Idiotypic determinants on human T cells and modulation of human T cell responses by anti-idiotypic antibodies

The role of idiotypic anti-idiotypic interactions in the regulation of the human T cell response to tetanus toxoid (TT) antigen was examined in three subjects. Rabbit anti-idiotypic (anti-Id) antisera were raised against IgG (Fab')2 anti-TT obtained 7 to 10 days after booster immunization with TT. F(ab')2 fragments of rabbit-anti-Id IgG were used in conjunction with fluorescein-conjugated goat anti-rabbit Ig in an indirect immunofluorescence assay to determine the frequency of Id-positive cells in T cell-enriched preparations. This frequency was 24, 29, and 38 per 10,000, respectively, in the three subjects studied. Significant contribution of contaminating B cell to fluorescence-staining was ruled out by capping experiments using goat anti-human Ig (GAHIG) and by double staining experiments using rhodamine-conjugated GAHIG. Absorption of anti-Id antisera with Epstein Barr virus (EBV)-transformed B cell lines from the IgG (Fab')2 anti-TT donor, but not with EBV-B cell lines from unrelated donors, removed their reactivity with the T cells. Rabbit anti-Id IgG caused minimal proliferation (two-threefold) of T cells and had no effect on T cell proliferation in response to TT antigen when added to the cultures. Preincubation of T cells for 48 hr with rabbit anti-Id IgG (Fab')2, but not with preimmune rabbit IgG (Fab')2, resulted in the generation of antigen-specific suppressor cells that inhibited T cell proliferation in response to TT, but not in response to diphtheria toxoid (DT). These cells also inhibited the synthesis of IgG anti-TT in response to in vitro stimulation with TT antigen, but not the synthesis of IgG anti-DT in response to DT antigen. Adsorption of T cells over plates coated with rabbit anti-Id IgG (Fab')2 enhanced the proliferative response of the T cells to TT, but not to DT antigen, and enhanced the helper activity of the T cells for the in vitro synthesis of IgG anti-TT but not of IgG anti-DT antibodies. These results suggest that idiotypic-anti-idiotypic interactions play a role in the human T cell response to antigen.     

Detection and titration of antitetanus antibodies using an automated passive hemagglutination method]

An automated reversed passive haemagglutination technique is described, using human erythrocytes sensitized by the chromic chloride method with higly purified tetanus toxoid. Such erythrocytes are agglutinated by the specific antibodies in a Technicon autoanalyzer. Clots are decanted, supernatant is hemolyzed and optical density is measured at 550 millimicron. Assay standards for comparison, ranging from 5 to 25 UI/ml, are prepared from human antitetanus immunoglobulins titrated by toxin neutralisation assay in mice. This method allows to screen donors' sera for presence of high titers of tetanus antibodies (larger than or equal to 5 UI) suitable for preparation of antitetanus immunoglobulins. 4,4% of donors have circulating antitetanus antibody levels corresponding to 5 UI/ml or more, by this method. The specificity of antibodies has been confirmed by the neutralisation assay in mice. The results obtained among 1.000 donors well agree with those of the counter-immuno-electrophoresis technique (C.E.P.). But, when, compared with C.E.P., the haemagglutination assay appears more objective, more quantitative and sensitive, and allows to get not only a rapid screening test but also a precise titration simultaneously.     

Idiotypic analysis of antibodies to hen egg-white lysozyme (HEL). III. Further studies on the idiotypic cross-reactivity among anti-HEL antibodies in mice

An isolated antibody preparation directed to the native hen egg-white lysozyme (HEL) from a single A/J mouse (#a-11), termed IDHELa-11, was inoculated into rabbits to produce anti-idiotypic sera (anti-Id). The antisera were extensively absorbed with normal A/J Ig to render then idiotype specific. Radioimmunoassay utilizing 125I-IdHELa-11 and the anti-ID sera (R103 and R104) was performed to examine the idiotypic cross-reactivity of the humoral immune response to HEL in various mouse strains and other animal species. Idiotypes shared by IdHELa-11 were detected in the sera of five mouse strains tested, but not in any of the examined sera of other animal species such as rats, goats, guinea pigs, or sheep, indicating the occurrence of species-specific cross-reactive idiotypes (CRI) of antibodies to HEL. Our experiments also suggested the presence of intrastrain CRI. The present experiments, along with our earlier results (15), suggest that idiotypic cross-reactivity among murine antibodies to HEL appears to be weak. Thus when R103 and R104 were the anti-Id sera used, the frequency of occurrence of CRI shared by IdHELa-11 in 5 micrograms of anti-HEL antibody in strain A mice was 74 ng and 111 ng; in other strains it was 25-44 ng and 60-98 ng, respectively.     

Detection of in ovo derived idiotypic antibodies. I. A model for maternal-neonatal idiotype network studies.

Idiotypic vaccines appear to have many advantages over conventional vaccines. Maternal Id or anti-Id that are passively transferred to the fetus or neonate could provide another avenue for vaccination. Based on this premise we have investigated the transmission of idiotypic antibodies from dam to embryo by inoculating laying hens and analyzing their egg yolks for the presence of Id, anti-Id, and anti-anti-Id. The Ag chosen for these studies was BSA. After isolation and concentration BSA Id titers were approximately 256,000. The presence of anti-Id antibody in yolk samples is characterized by the ability of anti-Id to inhibit BSA binding to Id. The anti-Id extracted from yolks inhibited BSA binding to Id by 7 to 53%. Not all of the anti-Id present in samples was binding to a paratope-associated Id because 11 to 16% of the fluoresceinated anti-Id bound to Id-coated beads in the presence of excess BSA. Because a portion of the anti-Id antibodies were internal images of the BSA Id, they should be able to mimic Ag. This idea was confirmed when anti-Id inoculated in hens caused the synthesis of antibodies that would bind BSA and could be detected in an ELISA. Binding of anti-anti-Id to BSA-coated wells could be inhibited by preincubation of anti-anti-Id with BSA in solution. The chicken model provides a novel system to investigate maternal-fetal and maternal-neonatal interactions in the idiotypic network and the cellular mechanisms involved in the ontogeny of the Id network in neonates.     

Anti-idiotype antibodies prevent antibody binding to mouse sperm and antibody-mediated inhibition of fertilization

Antibodies to components of sperm can interfere with sperm function and prevent fertilization by blocking specific steps of gamete interaction. It can be proposed that anti-idiotype antibodies (anti-Ids) that recognize determinants located close to or within the antigen-binding site of an anti-sperm antibody could block antibody binding to sperm antigen and antibody-mediated inhibition of fertilization. To test this hypothesis, rabbit polyclonal antibodies to idiotypic determinants of the monoclonal anti-sperm antibody M42.15 were developed and characterized. Previous studies demonstrated that M42.15 monoclonal antibody (mAb) inhibits fertilization in vitro and in vivo by inhibiting sperm-zona pellucida interaction. Anti-idiotype antibodies to M42.15 mAb (anti-Id M42) were isolated by affinity chromatography on immobilized M42.15 mAb. Binding specificity of anti-Id M42.15 was demonstrated in a solid-phase radioimmune binding assay and by specific immunoprecipitation of soluble M42.15 mAb. Anti-Id M42 competitively inhibited M42.15 mAb, but not P220.2 mAb, binding to mouse sperm, confirming that the anti-Id preparation contained antibodies directed against idiotopes within or adjacent to the antigen-binding site of the mAb. At equimolar concentrations, anti-Id M42 inhibited binding of 125I-labeled M42.15 mAb to sperm by greater than 80%. These results showed that anti-Id M42 efficiently blocked antibody binding to sperm and suggested that anti-Id M42 could be used to neutralize the anti-fertility activity of the M42.15 mAb. When tested in in vitro fertilization assays, anti-Id M42, but not rabbit immunoglobulin, prevented M42.15 mAb-induced inhibition of fertilization. Together, these results show that the inhibitory activity of anti-sperm antibodies capable of interfering with gamete interaction can be neutralized by anti-Id that recognize determinants close to the antigen-combining site of the anti-sperm antibody.     

Lack of pokeweed mitogen-induced IgE formation in vitro by human peripheral blood mononuclear cells: detection of cross-reacting idiotypic determinants on polyclonal Ig by antibodies to a single IgE myeloma protein

The ability of human peripheral blood mononuclear cells to synthesize IgE in vitro in response to pokeweed mitogen (PWM) is controversial. To determine whether the conflicting results obtained by different laboratories could be due to inherent qualitative differences in the anti-IgE antibodies used to measure low concentrations of IgE in culture supernatants, we compared the specificities of anti-IgE reagents prepared by various methods. Immunoadsorbent-purified antibodies were isolated from a goat antiserum to the lambda, IgE myeloma protein PS and a rabbit antiserum to the kappa, IgE protein Bed in three ways: 1) antibodies to IgE PS (anti-PS) were isolated from the goat antiserum by affinity chromatography with PS coupled to Sepharose 4B; these antibodies consisted of anti-epsilon chain-specific and anti-idiotypic antibodies to protein PS; 2) antibodies specific for the epsilon-chain (anti-epsilon) were purified by affinity chromatography with IgE myeloma proteins that were not used for immunization; and 3) antibodies to idiotypic determinants of proteins PS (anti-id PS) and Bed (anti-id Bed) were isolated on affinity columns with the respective myeloma proteins after absorption of the epsilon-chain-specific antibodies. These three types of antibodies were then used in a solid phase radioimmunosorbent test to quantitate the amount of "IgE" synthesized by peripheral blood mononuclear cells from nonatopic and atopic donors cultured for 7 days in the presence and absence of PWM. ANTI-PS antibodies detected a PWM-induced "IgE formation" in cell culture supernatants of both non-atopic and atopic donors. (ABSTRACT TRUNCATED AT 250 WORDS)     

Idiotypic variation in a human B lymphoma cell line

The Ig Id of a B cell lymphoma serves as a distinct marker of the malignant clone and thus as a tumor-specific target for antibody therapy. Somatic variation of the Ig genes expressed by B cell tumors can lead to loss of reactivity with anti-Id antibodies and escape of tumors from the therapeutic effects of such antibodies. In our study, we have used anti-Id antibodies to screen for variants within a cell line derived from a patient with a large cell lymphoma of the B cell type. Cells were simultaneously stained on their surface for idiotypic and for isotypic Ig determinants using reagents labeled with different fluorochromes. Tumor cells expressing intact Ig molecules with alteration of their idiotypic determinants were isolated with the fluorescence activated cell sorter. Idiotypic variation was an ongoing process in vitro with Id- variants being generated at a rate of 2.7 x 10(-4)/cell per generation and Ig- cells being produced at a rate of 1.31 x 10(-5)/cell per generation. Subcloned variants expressed subtle differences in reactivity with a panel of three non-cross-blocking anti-Id antibodies. Analysis of Ig gene rearrangements by the Southern blotting technique using a JH probe established that the variants and the original tumor cells were all clonally related. Immunoprecipitation of surface labeled Ig molecules from the variant subclones disclosed major alterations of the lambda-L chains with no gross alterations of the mu-H chains. Related studies have established that the tumor cells undergo rearrangement and expression of new lambda-L chain genes.     

Induction of allergen-specific IgE and IgG responses by anti-idiotypic antibodies

In order to explore idiotypic, anti-idiotypic, and anti-anti-idiotypic responses to allergens, BALB/c mice were immunized with affinity-purified human idiotypic antibodies directed against a highly purified shrimp allergen. This resulted in the production of anti-idiotypic antibodies which were quantitated by using rabbit idiotypic antibodies raised against the same purified allergen. The mouse anti-idiotypic antibodies recognized shrimp-specific human idiotypic antibodies of the IgE isotype from 18 of 20 individuals, and IgG antibodies from 14 of 20 shrimp-sensitive patients. Immunization of BALB/c mice with affinity-purified, allergen-specific anti-idiotypic antibodies induced anti-allergen IgE and IgG responses in the absence of the allergen. This paper thus presents evidence that anti-idiotypic antibodies raised against allergen-specific idiotypic antibodies may substitute for the original allergen in the induction of allergen-specific idiotypic antibodies. The demonstration of shared idiotopes on IgG and IgE antibodies in the sera of shrimp-sensitive patients supports the use of allergen-specific anti-idiotypic antibodies as surrogate allergens.     

Functional, structural, and antigenic similarities of guinea pig anti-phosphorylcholine antibodies.

The humoral immune response to PC was measured in guinea pigs. PC-vaccine stimulated IgM and IgG2, but little IgG1, anti-PC -antibodies. No memory was induced and immunization in CFA produced tolerance. PC-KLH, on the other hand, stimulated IgM, IgG2, and IgG1 anti-PC antibodies with carrier-specific memory. Hapten inhibition of plaque formation showed uniform binding patterns with minor, but significant, differences between antiPC-vaccine and anti-PC-KLH antibodies. The antibodies were characterized by IEF and idiotypic analyses. Early after immunization with PC-vaccine, guinea pigs had restricted IEF patterns which in inbred, but not outbred animals were indistinguishable between individuals. These patterns remained restricted but more individualized with time after immunization. Anti-PC-KLH antibodies showed more heterogeneity and individuality. However, these structurally heterogeneous antibodies reacted equivalently with rabbit anti-idiotype antisera and therefore must share common structural features, regardless of isotype or the genetic background of the guinea pig.     

Monoclonal anti-idiotopic antibodies against myasthenia-inducing anti-acetylcholine receptor monoclonal antibodies. Preponderance of nonparatope-directed antibodies affecting antigen binding

To analyze components of the idiotypic network in experimental autoimmune disease, we produced 17 isogeneic anti-idiotopic monoclonal antibodies (anti-Id) against two experimental autoimmune myasthenia gravis-producing anti-acetylcholine receptor (anti-AChR) monoclonal antibodies. We studied the binding of five of the anti-Id to the anti-AChR monoclonal antibodies bearing the complementary idiotopes (Id-mAb). They bound with Kd values ranging from 0.06 to 0.86 nM, values comparable to those of Id-mAb:AChR complexes (0.26 and 0.34 nM). All of the anti-Id tested moderately inhibited the binding of AChR to Id-mAb, whereas for each anti-Id, AChR either strongly inhibited anti-Id binding or had no effect on anti-Id binding. Hence, the inhibition of Id-mAb:AChR binding by anti-Id was not reciprocal with the inhibition of anti-Id:Id-mAb binding by AChR. For each anti-Id, the relative affinities of anti-Id and AChR for Id-mAb together with the lack of symmetry of inhibition by anti-Id compared to inhibition by AChR indicate that these two "ligands" are not competitive inhibitors. Consequently, anti-Id and AChR do not bind to overlapping sites on the Id-mAb, suggesting that the observed inhibition is mediated allosterically. This may be a common mechanism of anti-Id:Id binding, which would have important implications for the mechanism of anti-Id-induced suppression.     

Sodium periodate-induced suppressor T cells of the human in vitro antibody response

Combinations of T and B lymphocytes from normal individuals booster immunized 14–30 days previously with a combination of diphtheria and tetanus toxoids, synthesized IgG antitetanus toxoid, and IgG antidiphtheria toxoid antibodies when stimulated by pokeweed mitogen in vitro. The addition of 5 μg of soluble tetanus toxoid to the cultures during the first 2 days incubation resulted in greater than 90% suppression of the subsequent production of IgG antitetanus toxoid antibodies. The synthesis of IgM antitetanus toxoid antibodies, total IgG, total IgM, and IgG antidiphtheria toxoid antibodies were unaffected. Similarly, the addition of 5 μg of soluble diphtheria toxoid suppressed the synthesis of IgG antidiphtheria toxoid antibodies with no effect on the synthesis of IgG antitetanus antibodies. Allogeneic combinations of B and T lymphocytes were capable of mediating the suppression, and irradiation of the T cells caused only a partial and variable reversal of the suppression. The antigen-induced specific suppression of antibody synthesis could not be demonstrated in cultures stimulated with soluble T-cell-derived helper factors.     

Structural characterization of a cross-reactive idiotype shared by monoclonal antibodies specific for the human CD4 molecule.

A panel of mouse monoclonal anti-CD4 antibodies was characterized in terms of idiotypic expression by using specific anti-idiotypic antibody (anti-Id) reagents generated in rabbits immunized with anti-Leu3a, a monoclonal anti-CD4 which inhibits the human immunodeficiency virus (HIV) gp120 binding to CD4. Direct binding and competitive inhibition assays demonstrate that the majority of monoclonal anti-CD4 antibodies able to recognize CD4 epitopes overlapping the epitope recognized by anti-Leu3a expressed an antigen-combining site-related cross-reactive idiotype (IdX). Western blot analysis was used to demonstrate that this IdX is associated primarily with the light (L) chain of the monoclonal anti-CD4 antibodies. To further characterize the structural basis of the IdX, the nucleotide sequence of the variable region of the L kappa chain of anti-Leu3a was determined. Peptides corresponding to the first, second, and third complementarity determining regions (CDRs) of the L chain of anti-Leu3a were synthesized and used to immunize rabbits. All anti-peptide antisera recognized the immunizing peptide, the cognate anti-Leu3a molecule, and several other monoclonal anti-CD4 antibodies by direct binding assays. Western blot analysis utilizing the anti-CDR peptide reagents demonstrates that the reactivity to the monoclonal anti-CD4 antibodies was L chain-specific. The anti-Id generated by immunizing with the intact anti-Leu3a molecule failed to recognize the three L chain-derived CDR synthetic peptides, suggesting that the IdX requires the presence of the three-dimensional configuration of the L chain for its expression. The broad range of reactivity exhibited by the antipeptide antisera indicates that the majority of mouse monoclonal anti-CD4 antibodies characterized in this study utilize L chains encoded by a single germ line variable (V) region kappa (V kappa) chain gene or by V kappa genes that belong to the same gene family.     

Expression and synthesis of cross-reactive idiotypic antibodies by peripheral blood lymphocytes and their suppression by anti-idiotypes in patients with IgA nephropathy.

We have recently described that patients with IgA nephropathy present high serum levels of anti-BSA idiotypic antibodies that were well correlated with the existence of hematuria. Furthermore, these Id were found in circulating and renal deposited immune complexes. In the present work, we examined the expression of surface idiotypic determinants on PBL by flow cytometry and their in vitro production, using as reagent anti-idiotypic antibodies previously well characterized. The presence of cross-reactive Id-bearing cells was observed in 5 out of 6 patients studied, with frequencies ranging from 3 to 12% of lymphocytes. After 7 days of culture, the spontaneous synthesis of idiotypic antibodies by PBL was found elevated in 6 out of 13 (46%) patients. A major Id cell expression and production was noted in patients with active disease as defined by hematuria. The preincubation of PBL with 20 and 50 micrograms of anti-idiotypic antibodies/2 x 10(6) cells for 3 days induced a significant inhibition of cross-reactive Id production in a dose-dependent fashion, with a degree of suppression between 12 and 50% in five out of six patients studied. In the above assays, as negative controls, we used the anti-Id antibodies previously adsorbed on an Id-Sepharose column. On the whole, these results suggest that patients with IgA nephropathy present dysfunctions in the Id-Anti-Id network that could play an important role in the pathogenesis of this disease.     

Interspecies cross-reactivity of monoclonal antibodies to various epitopes of human plasminogen

The immunological cross-reactivities of three conformationally specific monoclonal antibodies to distinct epitopes on human plasminogen toward plasminogens purified from 14 additional species have been examined. Antibody 10-F-1, which is produced against an epitope on the kringle 4 region of human plasminogen, shows a high degree (greater than 80%) of cross-reactivity against baboon, goat, monkey, ovine, and rabbit plasminogens; more limited (20-50%) cross-reactivity against bovine, equine, goose, guinea pig, mouse, rat, and porcine plasminogens; and little comparable cross-reactivity against canine and chicken plasminogens. Antibody 10-H-2, generated to an epitope of the kringles 1-3 region of human plasminogen, shows extensive cross-reactivity (72%) only toward monkey plasminogen, more limited (22-35%) cross-reactivity toward equine and rabbit plasminogens, and much less cross-reactivity toward any other of the above plasminogens. Antibody 10-V-1, also produced against an epitope on the kringle 1-3 region of human plasminogen, which is distinct from the 10-H-2 epitope, shows extensive cross-reactivity (72-100%) with baboon, monkey, and rabbit plasminogens; more limited cross-reactivity with equine (48%) and mouse (28%) plasminogens; and a low level of such reactivity with the remaining plasminogens. These studies show that the extent of interspecies cross-reactivity of various plasminogens greatly depends upon the epitope in question. The K4 region of these molecules appears more extensively conserved than the K1-3 region, at least in regard to the particular epitopes examined in this study.     

Selection of antibody repertoires by anti-idiotypes can occur at multiple steps of B cell differentiation

These experiments were designed to evaluate whether alterations in Id expression after anti-Id treatments result from direct modulation of Id-producing B cells, and whether idiotypic selection operates in bone marrow or spleen B cells. By using the NPb Id model, we have studied the functional behavior of isolated LPS-reactive B cells transferred from B6 mice into histocompatible LPS-NR B10.Cr hosts and primed with LPS conjugates of anti-Id antibodies. We have found that previous anti-idiotypic manipulation of host mice by neonatal administration of suppressive doses of Ac 38 antibodies, or adult injection of enhancing doses of Ac 146 antibodies, modulated the T cell-independent Id response of either immature bone marrow or mature splenic responding cells, transferred from normal, untreated donors. These results are interpreted to suggest that selection of antibody repertoires by anti-Id may occur at multiple steps of B cell differentiation.     

Anti-immunoglobulin antibodies. II. Expression of individual and cross-reactive idiotypes on syngeneic and homologous anti-idiotype antibodies

Syngeneic anti-(anti-Id) antibodies were prepared against BALB/c anti-A48Id antibodies, BALB/c anti-460Id monoclonal antibodies, and A/J anti-J558 IdI monoclonal antibodies. With these anti-(anti-Id) antibodies we identified cross-reactive idiotypes on syngeneic and homologous anti-A48Id and anti-460Id antibodies. By contrast, tbe idiotypic determinants of A/J anti-J558 IdI monoclonal antibodies were not shared by other syngeneic, homologous, or xenogenic anti-J558 IdI or IdX antibodies. These results suggest that idiotype-antiidiotype reactions that serve as regulatory controls within the immune system are characteristic for each particular antigen system, strain, or species and that such interactions make the system self-limited with respect to the whole antild repertoire.     

Mouse monoclonal antibodies to chicken VH idiotypic determinants reactivity with B and T cells

We have prepared mouse monoclonal antibodies against idiotypic (Id) determinants on chicken antibodies to N-acetylglucosamine (NAGA) and p-aminobenzoic acid (PABA) made by inbred line EL 6(3) birds. The monoclonal anti-NAGA Id antibody, termed CId-1, reacted with affinity purified antibodies to NAGA, but not with antibodies specific for PABA, arsanilic acid (Ars), phosphorylcholine (PC), or with normal chicken IgG and IgM. The monoclonal anti-PABA ID antibody, termed CId-2, reacted with anti-PABA antibodies and to a lesser extent with anti-Ars antibodies, but not with anti-NAGA, anti-PC, and normal IgG and IgM. The Id determinants were found among antibodies to NAGA and PABA made by outbred and inbred lines of White Leghorn chickens. The binding of the CId-1 and CId-2 antibodies to intact homologous anti-NAGA and anti-PABA antibodies, respectively, was not hapten-inhibitable in either case. Both anti-Id antibodies reacted specifically with isolated homologous heavy chains, suggesting VH Id specificities. The monoclonal CId-1 and CId-2 antibodies were reactive by immunofluorescence with approximately 0.9 and 0.2%, respectively, of the circulating lymphocytes and with approximately 0.4 and 0.15 of plasma cells. CId-1+ and CId-2+ bursal cells were first detected on the 16th and 14th days of incubation, respectively; both reached maximal frequencies by the 17th day of incubation. The CId-2 antibody reacted exclusively with immunoglobulin-positive cells. The CId-1 antibody also reacted with a subpopulation (0.4%) of immunoglobulin-negative lymphocytes from normal and agammaglobulinemic chickens, and thus would appear to recognize an idiotypic determinant expressed by certain clones of B and T cells.     

Xenogeneic antibodies with apparent public idiotypic specificity for anti-Ia.7 antibodies are directed in part against V kappa 21D and E subgroup marker

Public idiotypes (IdX) expressed on monoclonal antibodies (mAb) against a monomorphic alpha-chain determinant of the I-E molecule (Ia.7 epitope cluster I) have been studied by using xenogeneic anti-Id reagents derived from pig, rabbit, and rat. IdX+ anti-Ia.7 mAb were recently demonstrated to be structurally related by a high frequency expression of the V kappa 21E light chain subgroup. This raised the question of whether V region determinants of the IdX were related to V kappa 21E sequences or whether they were unique to hypervariable regions of Ia.7 binding antibodies. To clarify this question, the possible association between the expression of the public Id (IdX(s)Ia.7) and the presence of V kappa sequences (V kappa 21E and/or J kappa segment) was examined. The reactivity of the anti-Id reagents with a random panel of 28 myeloma products (each containing a light chain from one of the different V kappa 21 subgroups) was studied by assaying the ability of these mAb to inhibit the binding between the anti-Id and anti-Ia.7 mAb. This analysis demonstrates that what has previously been defined as IdX Ia.7 includes determinants shared by V kappa 21E and V kappa 21D light chain V regions. The structures recognized are expressed irrespective of the J kappa segment. In addition, this study demonstrates interspecies variation in immune responses to such V kappa 21E antigenic determinants. Additional IdX components are found on anti-Ia.7 mAb but not on other V kappa 21E or D proteins. Thus V region subgroup considerations have crucial implications for Id characterization. In addition, this work describes the first division of the V kappa 21 subgroup into component parts by a mAb.