Idiotypic analysis of anti-I-Ak monoclonal antibodies. III. T- and B-cell responses to anti-Ia idiotopes are not modulated by syngeneic anti-idiotypic monoclonal antibodies

To investigate whether anti-idiotypic (anti-Id) antibodies activate T cells either directly or indirectly, we examined the ability of syngeneic anti-Id monoclonal antibodies (mAbs) to regulate idiotype (Id) expression, antigen-binding antibody production, and T-cell reactivity to antigen. Our idiotypic system consists of an anti-I-A mAb that carries an infrequently expressed Id. Using three syngeneic anti-Id mAbs (Ab2), we previously defined the idiotype of the 11-5.2.1.9 (11-5) anti-I-Ak mAb. Two of these mAbs, IIID1 and IA2, recognize the same or closely related epitopes on 11-5 and cross react with two additional anti-I-Ak mAbs, 8B and 39J; the third anti-Id mAb, VC6, recognizes a distinct epitope shared by 11-5 and 8B. In the present study, BALB/c (H-2d) mice were primed with varying doses of these anti-Ids and were then boosted with C3H (H-2k) spleen cells. Among 130 such primed mice, the syngeneic anti-Ids when tested at priming doses between 10 ng and 10 micrograms were unable to induce Id production. The priming anti-Id mAbs persisted in the serum of the mice and were detectable as late as 40 days after priming. Ab1 expression was not modulated in BALB/c mice immunized with KLH-coupled Ab2, however, this immunization elicited the production of Ab3 which shared idiotypes with 11-5, 8B, and 39J. BALB/c anti-C3H alloreactive T-cell clones were also not induced by anti-Id priming, nor could they be shown to bind directly to the three Ab2 used. Nevertheless, the proliferative response of one anti-I-Ak specific T-cell clone that recognizes the same epitope as 11-5, 8B, and 39J, was inhibited by the IIID1 and IA2 Ab2. Thus, a T cell can express an idiotype shared by a B cell, but the linked recognition of an Id-associated carrier determinant(s) by an alloreactive T cell is required to elicit an anti-Id antibody response. These results favor the possibility that the activation of T cells is not dependent upon their ability to bind to anti-Id, but rather on their capacity to respond to epitopes of Id-anti-Id antigen-antibody complexes formed on B cells.     

Analysis of the anti-alpha 1 leads to 3 dextran response with monoclonal anti-idiotype antibodies

The antibody response to alpha 1 leads to 3 dextran (DEX) in BALB/c mice consists of a family of closely related yet highly heterogeneous molecules. Although these antibodies have been previously characterized both idiotypically and structurally, detailed analysis of responding clones has not been possible using conventional anti-idiotype antibodies. Monoclonal syngeneic and allogeneic anti-idiotype antibodies (MAIDs) specific for anti-DEX antibodies were used in this study to dissect the serum antibody response to DEX in BALB/c mice. The constructed MAIDs showed considerable heterogeneity by isoelectric focusing and by their binding characteristics to a series of DEX specific myeloma and hybridoma proteins. The predominant heavy chain isotype of these MAIDs was gamma 1. These antibodies were used to identify individual idiotypic structures (IdI) on J558, or M104E as well as cross-reactive determinants common to both (IdX). Although both IdX and IdI MAIDs were obtained, IdI specific antibodies were obtained more frequently. BALB/c mice immunized with DEX produced antibodies expressing both IdI but in highly variable amounts. A large percentage of, but not all DEX specific antibody, could be accounted for by IdX bearing antibodies. Suppression of adult and neonatal mice by IdI specific MAIDs was effective with precise elimination of only those clones expressing IdI determinants leaving the total lambda bearing anti-DEX response intact. Suppression of adults and neonates by an IdX specific MAID resulted in a temporary and partial suppression of the total lambda bearing anti-DEX response along with total suppression of the IdX portion of the response. Unlike other systems these monoclonal antibodies produce only suppression, and under a variety of conditions enhancement of anti-DEX responses has not been observed.     

Evidence that anti-idiotype induced immunity to experimental African trypanosomiasis is genetically restricted and requires recognition of combining site-related idiotopes

The ability of anti-idiotypic (anti-Id) antibodies to immunize mice against African trypanosomiasis independent of antigen has been confirmed. Of three allogeneic anti-Id antibodies raised against three protective monoclonal antibodies, each with specificity for the variant surface antigen of a clone of Trypanosoma rhodesiense, only one (anti-7H11 Id) was effective in immunizing BALB/c mice against homologous challenge. The immunity was associated with the more rapid and enhanced expression of the corresponding Id after infection. The immunity was restricted to mice bearing genes linked to Igh-Ca, which appeared to control expression of this Id both in response to infection and anti-Id treatment. Another Id, 11D5, appeared to be under similar genetic control. Anti-11D5 Id, however, was ineffective in immunizing mice against infection despite inducing high levels of Id bearing molecules before challenge. The immunizing potential of the respective anti-Id antibodies appeared to be related to the relative concentrations of antibodies reactive with idiotopes near to or within the antigen-combining site, which, in turn, determined the relative proportion of Id-bearing clones activated that had antigen binding activity.     

Junctional diversity of H and L chains allows the coexpression of two mutually exclusive idiotopes (IdI104 and IdI558).

The molecular basis for the unexpected coexpression of the individual Id (IdI)558 and IdI104 Id by anti-alpha(1-3) DEX antibody (Ab) (126.33 and 414.2) derived from the MPW wild mouse strain has been investigated by the comparison of the structures of their VH and V lambda 1 chain regions with those of two other MPW-derived Ab (262.9 and 16.3) expressing either IdI558 or IdI104 Id. Our data show that 262.9 and 16.3 Ab display identical V lambda 1 and very similar VH regions when compared with BALB/c anti-alpha (1-3) dextran Ab expressing IdI104 or IdI558, respectively. The two Ab (414.2 and 126.33) that express both IdI104 and IdI558 Id display two main features. First, their VH CDR3 are different from those found in IdI104 or IdI558 expressing anti-alpha(1-3) dextran Ab. Second, their V lambda 1 are identical to those from BALB/c origin except for the presence of an additional residue, a phenylalanine at position 95A of CDR3. This additional residue is encoded by the V lambda 1 gene segment and results from a hitherto undescribed V lambda 1-J lambda 1 junction. The alteration of the length of the V lambda 1 CDR3 loop, in conjunction with particular residues within VH CDR3, allows the coexpression of two Id that were found to be mutually exclusive in laboratory mice.     

Monoclonal anti-idiotypic antibodies induce humoral immune responses specific for simian virus 40 large tumor antigen in mice.

Mouse monoclonal anti-Id antibodies were generated against a mouse mAb (Ab-1) preparation specific for SV40 large tumor Ag (T-Ag). Four monoclonal anti-Id preparations each inhibited the binding of the monoclonal anti-SV40 T-Ag Ab-1 preparation to SV40 T-Ag. These anti-Id preparations appeared to recognize similar idiotopes on the monoclonal anti-SV40 T-Ag Ab-1 based on competitive cross-inhibition studies. One of these anti-Id preparations, designated 57B, was examined further for its in vivo modulatory capacity in mice. This anti-Id induced an Ab-3 response in BALB/c mice that recognized SV40 T-Ag (Ag+) and expressed an Id that was shared by the monoclonal anti-SV40 T-Ag Ab-1 preparation (Id+). The Id expressed on the Ab-3 differed from the Id induced in BALB/c mice immunized with the nominal SV40 T-Ag. Furthermore, characterization of the humoral immune response induced by anti-Id immunization indicated that the Ab-3 also recognized different epitopes on SV40 T-Ag when compared to the anti-SV40 T-Ag Ab-1 preparation used to generate the anti-Id. These studies indicate that monoclonal anti-Id can be used to induce humoral immune responses to a viral encoded tumor-associated Ag in vivo with 1) and Id specificity that differs from that expressed on antibodies produced by immunization with the nominal Ag and 2) an epitope specificity distinct from the Ab-1 preparation used for the production of the anti-Id.     

Shared idiotope on monoclonal anti-Ia.7 antibodies reactive with determinants in a structural domain of the I-E molecules

In previous studies, heterologous anti-idiotypic (anti-Id) antisera against the C3H.SW 14-4-4S or the A.TH 41.A anti-Ia.7 monoclonal antibodies (mAb) were shown to identify an interstrain cross-reactive idiotypic specificity (IdX.Ia.7) expressed on monoclonal or conventional anti-Ia.7 alloantibodies. The objective of the present investigation was to characterize further this IdX at the idiotopic level. To this end, 11 hybridomas producing IgG1, IgG2a, or IgM anti-Id mAb were derived from a rat immunized with a mixture of 10 A.TH or A.BY anti-Ia.7 mAb. The specificity of the latter anti-Id mAb was determined by direct Id binding radioimmunoassay (RIA) with the use of a panel of 52 anti-Ia mAb derived from hybridomas produced in various inbred mouse strains. These rat anti-Id mAb recognized idiotopes expressed on i) all anti-Ia.7 mAb against determinants in the topographic domain I of the I-Ek molecule but not on 18 other anti-I-Ek mAb directed at epitopes in domains II or III; ii) three of 19 anti-I-Ak mAb; and iii) one A.TL-derived anti-I-As mAb. Competitive Id binding assays revealed that among the 14 IdX+ anti-Ia.7 mAb, one (81.B) was bound to a lesser extent by various rat anti-Id mAb, suggesting that heterogeneity probably exists in this antibody family. By contrast, two isologous (B10.S(7R)) anti-Id mAb to the IdX.Ia.7+ mAb 41.A displayed a specificity restricted to 41.A individual idiotopes (IdI). Rat anti-IdX.Ia.7 and mouse anti-41.A IdI mAb inhibited the binding of 125I-labeled mAb 41.A to CBA spleen cells. These two sets of mAb bound in a noncompetitive fashion to mAb 41.A-coated plates, indicating that their corresponding public or private idiotopes were spatially distinct. These data may have implications for in vivo manipulations of anti-Ia immune responses.     

V kappa gene usage, idiotype expression, and antigen binding among clones expressing the VHX24 gene family derived from naive and anti-idiotype immune BALB/c mice

The BALB/c myeloma protein ABPC48 binds beta(2-6)-linked fructosans and expresses genes derived from the VHX24 and V kappa 10 gene families. We have selected 30 hybridomas expressing the VHX24 gene family derived from mitogen-stimulated spleen cells of naive BALB/c mice and mice injected at birth with the syngeneic monoclonal anti-ABPC48Id, IDA10. The majority of mAb with kappa L chains uses V kappa 1. Antibodies reacting with IDA10 use both V kappa 10 and V kappa 1. Most of these VHX24+ mAb reacted with one or more members of a limited panel of predominantly polysaccharide Ag that have been previously observed to interact with antibodies expressing the VHX24 gene family. Nucleotide sequencing of selected VH and V kappa genes shows a very low frequency of somatic mutation. The effect of neonatal anti-Id injection on VHX24-V kappa pairing and Id expression is discussed.     

Influence of the immunoglobulin heavy chain locus on expression of the VK1GAC light chain

The VK1GAC light chain represents the dominant V kappa structure employed in the antibody response of A/J mice to streptococcal group A carbohydrate ( GAC ). Two anti-idiotypic antisera, anti- Id5 and anti- Id20 , with specificity for the VK1GAC light chain were used to examine anti- GAC antibody responses in a series of inbred mouse strains that differ at the heavy chain constant region ( IgCH ) allotype locus. Both idiotypes were expressed in normal and immune sera from mice of most IgCH allotypes, except IgCHb (C57BL/6J) and IgCHf (CE/J). C57BL/6J mice expressed Id5 , but not Id20 , whereas CE/J mice did not express either idiotype. Testing of recombinant inbred strains between BALB/c and C57BL/6 indicated that the pattern of idiotype expression did not correlate with IgCH allotype. The C X B recombinants expressed all three idiotype patterns that were observed in the panel of inbred strains. Testing of allotype congenic mice between BALB/c and C57BL/6 showed that CB.20 and BC.8 mice were Id20 -, whereas BAB-14 mice were Id20 +, indicating that both VH and background (V kappa or regulatory) loci must be derived from BALB/c to obtain Id20 expression. The difference in the frequency of idiotype expression observed between BALB/c and BAB-14 mice indicates that the IgCH locus may exert a quantitative influence on the expression of this light chain. To examine the Id20 -, Id5 + antibodies of C57BL/6 mice, anti- GAC hybridomas were prepared. Of 16 C57BL/6-derived anti- GAC monoclonal antibodies, six were reactive with anti- Id5 and not with anti- Id20 . Isoelectric focusing of the purified kappa light chains from three of these antibodies revealed two distinct spectrotypes that co-migrated with the two known VK1GAC spectrotypes observed with A/J anti- GAC light chains. Idiotypic analysis of in vitro recombinants between the heavy and light chains of A/J and C57BL/6 monoclonal antibodies demonstrated that the C57BL/6 light chains were idiotypically similar to A/J light chains when they were free in solution or paired with A/J heavy chains. These results demonstrate that C57BL/6 mice can express a light chain that is very similar, if not identical, to the VK1GAC light chain, although the light chain is expressed in lower frequency and is paired with a distinct VH structure, which can mask expression of one of the VK1GAC idiotypes. These effects on V kappa expression map to at least three genetic loci: VH, CH, and an unlinked locus.     

Comparison of the murine humoral immune response to recombinant simian virus 40 large tumor antigen: Epitope specificity and idiotype expression

Baculovirus-derived recombinant simian virus 40 (SV40) large tumor antigen (SV40 T-Ag) was used to immunize inbred strains of mice to compare the humoral immune responses. Specifically we examined the epitope specificities and idiotype (Id) expression on anti-(SV40 T-Ag) responses induced in BALB/c and C57BL/6 inbred strains of mice. The predominant SV40 T-Ag epitopes recognized by the anti-(SV40 T-Ag) responses appeared to differ between these two inbred strains, this being based on the ability of sera to inhibit the binding of several murine monoclonal antibodies specific for SV40 T-Ag. In addition, anti-(SV40 T-Ag) responses produced in C57BL/6 mice failed to express a previously described cross-reactive Id expressed in the anti-(SV40 T-Ag) response in BALB/c mice. This cross-reactive Id is detected by a mouse monoclonal anti-Id, designated 58D, which has been shown to represent a potential focal point for manipulating the humoral immune response to SV40-induced tumors in BALB/c mice. Together, these data indicate that the functional duality of the humoral immune response, as assessed by epitope recognition and Id expression, differs between these two inbred strains of mice when immunized with a recombinant SV40 T-Ag.     

Generation and specificity of monoclonal anti-idiotypic antibodies against human HIV-specific antibodies. I. Cross-reacting idiotopes are expressed in subpopulations of HIV-infected individuals

In this study we have generated monoclonal anti-idiotypic antibodies against human monoclonal and polyclonal anti-HIV antibodies in seropositive sera. A human anti-gp41 mAb (H2, IgM kappa) was used to immunize BALB/c mice and to prepare hybridoma anti-antibodies that react with H2 and not with normal human IgM. Similar monoclonal anti-antibodies were made in BALB/c mice immunized with Ig fraction prepared from a pool of HIV-seropositive sera. Both kinds of anti-idiotypic antibodies reacted with antibodies in pools of seropositive sera and with individual seropositive sera but not with normal human Ig or seronegative sera. The Id-positive Ig from single donors were isolated on two different anti-Id immunoabsorbents and shown to bind to p24 and gp120, respectively. The detection and isolation of idiotypically cross-reactive human anti-HIV antibodies from seropositive donors demonstrated, for the first time, the existence of shared Id expressed by antibodies against HIV Ag. The utility of cross-reacting anti-idiotypic antibodies as tools to dissect the network regulation of the anti-viral immunity in AIDS is discussed.     

Properties of anti-idiotypic T cell lines propagated with syngeneic B lymphocytes. I. T cells bind intact idiotypes and discriminate between the somatic idiotypic variants in a manner similar to the anti-idiotopic antibodies

We describe the development of T cell lines possessing a binding specificity for syngeneic T15 idiotopes (Id) expressed on phosphorylcholine (PC)-reactive Ig. The lines were obtained by cultivation of BALB/c splenic T cells with T15 Id+ stimulator cells BCg3R-1d, a BALB/c lymphoma transfected with genomic sequences mu and kappa with S107 (T15) variable regions. Resulting Thyl-2+, L3T4+ cell lines depend on the T15 Id+ BCg3R-1d cells for growth and demonstrate the ability to bind TEPC15, a S107 germline-encoded, PC-specific Ig alpha. The specificity of the 125I-TEPC15 binding was studied in a competitive RIA with various unlabeled Ig. The isolated H and L chains of TEPC15 failed to inhibit the 125I-TEPC15 binding, and the T15-, PC-binding proteins M603 (alpha) and M511 (alpha) inhibited the binding either poorly or not at all. Moreover, the T cell lines had a discriminatory binding specificity for various T15+ Ig that are somatic variants of TEPC15 and that differ from each other in discrete, conformational Id (epitopes) detectable with specific monoclonal anti-Id. The T cell lines could be grouped according to their binding patterns, which were comparable to the recognition patterns of certain monoclonal anti-Id. These data suggest the existence of T cells with specificity for serologically-defined determinants of syngeneic idiotypes.     

Autoimmunity induced by HgCl2 in Brown-Norway rats. II. Monoclonal antibodies sharing specificities and idiotypes with mouse natural monoclonal antibodies

Spleen cells derived from BN rats receiving HgCl2 were fused with the nonsecreting rat myeloma cell line IR983F. We screened 59 supernatants from immunoglobulin-secreting hybrids for antibody activity against actin, tubulin, autologous and heterologous myosin, myoglobin, dsDNA, peroxidase, and the haptens TNP, NIP, NNP, and NBrP. Six monoclonal antibodies (mAb) were found to react with antigen(s) of the panel. At least three groups of antibody specificities were identified: clones reacting with TNP (1 IgM, 1 IgE); clones reacting with horseradish peroxidase (1 IgM); and clones possessing widespread reactivity for several antigens as found for mouse natural autoantibodies (2 IgM, 1 IgE). We also analyzed the idiotypic (Id) determinants of the 59 mAb by using anti-Id antibodies described elsewhere prepared in rabbits against the BALB/c D23 natural monoclonal autoantibody and recognizing a BALB/c recurrent Id (Id D23) of natural polyspecific autoantibodies. We found that all rat mAb that possessed widespread reactivities bore this Id. We performed similar studies in sera from normal and mercury-stimulated rats. The results indicate a role for HgCl2 in the stimulation of natural antibodies producing cells and the existence of interspecies cross-reactive Id among mouse and rat natural antibodies.     

Complete heavy and light chain variable region sequence of anti-arsonate monoclonal antibodies from BALB/c and A/J mice sharing the 36-60 idiotype are highly homologous

Structural and serologic studies on murine A/J monoclonal anti-arsonate antibodies resulted in the identification of a second idiotype family (Id36-60) in addition to the predominant idiotype family (IdCR). Id36-60, unlike IdCR, is a dominant idiotype in the BALB/c strain but is a "minor" idiotype in the A/J strain. The complete heavy and light chain variable region (VH and VL) amino acid sequences of a representative Id36-60 hybridoma protein from both the A/J and BALB/c strains have been determined. There are only four amino acid sequence differences between the VH of antibody 36-60 (A/J) and antibody 1210.7 (BALB/c). Two of these differences arise from single nucleotide changes in which the A/J and BALB/c Id36-60 VH germline gene sequences differ. The two other differences are the result of somatic mutation in hybridoma protein 36-60. In addition, Id36-60 heavy chains employ the same D and JH3 segments in both strains. The entire Vk2 VL of 36-60 and 1210.7 differ by only two amino acids, suggesting that like the heavy chains, they are derived from highly homologous VL genes. The same Jk segment is used in both antibodies. A comparison of the amino acid sequence data from Id36-60-bearing hybridomas suggests that a heavy chain amino acid difference accounts for the diminished arsonate binding by the 1210.7 hybridoma protein. Because the 1210.7 heavy chain is the unmutated product of the BALB/c VH gene, somatic mutation in VH may be required to enhance Ars affinity in this system.     

Detection of private and recurrent idiotopes on natural anti-tubulin antibodies by monoclonal anti-idiotopic antibodies

Two monoclonal anti-idiotopic antibodies (anti-Id) were raised in mice against a human monoclonal IgA,K displaying a monospecific anti-tubulin (anti-alpha- and anti-beta-tubulin) activity. One anti-Id (IgG,K) recognized a private idiotope, TID 3.2, present only on the IgA,K immunogen, close to or within the antigen-combining site. The other anti-Id (IgM,K) recognized a recurrent idiotope, TID 7.1, outside the paratope and present in normal human and BALB/c mouse serum, on 2 of 11 polyspecific human monoclonal immunoglobulins and on 6 of 11 murine natural monoclonal auto-antibodies exhibiting a widespread anticytoskeletal protein-binding activity. Both the idiotopes were absent on two induced anti-tubulin antibodies exhibiting a monospecific anti-alpha- and anti-beta-tubulin specificity. Utilizing competitive and additivity immunoassays, we could show that the polyspecific human and mouse anticytoskeletal antibodies tested, whether bearing the TID 7.1 Id or not, appeared to compete in variable degrees for epitopes on the tubulin molecule recognized by the monoclonal IgA,K but distinct from the epitopes recognized by the induced monospecific anti-tubulin antibodies. The high incidence of the recurrent TID 7.1 idiotope in man and mouse suggests an important physiologic and perhaps regulatory function of this idiotope. Furthermore our data suggest that a restricted family of germ-line genes, highly conserved during phylogeny, may encode for these idiotope-bearing Ig molecules.     

Immune response to human immunodeficiency virus. In vivo administration of anti-idiotype induces an anti-gp160 response specific for a synthetic peptide

Anti-idiotypic antibodies (anti-Id) to chimpanzee antibodies directed against a synthetic peptide corresponding to a native epitope associated with gp41 of human immunodeficiency virus (HIV) envelope glycoprotein were produced in rabbits. The peptide was analogous to amino acid sequences 735 to 752 from the human T cell leukemia virus-IIIB isolate of HIV. Characteristics of the anti-Id preparation included: 1) detection of a shared determinant present on a second chimpanzee and one of three rabbit antibody preparations directed against the synthetic peptide, 2) failure to recognize an idiotype (Id) in BALB/c mouse antisera to the peptide, and 3) partial inhibition of the homologous chimpanzee Id preparation from binding either peptide or a recombinant HIV gp160 preparation. Immunization of BALB/c mice with the anti-Id induced an antipeptide response which bound a recombinant gp160 preparation without subsequent peptide or gp160 exposure. The anti-gp160 containing sera from mice immunized with anti-Id were able to inhibit the Id-anti-Id reaction indicating that an Id-positive antibody response was induced. This Id is not normally expressed in the murine anti-gp 160 immune response to the synthetic peptide and suggests that this anti-Id may activate normally silent clones. This study indicates that Id networks may be operational during the immune response to HIV epitopes. Alternatively, anti-Id may be useful in altering the serologic characteristics of an antibody response to HIV and may offer potential for modulating the immune response in this viral infection.     

Characteristics of murine monoclonal anti-CD4. Epitope recognition, idiotype expression, and variable region gene sequence.

We have characterized a series of mouse monoclonal anti-CD4 and describe both their CD4 epitope recognition and Id expression. We also determined the V region gene sequences of these antibodies in an attempt to correlate epitope recognition and Id expression with V region sequence. All of these preparations recognize epitopes that cluster around the HIV gp120 binding site on the human CD4 molecule. However, we observed differences in epitope recognition among the anti-CD4 preparations, based on either competitive inhibition assays or functional assays, such as syncytium inhibition. Analysis of Id specificities using a polyclonal anti-Id generated against anti-Leu 3a indicated that five of the seven monoclonal anti-CD4 expressed a shared Id. Based on V region gene sequences, the V region kappa-chain (V[kappa]) from each of the seven antibodies was encoded by the V[kappa]21 gene family and expressed the J[kappa]4 gene segment. Those preparations that expressed the shared Id with anti-Leu 3a have virtually identical V[kappa] sequences, with a high degree of homology in the CDR. The VH region gene sequences of six of the seven antibodies also shared overall homology and appeared to be encoded by the J558 VH gene family. The seventh anti-CD4 VH region is encoded for by the VHGAM gene family. The majority of these antibodies used JH3 gene segment, although the JH2 and JH4 gene segments were also represented. In addition, several of these antibodies share a common sequence organization within their V-D-J joining regions that appears to involve N and P sequences to generate unique D segments. Together, these data suggest that differences in epitope recognition among the monoclonal anti-CD4 may reflect sequence variability primarily within the CDR3 region of both V[kappa] and VH. The basis for the detection of a shared Id most likely reflects the high degree of homology within the V[kappa] region sequences. In addition, these data, which are based on a limited analysis, suggest the possible restricted use of V region germ-line gene families in the secondary antibody response of BALB/c mice to specific epitopes on the human CD4 molecule.     

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.     

Anti-idiotype-induced, lipopolysaccharide-specific antibody response to Pseudomonas aeruginosa. II. Isotype and functional activity of the anti-idiotype-induced antibodies

We recently developed a murine anti-idiotypic mAb that functioned as a molecular mimic of the O-specific polysaccharide side chain (Ps) of Pseudomonas aeruginosa LPS in vitro, and which induced Ps-specific antibodies in syngeneic BALB/c ByJ mice. In the current studies, we demonstrate that these anti-Id-induced, Ps-specific antibodies fix complement to the bacterial cell surface, and protect neutropenic mice from fatal P. aeruginosa sepsis. The isotypic distribution of the anti-Id-induced antibodies, however, resembles the restricted pattern (IgM and IgG3) seen after administration of purified Ps to mice. The immunogenicity and number of isotypes of Ps-specific antibody produced could be enhanced by conjugating the anti-Id to keyhole limpet hemocyanin. Finally, the anti-Id administered before immunization with purified Ps, primed BALB/c ByJ mice for production of other Ps-specific antibody isotypes (IgG1). These studies show that this anti-Id induces functional anti-Ps antibodies in syngeneic mice, and when used in conjugate form or as a priming agent before Ps immunization, yields an antibody response consistent with "T cell dependence." These immunization strategies may be useful for the induction of polysaccharide-specific antibodies in man.     

Expression of a cross-reactive idiotope on naturally occurring murine antibodies against 3-fucosyllactosamine, levan, and dextran

We recently identified a cross-reactive Id (6C4) that is expressed on the H chain of many BALB/c mAb against the 3-fucosyllactosamine (3-FL) determinant, Gal(beta 1-4) (Fuc(alpha 1-3] GlcNAc-R. The VH segments of seven mAb that we recently sequenced are encoded by VH441, which also encodes VH segments of antibodies against galactan, levan, and dextran. To analyze the expression of the 6C4 Id on naturally occurring anti-carbohydrate antibodies, we isolated 6C4+ antibodies by affinity chromatography from pools of normal BALB/c serum. Approximately 20 to 30% of antibodies against 3-FL and levan, and all antibodies against dextran, were removed from the sera by passage over a column containing mAb 6C4. Absorption of the eluate with 3-FL beads removed anti-3-FL antibodies but not anti-dextran or anti-levan. The expression of a cross-reactive Id on naturally occurring antibodies against several carbohydrate Ag suggests that these antibodies may participate in an Id network. We also reported previously that BALB/c mice have naturally occurring anti-3-FL antibodies and respond well to immunization against this determinant, whereas C57BL/6 mice do neither. To examine the role of the Igh-C allotype in the regulation of the anti-3-FL response, we studied congenic strains of BALB/c and C57BL/6 mice. Both congenic strains produced anti-3-FL antibodies in response to immunization, but only C.B-20 mice exhibited naturally occurring antibodies. These data suggest that the naturally occurring and elicited antibody responses against 3-FL are differentially regulated.     

Induction of an anti-hepatitis B surface antigen response in mice by noninternal image (Ab2 alpha) anti-idiotypic antibodies

Anti-idiotype antibodies to a mouse monoclonal antibody A-12 directed against HBsAg were produced in rabbits. The anti-Id consisted of an Ab-2 alpha preparation that did not display any detectable internal image activity. Immunization of BALB/c mice with the anti-Id (Ab-2 alpha) coupled to KLH induced an anti-HBs response without subsequent HBsAg exposure. No anti-HBs was detected in control groups of mice immunized with other rabbit anti-Id-KLH preparations. The anti-HBs containing sera from mice immunized with the Ab-2 alpha were able to inhibit the Id-anti-Id reaction, indicating that an Id-positive, anti-HBs response was induced. This idiotype is not normally expressed during the murine immune response to HBsAg and suggests that noninternal image anti-Id activates silent clones. This study, along with our previous results obtained with the use of internal image anti-Id, suggests that there is more than one Id network operational during the BALB/c murine immune response to HBsAg.