anti-Id mAb Research Articles

HLA (A*0201) Mimicry by Anti-Idiotypic Monoclonal Antibodies

Soluble MHC Ags and anti-Id (anti-anti-MHC) Abs have both been shown to inhibit MHC alloantigen-specific B cell responses in vivo. We hypothesized that some anti-idiotypic Abs function as divalent molecular mimics of soluble HLA alloantigen. To test this idea, we studied two well-defined anti-idiotypic mAbs, T10-505 and T10-938, elicited in syngeneic BALB/c mice by immunization with CRll-351, an HLA-A2,24,28-specific mAb. Each anti-Id induced "Ab-3" Abs in rabbits that cross-reacted with HLA-A2 but not with HLA-B Ags. Furthermore, each anti-Id could bind to and block Ag recognition by Ha5C2.A2, a human homologue of mAb CRll-351. Both anti-Id mAb displayed weak reactivity with the human mAb SN66E3, which recognized an overlapping but distinct determinant of HLA-A2 Ags; neither reacted with human mAb MBW1, which recognized a nonoverlapping HLA-A2 determinant. Amino acid sequence comparison of mAb CRII-351 heavy and light chain variable region complementarity-determining regions (CDRs) with those of mAb Ha5C2.A2 and SN66E3 revealed short regions of homology with both human mAb; a large insert in the light chain CDR1 of mAb SN66E3 distinguished it from both CRll-351 and Ha5C2.A2. The amino acid sequences of mAb T10-505 and T10-938, which differed markedly from each other, revealed no homology to the alpha2 domain sequence of HLA-A*0201 that contains the CRll-351 mAb-defined epitope. We conclude that structurally different anti-Id Abs can mimic a polymorphic conformational epitope of an HLA Ag. In the case of T10-505 and T10-938 mimicry was not based on exact replication of the epitope by the hypervariable loops of the anti-Id mAb.

Improved tumor targeting of rhenium-186-labeled anti-human high-m.w. melanoma-associated antigen monoclonal antibody 763.74 following purification with anti-idiotypic monoclonal antibody MK2-23.

Because of its high-energy beta emissions and imageable gamma emissions, 186Re represents an attractive isotope to radiolabel monoclonal antibodies (MAbs) recognizing human tumor-associated antigens (TAAs) for radioimmunoscintigraphy (RIS) and radioimmunotherapy (RIT) of patients with malignant diseases. Application of 186Re, however, is hindered by the frequent denaturation of MAbs following exposure to the strong reducing conditions employed in the labeling procedures. To overcome this problem, we have utilized a direct labeling procedure and combined it with affinity chromatography over columns of immobilized anti-idiotypic (anti-id) MAbs which recognize idiotopes in the antigen-combining site of the radiolabeled MAb. The validity of the procedure was demonstrated with the anti-high-m.w. melanoma-associated antigen (HMW-MAA) MAb 763.74 and its corresponding anti-id MAb MK2-23. Utilizing this approach, MAb 763.74 was labeled to specific activities of 2.8 +/- 0.6 mCi/mg with 186Re. Furthermore, its immunoreactivity, which was reduced to about 30% following labeling with 186Re, was improved to about 50% by affinity chromatography over columns of anti-id MAb MK2-23. The improvement in immunoreactivity of 186Re MAb 763.74 resulted in a significant (p < 0.05) increase in targeting to human melanoma tumors grown in nude mice and an increase in sensitivity of RIS. Our results suggest that direct labeling of anti-TAA MAb with 186Re coupled with purification over affinity columns of anti-id MAb may facilitate the application of 186Re to RIS and RIT.

Comparisons of the Ability of Human IgG3 Hinge Mutants, IgM, IgE, and IgA2, to Form Small Immune Complexes: A Role for Flexibility and Geometry

Various native and hinge-modified forms of Ig with identical Ids were reacted with an anti-Id mAb, and the resultant immune complexes were analyzed by negative stain immunoelectron microscopy. Complexes were scored for their geometry (linear versus ring complexes) and size (dimer, trimer, etc.). Ring dimers are the thermodynamically most favorable configuration, unless inhibited by steric and/or flexibility constraints. We found ring dimerization to correlate with the length of the upper, but not middle or lower, hinge. In contrast, the geometry and size of complexes of those molecules lacking formal hinges were unpredictable. A hingeless IgG mutant and native IgE readily formed ring dimers. Remarkably, monomeric IgM formed more ring dimers than any of the other Igs tested, including IgG3. We also tagged the Fab arms and measured the mean Fab-Fab angles and the degree of angular variation for each type of Ig. Surprisingly, IgM proved the most flexible by this assay. In hinged Igs, there was a correlation between length of the upper hinge and Fab-Fab flexibility. In contrast, we found no correlation between the mean Fab-Fab angle in uncomplexed Igs and their ability to dimerize with anti-Id mAb. These data suggest that the physicochemical methods typically used to evaluate molecular flexibility are often of low predictive value when tested in a functional assay.

Monoclonal Antibody Therapy of B Cell Lymphoma: Signaling Activity on Tumor Cells Appears More Important Than Recruitment of Effectors

AbstractDespite the recent success of mAb in the treatment of certain malignancies, there is still considerable uncertainty about the mechanism of action of anti-cancer Abs. Here, a panel of rat anti-mouse B cell mAb, including Ab directed at surface IgM Id, CD19, CD22, CD40, CD74, and MHC class II, has been investigated in the treatment of two syngeneic mouse B cell lymphomas, BCL1 and A31. Only three mAb were therapeutically active in vivo, anti-Id, anti-CD19, and anti-CD40. mAb to the other Ags showed little or no therapeutic activity in either model despite giving good levels of surface binding and activity in Ag-dependent cellular cytotoxicity and complement assays, and in some cases inhibiting cell growth in vitro. We conclude that the activity of mAb in vitro does not predict therapeutic performance in vivo. Furthermore, in vivo tracking experiments using fluorescently tagged cells showed that anti-Id and anti-CD40 mAb probably operate via different mechanisms: the anti-Id mAb cause growth arrest that is almost immediate and does not eliminate cells over a period of 5 or 6 days, and the anti-CD40 mAb have a delayed effect that allows tumor to grow normally for 3 days, but then abruptly eradicates lymphoma cells. This work supports the belief that mAb specificity is critical to therapeutic success in lymphoma and that, in addition to any effector-recruiting activity they may possess, in vivo mAb operate via mechanisms that involve cross-linking and signaling of key cellular receptors.

Induction of Tolerance in B-1 Cells for Bromelain-Treated Mouse Red Blood Cells by a Transient Presence of Anti-Idiotype Antibodies in Neonatal and Adult Mice

Murine B-1 cells are thought to develop from Ig- progenitors early in ontogeny and to expand by self-renewal. To examine the early development of Ig+ precursors of B-1 cells for bromelain-treated mouse RBC, the transient presence of RidA, a rat anti-Id mAb for V(H)11/Vkappa9-type anti-bromelain-treated mouse Abs, was produced in neonatal mice. The presence of RidA during days 0 to 10 of age resulted in an 80% reduction in peritoneal RidA-Id+ B cells and B cells secreting RidA-Id+ Ig after LPS stimulation in 8-wk-old mice. This suggests that most Ig+ precursors for adult RidA-Id+ B cells already exist in 10-d-old mice. However, RidA injected into mice on day 10 had to persist for >4 days to result in a significant reduction in adult B cells. Similarly, although RidA injected into adult mice bound immediately to membrane Ig (mIg) of the peritoneal RidA-Id+ B cells, a RidA persistence for >4 days was required to suppress LPS reactivity of peritoneal and splenic B cells. The binding of RidA to mIg preexisting on B cells has no apparent effect on the ability of neonatal B cells to expand clonally or on the ability of adult B cells to secrete RidA-Id+ Ig after LPS stimulation. Both abilities evidently are suppressed by the accumulation of reaction between freshly expressed mIg and RidA.

Anti-tumor effect of internal image bearing anti-idiotypic monoclonal antibody in relation to GM3 ganglioside.

Anti-idiotypic antibodies are a new type of useful tools for the possible treatment of cancer patients, since some act as antigen specific immunomodulators. Anti-idiotypic monoclonal antibody (anti-Id MAb) D704 (Ab2) was established which bore the internal image of the determinant defined by MAb M2590 (Ab1) against a sialic acid residue on GM3 ganglioside. In an in vivo syngeneic tumor system, anti-Id MAb D704 was more effective in preventing tumor progression, as compared with anti-GM3 MAb or no treatment. Significant suppression of tumor growth and prolongation of survival by administration of anti-Id MAb D704 in an animal group inoculated with 1 x 10(4)/mouse melanoma cells were seen, but not in a group inoculated with 5 x 10(4) cells/mouse. In an active specific immunotherapy protocol utilizing Ab2, the activity of anti-anti-Id antibodies (Ab3) specific for GM3 (antigen) which has a weak immunogenicity only, was maintained for more than 3 months. Ab2 generated cellular anti-tumor immune responses, including delayed type hypersensitivity (DTH) reaction. Immunohistological analysis indicated a marked infiltration of CD4 and CD8 positive cells into the DTH sites. Our results suggest that internal image bearing anti-Id MAbs have a therapeutic potential against tumors if the number of melanoma cells is relatively low or if hosts are at an early stage of melanoma progression.

Anti-idiotype monoclonal antibodies specific for the MOPC167 anti-phosphocholine transgene-encoded antibody.

Four rat x mouse hybridomas secreting monoclonal anti-idiotypic (anti-Id) antibodies (MAb) specific for the transgene-encoded antibody of the 207-4 transgenic mouse line, which carries the VH1/V kappa 24 gene segments of the IgA, phosphocholine-(PC) specific MOPC167 myeloma, were developed from a fusion of Ag8-X63.653 mouse cells with spleen cells from a rat immunized with MOPC167 and HPCM27 anti-PC antibodies. The anti-Id MAb were shown by ELISA to be specific for PC-binding proteins of VH1/V kappa 24 H and L chains of various isotypes. They did not bind VH1/V kappa 22, VH1/V kappa 8, or VH1/V kappa 1 PC-binding proteins or other IgA or IgM myeloma proteins. Analysis by flow cytometry demonstrated that these MAb bind to the transgene-encoded membrane immunoglobulin (sIgM) as expressed on > 95% of the B220 positive 207-4 spleen cells. All four MAb were able to inhibit the binding of MOPC167 to PC conjugated to bovine serum albumin. Differences in fine specificity of binding were demonstrated by differential staining of spleen cells of the 216-7 mu kappa delta Mem MOPC167 transgenic mice. In these mice endogenous H chains associate with the transgene encoded L chain to form MOPC167 crossreactive idiotopes. Two of the MAb, 28-4-3 and 28-6-20, stained significant numbers of cells, while MAb 28-5-15 did not bind to 216-7 cells. Three of the MAb, 28-5-15, 28-6-20, and 28-4-3, when conjugated to Sepharose beads, were able to induce DNA synthesis in cultures of 207-4 transgenic spleen cells. None of the MAb were able to induce an antibody response in vivo. These MAb should prove useful in staining PC-transgenic B cells for flow cytometry studies and in defining early cellular events in the activation of idiotype positive B cells by anti-Id antibodies.

Flexibility of human IgG subclasses.

A variable region (Id)-matched set of genetically engineered human IgG1, -2, -3, and -4 subclass molecules was analyzed by electron microscopy for hinge-mediated differences in flexibility. The hinge-mediated bending was studied, as was the ability of the subclasses to form immune complexes with two anti-Id mAbs. The data show that the rank order (most to least flexible) of the IgG subclasses for hinge-folding mode of flexibility between Fab arms is IgG3 > IgG1 > IgG4 > IgG2. The mean Fab-Fab angles for the subclasses are IgG3, 136 degrees; IgG4, 128 degrees; IgG2, 127 degrees; and IgG1, 117 degrees. Fab-Fc angles were similarly analyzed. By sampling of equimolar mixtures of Id-bearing IgGs and each of two anti-Id mAb after incubation over time (1.5 min to 3.5 h), different kinetic profiles of immune complex formation of defined geometry were documented. Both anti-Id mAbs displayed unique kinetic profiles when complexed with the four IgG subclass molecules but also shared important features. Most notable was the higher propensity to form closed bivalent ring Id-anti-Id dimers with IgG3 than with IgG2 and IgG4. IgG1 was intermediate in its ability to form such dimers.

Active specific immunotherapy with mouse anti-idiotypic (anti-id) mAb MK2???23

Active specific immunotherapy with mouse anti-idiotypic (anti-id) mAb MK2???23

Purification by affinity chromatography with anti-idiotypic monoclonal antibodies of immunoreactive monoclonal antibodies following labeling with 188Re.

Because of its high energy beta emissions and imageable gamma emissions, 188Re represents an attractive isotope to radiolabel monoclonal antibodies (mAb) recognizing human tumor-associated antigens for radioimaging and radioimmunotherapy in patients with malignant diseases. The application of 188Re is, however, hindered by the denaturation of a sizable proportion of antibody molecules during the labeling process. To overcome this problem, we have combined radiolabeling of mAb with 188Re with purification of immunoreactive 188Re-labeled mAb by affinity chromatography over columns of anti-idiotypic (anti-id) mAb. Utilizing the anti-high-molecular-weight melanoma-associated antigen (HMW-MAA) mAb 763.74 as a model system, we found that the immunoreactivity of mAb 763.74 labeled with 188Re to a specific activity of 1 mCi/mg increased from about 50% to at least 80% following passage over columns of immobilized anti-id mAb. Moreover, between 90-100% of immunoreactive mAb contained in radiolabeled preparations could be recovered from anti-id mAb columns. These results indicate that the procedure we have described may facilitate the application of 188Re for immunoscintigraphy and immunotherapy of malignant diseases.

Induction and characterization of monoclonal anti-idiotypic antibodies to tick-borne encephalitis virus neutralizing antibody.

Seven monoclonal anti-idiotypic antibodies (anti-ID MAbs, Ab2) were generated against virus-neutralizing and hemagglutination-inhibiting monoclonal antibody (Ab1) specific for tick-borne encephalitis (TBE) virus. Six of these anti-ID MAbs inhibited the binding of Ab1 to the virus antigen, thus classifying these anti-ID antibodies as Ab2 beta or AB2 gamma. Inhibition tests with heterologous anti-TBE sera revealed that these anti-ID MAbs were not recognized by anti-TBE antibodies and therefore they do not carry an internal image of TBE virus antigen. Hence, the anti-ID MAbs may be classified as Ab2 gamma type. None of the anti-ID MAbs induced production of antiviral antibodies nor protective immunity in syngeneic Balb/c mice. Using these anti-ID MAbs four nonoverlapping idiotopes were identified on Ab1 variable region.

Idiotypic diversity and variable region gene usage by mouse anti-HLA-DQ3 mAb.

The anti-HLA-DQ3 monoclonal antibodies (mAb) KS13, SO1, SO2, SO3, SO4, and SO5 recognize spatially close but distinct antigenic determinants, since they crossinhibit each other in their binding to HLA-DQ3 antigens, but do not share idiotopes recognized in their antigen combining site by syngeneic and anti-id antisera and mAb. Furthermore, mAb SO1, SO3, SO4, and SO5 react also with HLA-DQ allospecificities other than HLA-DQ3. Sequence analysis of the heavy (VH) and light (VL) chain variable region of the six mAb revealed preferential usage of VH 36-60 and VK 12/13 gene families. However, the individual VH and VL germline gene usage by the six mAb is diverse and the utilization of D, JH, and JL gene segments is heterogeneous. The diverse usage of VH and VL gene segments and heterogeneous amino acid sequences of VH and VL CDR, together with the heterogeneous idiotypic profile, may reflect the complexity of the determinants recognized by the six mAb on HLA-DQ3 antigens. The results we have presented provide for the first time information about the structural basis of the diversity of antibodies recognizing human histocompatibility antigens.

Antiantiidiotypic Response and Clinical Course of the Disease in Patients with Malignant Melanoma Immunized with Mouse Antiidiotypic Monoclonal Antibody MK2-23

An active specific immunotherapy trial has been implemented in patients with malignant melanoma utilizing the mouse anti-id MAb MK2-23. The latter bears the internal image of the determinant by the anti-HMW-MAA MAb 763.74. The immunogenicity of anti-id MAb MK2-23 is enhanced by conjugation to a carrier and administration with an adjuvant. Anti-id MAb MK2-23 induced humoral anti-HMW-MAA immunity in about 60% of the immunized patients. The latter was associated with a statistically significant survival prolongation. It is suggested that anti-HMW-MAA immunity may have a beneficial effect on the clinical course of the disease by inhibiting the function of HMW-MAA in the biology of melanoma cells.

Molecular cloning of cDNAs for immunoglobulin variable regions of a monoclonal anti-idiotypic antibody specific for sigma receptors

In previous work on sigma (σ) receptors, we established a hybridoma cell line, 10G9, producing a monoclonal anti-idiotypic antibody (anti-Id mAb) to the monoclonal anti-haloperidol antibody. The anti-Id mAb showed specific binding affinity to σ receptors in immunoprecipitation and competition experiments. Here we isolated and sequenced cDNA clones for the variable regions of the anti-Id mAb. These combinations of segments and formation of the unique CDR3 were considered to be the structural basis for the diversity of the antigen-binding site of our anti-Id mAb and, therefore, for its binding activity to a receptors.

Human high molecular weight-melanoma associated antigen mimicry by mouse anti-idiotypic MAb MK2-23. Immunohistochemical analysis of the reactivity of anti-anti-idiotypic MAb with surgically removed melanoma lesions.

The mouse anti-id MAb MK2-23 bears the internal image of the antigenic determinant defined by anti-HMW-MAA MAb 763.74. Active specific immunotherapy with anti-id MAb MK2-23 is associated with a statistically significant prolongation of survival in patients with malignant melanoma. Characterization of the immune response elicited by anti-id MAb MK2-23 may contribute to our understanding of the mechanisms underlying the apparent beneficial clinical effects of active specific immunotherapy with anti-id MAb MK2-23. Therefore, 8 HMW-MAA binding anti-anti-id MAbs elicited with MAb MK2-23 were characterized in their reactivity with a large panel of surgically removed benign and malignant tumors and of normal tissues. The 8 anti-anti-id MAbs displayed subtle differences in their immunoperoxidase staining of both benign and malignant lesions and of normal tissues. The diversity in the fine specificity of the 8 anti-anti-id MAbs is likely to reflect the few somatic mutations which occur in the amino-acid sequence of the variable regions of their heavy and light chains in the course of the immune response to MAb MK2-23. The reactivity patterns of the 8 anti-anti-id MAbs with the tissue substrates are similar, although not superimposable upon that of anti-HMW-MAA MAb 763.74 elicited with melanoma cells. This difference may reflect the imperfect mimicry by anti-id MAb MK2-23 of the antigenic determinant defined by anti-HMW-MAA MAb 763.74.

Internal Image-Bearing Anti-ldiotypic Monoclonal Antibodies

Five anti-idiotypic (Id) monoclonal antibodies (MAbs) (Ab2) were prepared from a BALB/c mouse immunized with anti-carcinoembryonic antigen (CEA) MAb MA208 (Ab1) in a syngeneic system. These anti-Id MAbs appear to recognize unique idiotopes at the combining site of MAb MA208, because they were specifically reactive with MAb MA208 and showed inhibitory activity against the binding of MAb MA208 to CEA. These MAbs were divided into three groups according to the analysis of anti-anti-Id antibodies (Ab3) induced with each anti-Id MAb. Anti-anti-Id MAb M7-625 antiserum (Ab3) reacted with purified CEA in a binding assay and in Western blot analysis, and competed with Ab1 binding to CEA. Furthermore, the binding of anti-Id MAb M7-625 to MAb MA208 was inhibited with CEA, indicating that Ab2 mimics the structure of the epitope in CEA which was recognized with Ab1. These serologic findings suggest that anti-Id MAb M7-625 carries the internal image of the antigen. According to the amino acid sequences of complementarity determining region (CDR) 1, 2 and 3 of the MAb M7-625 variable region, homology of amino acid sequences exists between CDR2 in the H chain (5 amino acids of 10) and domain III of CEA (545-554). Seven anti-Id MAbs were then generated using anti-CEA synthetic peptide MAb P1-356 to analyze further the epitope structure of CEA. These anti-Id MAbs were divided into four groups. Serological analyses as described above suggested that among them, anti-Id MAb M315 had an internal image. We therefore prepared anti-anti-Id MAbs using anti-Id MAb M315. Among them, anti-anti-Id MAb 11B2 reacted directly with CEA and competed with MAb P1-356 in the competition assay. In addition, MAb 11B2 stained both cultured CEA-producing cells and colonic cancer tissues, suggesting that MAb 11B2 is Ab1 like Ab3. These MAbs (Ab1-3) will be of use for the structural analysis of the internal image.

Monoclonal Antibody Therapy of Leukemia and Lymphoma

Publisher Summary Through careful clinical trials, a number of obstacles to effective therapy with monoclonal antibodies (mAbs) have been identified. New technologies, however, have continued to provide strategies to overcome many of these obstacles. Four approaches—mAbs can be used as mediators of immune effector function, anti-idiotype (anti-id) mAbs can both target unique surface immunoglobulins expressed on B-cell neoplasms and mimic an antigen triggering tumor immunity, mAbs can be directed against receptors that control tumor growth, and mAbs can serve as vehicles to carry cytotoxic agents—to the therapeutic use of mAbs have been developed. Among the most promising applications of mAb-based therapies are those that take advantage of multiple mechanisms of tumor killing, such as the use of radioconjugates in combination with high-dose chemotherapy and TBI prior to bone marrow transplantation (BMT), the use of mAb-based therapies to eliminate residual disease, and the use of mAbs to purge tumor cells from bone marrow prior to reinfusion after dose-intensive therapy. The immunophenotypic characterization of the various stages and lineages found during hematopoietic differentiation has formed the basis for the selection of therapeutic mAbs. Leukemia- and lymphoma-associated antigens, however, are neither tumor specific, nor are they always stage or lineage specific. Therefore, myelosuppression—that affects the treatment of leukemia greatly—is often associated with mAb therapy of hematologic malignancies.

Production and characterization of monoclonal anti-idiotypic antibodies to muramylpeptide

Hybridomas producing monoclonal anti-idiotypic antibodies (anti-id MAbs) to N-acetylglucosaminyl-β1-4- N-acetylmuramyl-alanyl- d-isoglutamine (GMDP) were developed. Three clones of hybridomas demonstrated the properties characteristic for the Ab2β type of anti-id antibody: they bound to Fab-fragments of high-affinity MAb to GMDP; dose-dependent inhibition of this binding by GMDP was observed; immunization of mice with these MAbs resulted in production of GMDP-specific antibodies. When these antibodies were used to stain blots from SDS-PAGE of macrophage lysate, the same receptor proteins were specifically stained as upon staining with 125I-labelled GMDP derivative.

Analysis of B Cell Repertoire Specific to the Neutralizing Epitopes of Glycoprotein 120 in HIV-Infected Individuals

This study describes the clonotypic analysis of neutralizing anti-gp120 antibodies elicited in HIV-infected individuals by a panel of anti-idiotype monoclonal antibodies (anti-Id MAbs). Sera from 80 HIV-infected individuals at various clinical stages of HIV-infection were tested for reactivity to 19 anti-Id MAbs in ELISA. Anti-idiotype MAbs reacted with between 0 and 26% of sera. Among the 13 idiotypes specific for anti-CD4 site antibodies, 4 were expressed in 15 to 20% of individuals, whereas 2 of 4 idiotypes specific for anti-V3 antibodies were expressed in 15 to 26% of the cases. These data suggest that each HIV-infected individuals has a diverse B cell repertoire to a given neutralizing epitope cluster and that certain clonotypes are more prevalent than others. To correlate the binding activity in ELISA with anti-gp120 specificity, the idiotype-positive antibodies (Id+ Abs) from representative serum samples were isolated by anti-Id MAb-Sepharose affinity columns. In most cases, the epitope specificity and the neutralizing properties of the isolated Id+ Abs correlated with that of anti-gp120 antibodies used for the generation of anti-Id MAbs. We propose that these anti-Id MAbs may be used to identify and measure neutralizing anti-gp120 antibodies of defined specificity in the sera of HIV-infected individuals, HIV-vaccinated individuals, and in HIV-infected mother-infant pairs.

Structural profile of idiotype, anti‐idiotype and anti‐anti‐idiotype monoclonal antibodies in the HLA‐DQ3 antigenic system

Interest in the characterization of idiotype cascades in the HLA antigenic system has been stimulated by their potential role in the immune response to mismatched HLA allospecificities and in the survival of kidney allografts. Since no information is available about the structural organization of idiotypic cascades in the HLA system, we have sequenced the variable regions of the heavy (VH) and light (VL) chains of mouse anti-HLA-DQ3 monoclonal antibody (mAb) KS13 elicited by cell membrane-bound antigens, of syngeneic anti-HLA-DQ3 mAb S2B154 elicited by anti-idiotypic (anti-id) mAb K03-34 and of five syngeneic anti-id mAb elicited by mAb KS13. mAb KS13 and S2B154, which have been previously shown to be very similar in their specificity and idiotypic profile, share several structural characteristics. Their VH and VL regions are encoded by the same VH, VK and JH genes, display relatively similar V(D)J rearrangements and differ only through a few amino acid substitutions. Among the five anti-id mAb elicited by mAb KS13, mAb R1-38 and R18-9 utilize multiple genetic elements that are different from those used by anti-id mAb KO3-34, K03-256 and K03-335. These results indicate that diverse V region combinations can confer an anti-id specificity in the antigenic system analyzed. mAb K03-34, K03-256 and K03-335 originate from the same B cell clone, since they use the same V, D and J genes and possess identical V(D)J rearrangements. The latter three anti-id mAb differ only by point mutations, which have dramatic effects on the HLA-DQ3 antigen mimicry properties of the three anti-id mAb. mAb K03-34 is the only one to induce anti-HLA-DQ3 antibodies both in syngeneic and xenogeneic hosts. The antigen mimicry properties of anti-id mAb K03-34 depend upon its three-dimensional conformation, since no significant amino acid sequence homology has been found between its VH and VL regions and alpha 1 and beta 1 domains of HLA-DQ3 antigens.