Idiotype Expression Research Articles

Influence of paternal immunity on idiotype expression of offspring.

The immune response of the rat to group A streptococcal carbohydrate (SACHO) and an associated idiotype, Id-1, was used to examine the effect of paternal immunity on Id-1 and SACHO-specific antibody expression by the offspring. First litters, conceived before immunization of the father, had significantly higher Id-1 levels than litters conceived by the same parental pairs after hyperimmunization of the father (P greater than 0.01). Total anti-SACHO levels were not affected. The effect appeared to be independent of the level of Id-1 expressed by the father or grandfather. No significant difference in Id-1 production was found between offspring of actively immune, neonatally Id-1 suppressed fathers and fathers expressing high levels of Id-1. We suggest that the paternal immunoregulatory influence acts via the maternal immune system to modify the idiotype repertoire expressed in the immune response of the offspring, and is not the result of genetic transmission of a trait acquired by the father. Some possible mechanisms of transmission are discussed.

Idiotypic manipulations: Hierarchy in idiotype expression in rabbits immunized against Micrococcus luteus

Idiotypic cross-reactions were analyzed among three series of anti-peptidoglycan antibodies of the Micrococcus luteus system. The reference idiotype Abl was an antibody fraction isolated from an isoelectric focusing preparative column. Cross-reactive idiotypes, Abl', were induced through the immunization chain (Abl-Ab2-Ab3). Idiotypic antibodies ofAbl-Fl type were obtained from offspring of female rabbits, actively producing Ab3 during pregnancy. Finally, Abl CRI were cross-reactive idiotypes with Abl found in a random population of rabbits immunized with M. luteus. Three idiotopes could be characterized within Abl antibody. Abl' usually expressed two of these idiotopes, but never the third specificity which is “private” to Abl. Abl-Fl shared one or two idiotopes with Abl and Abl' antibodies. Only one common idiotope appeared to be present on Abl CRI. Finally, this idiotope, IdX, could be detected by radioimmunoassay in 20% of rabbits immunized with micrococcal vaccine. It appears that a recurrent idiotype of anti-peptidoglycan antibodies can be preferentially amplified through idiotypic manipulations. On the other side, cascade immunizations lead to the expression on Abl' and on some Abl-Fl of a second idiotypic specificity, shared with Abl. This hierarchy of idiotype expression may well be important in the regulation of antibody synthesis through idiotypes.

A major anti-myoglobin idiotype. Influence of H-2-linked Ir genes on idiotype expression.

A rabbit antiidiotypic antiserum raised against an A.SW IgG1K monoclonal anti-sperm whale myoglobin (Mb) antibody, HAL19, and extensively absorbed with normal mouse immunoglobulin and MOPC 21 (IgG1K), was found to detect a common or major anti-Mb idiotype expressed by some but not all anti-Mb monoclonal antibodies, regardless of immunoglobulin G (IgG) subclass, and by 40-50% of the anti-Mb antibodies in immune serum from five high responder strains of mice representing five different Igh allotypes. It did not inhibit antibodies to three unrelated protein antigens. The fraction of antibodies expressing this idiotype, denoted IdHAL19, was regulated by H-2-linked genes that correlated exactly in four independent haplotypes and an F1 with the known Mb immune response (Ir) genes and may be identical to these. Whereas less than 50% of antibodies from high responder mice were inhibitable by anti-IdHAL19, greater than 80% of antibodies from low responder mice, tested at comparable final antibody concentration, were inhibitable. This result was true for both low responder haplotypes, H-2b (B10) and H-2k (B10.BR). The idiotype was found to be present on antibodies that bound to native Mb but not fragments 1-55 or 132-153 of Mb or a denatured form, S-methyl Mb. This specificity for native Mb paralleled that of the monoclonal idiotype HAL19 itself. Therefore, the production of antibodies specific for native in contrast to denatured Mb was studied in H-2-congenic high and low responder strains. Strikingly, low responders produced antibodies that reacted almost exclusively with the native conformation, whereas a larger proportion of antibodies from high responder mice also reacted with the denatured form, S-methyl Mb. Bypassing of the Ir gene defect by immunization with Mb attached to a carrier, F gamma G, resulted in low responder antisera resembling higher responder sera in both idiotype expression and conformational specificity. The simplest explanation of these results is that H-2-linked Ir genes control antibody fine specificity, which is reflected in the idiotypes of the variable regions expressed. We suggest that low responder mice produce a more limited repertoire of antibodies consisting primarily of IdHAL19-positive antibodies specific for the native conformation of Mb. High responder mice produce a greater diversity of antibodies to Mb, so that the IdHAL19-positive, conformation-specific population represents a smaller proportion of the total. Similarly, the use of carrier-specific helper T cells in low responder mice results in a greater diversity of antibodies, which dilutes out the IdHAL19 subset.(ABSTRACT TRUNCATED AT 400 WORDS)

Analysis of nondominant idiotypes expressed during alloimmune responses

The antibody response of Lewis rats (RT1.A 1) to class I MHC antigens of the Brown Norway rat (RT1.A n) was studied. Diversity of the serum alloimmune response was analyzed using syngeneic anti-idiotype raised against monoclonal antibodies of the same specificity. Cross-reactive idiotypes were detected on approximately one in one thousand Lewis anti-RT1.A n serum antibodies, at concentrations ranging from 20 to 600 ng/ml. The kinetics of idiotype expression coincided with that of total anti-BN antibody production, suggesting that both were regulated by the same mechanism. To determine whether humoral anti-idiotype was involved in such regulation, sera from these animals were screened for anti-idiotype content. Using an RIA sensitive to 20 ng/ml, no humoral anti-idiotype could be detected during any phase of the alloimmune response.

Internal complementarities in the immune system: regulation of the expression of helper T-cell idiotypes.

More than half of BALB/c helper T lymphocytes specific for 2,4,6-trinitrophenyl (TNP)-modified syngeneic spleen cells are inhibited in their proliferative responses to antigen-presenting cells and in their cooperation with B lymphocytes by monoclonal anti-idiotypic antibodies directed to a TNP-binding BALB/c myeloma protein (MOPC 460). This inhibition is specific for anti-TNP-self helper cells of BALB/c origin and is controlled by IgCh-linked genes, as it is not observed with CB.20 helper cells of the same specificity. In contrast, anti-TNP-self helper cells prepared from BALB/c mice that were chronically suppressed with anti-mu chain antibodies and possessed no B lymphocytes were not inhibited by anti-idiotypic antibodies. We conclude that the B-cell antibody repertoires contribute to the selection of the (idiotypic) T-helper-cell repertoires.

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.

Use of JH4 joining segment gene by an anti-arsonate antibody that bears the major A-strain cross-reactive idiotype but displays diminished antigen binding.

One of the antibody families utilized by the A/J mouse in its response to p-azophenylarsonate (Ars) is characterized by the expression of the major anti-arsonate cross-reactive idiotype (CRI) of the A strain. This family has been termed the Ars-A family. A hybridoma antibody (HP 101F11 ) obtained after immunization of an A/J mouse with Ars was identified initially as displaying the CRI, but was subsequently found to bind antigen at a level much lower than most members of the Ars-A family. The results of binding studies suggested that HP 101F11 possesses reduced avidity for antigen. When isolated light and heavy chains were allowed to recombine with the heavy and light chains of a strongly antigen-binding, strongly CRI-positive antibody of the Ars-A family (HP 93G7 ), the low level of antigen binding by HP 101F11 was found to be due to a structurally variant heavy chain. Whereas antibodies of the Ars-A family with normal avidity for antigen had been shown to use the JH2 joining segment gene, amino acid sequence analysis of HP 101F11 revealed that this antibody has a JH segment with a sequence identical to that encoded by a portion of a different JH gene, JH4 . The implication that 101F11 uses the JH4 gene instead of JH2 was supported by the observation that the productively rearranged gene is associated with an Eco R1 restriction fragment 0.95 Kb smaller than the corresponding fragments of Ars-A hybridomas with normal avidity for antigen. The size difference of 0.95 Kb corresponds exactly to the known distance between the JH2 and JH4 genes in BALB/c germline DNA. In addition to the structural differences immediately attributable to the use of JH4 , HP 101F11 has shown an amino acid interchange in the DH segment, and a single amino acid deletion at the DH-JH boundary. These results show that variation among members of the Ars-A family in the DH and/or JH segments provides alternative structural forms of Ars-A antibodies upon which selective processes can operate during the course of an immune response.

Expression and regulation of two idiotype families and subsets within an idiotype family among BALB/c antibodies against p-azophenylarsonate.

The expression and regulation of the two different idiotype (id) families associated with the anti-p-azophenylarsonate (Ar) antibodies of BALB/c mice were examined. Both families ( 5AF6 and 3C6 ) represented cross-reactive idiotypes (CRI) expressed in the anti-Ar of most individual BALB/c mice. In response to keyhole limpet hemocyanin-Ar, an average of about 28% of BALB/c anti-Ar had 5AF6 family idiotopes, while 3C6 family was expressed on about 16% of BALB/c anti-Ar antibodies. Suppression induced by anti-idiotype treatment against one family did not suppress the expression of the other family, suggesting that the two families were regulated independently. However, the relative expression of one family could influence the expression of the other, because depression of the 5AF6 family tended to increase the expression of the 3C6 family of anti-Ar. Analysis of the 5AF6 family showed that a majority of BALB/c mice produced antibodies bearing most or all of the idiotopes associated with the family, but that a subset of about 35% of the antibodies synthesized lacked idiotopes associated with a monoclonal anti-Ar member of this family, 2.4. Treatment of mice with anti-idiotypes prepared against two different monoclonal anti-Ar of the 5AF6 family produced different effects: one enhanced while the other suppressed idiotype expression, suggesting that there are differences in the idiotopes associated with these two regulatory pathways. Additionally, results indicated that subsets of antibodies within the 5AF6 idiotype family could be regulated independently of each other.

Tetraparental rabbits chimeric for their lymphoid system—II. Idiotype expression

In order to test some aspects of the network theory of “idiotype-anti-idiotype” interactions, we studied the idiotypic specificities of anti-DNP antibodies produced by tetraparental rabbits, chimeric for their lymphoid system, under two different experimental conditions. In the first, the tetraparental rabbits were simply immunized against the hapten and we observed that the antibodies, totally different in their allotypy, produced by the lymphocytes of the two constitutive parental strains remained idiotypieally independent in three cases out of three. In the second experimental regimen, the tetraparental rabbits underwent an anti-anti-idiotypic immunization before the anti-hapten one. In two out of the four cases studied we observed that anti-DNP antibodies different in their a- and b-series allotypic specificities can bear at least related idiotypic specificities.

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

Using hapten-reversible inhibition of plaque formation as an assay for auto-anti-idiotype antibody (anti-Id) and as a means for following idiotype (Id) expression, we have obtained evidence that following immunization with trinitrophenyl (TNP) conjugates (a) there are differences in Id expression in the anti-TNP antibody response to different TNP conjugates although there is some overlap; (b) different strains, although showing some differences in Id expression, tend to produce cross-reactive Ids, thus no obvious allotype linked inheritance of Id expression is observed in this heterogeneous immune response; (c) the auto-anti-Id produced following immunization with TNP- Brucella abortus or TNP-Ficoll tends to be of the IgG 2a and IgG 2b isotypes.

Idiotypic cross-reaction between MRL monoclonal autoantibodies with different antigen specificity

The idiotypic determinants borne by Y2, a monocional anti-Sm antibody of MRL mouse strain origin, were investigated to elucidate mechanisms for the generation of autoantibodies. Using an ELISA assay, a rabbit anti-Y2 anti-idiotypic antiserum was tested for binding to other MRL autoantibodies to identify cross-reactive determinants. In this way, a shared idiotype was demonstrated for Y2 and 4K1, 4K1 is an IgG2bK monoclonal antibody derived from a fusion with spleen cells of an MRL- lpr lpr mouse expressing anti-Sm. 4K1 displayed an antigenic specificity distinct from anti-Sm, however, with reactivity by fluorescent antinuclear antibody analysis to a subcellular component localized to the perinuclear region. The expression of the Y2-4K1 idiotype was assessed using a sensitive inhibition ELISA. The Y2-4K1 determinant was present in sera of MRL- lpr lpr mice irrespective of the presence of anti-Sm. Sera of some but not all normal mouse strains also demonstrated this determinant. These results indicate that while some idiotypes may be associated with autoantibodies, they are not restricted to the autoimmune setting. This pattern of expression suggests that autoantibodies are part of larger idiotype-bearing antibody families with structural similarity to antibodies produced by normal mice. The autoimmune setting may promote the emergence of idiotype-bearing molecules with autoantibody activity and aberrant regulation.

Immunologic memory to phosphocholine. V. Hybridomas representative of group II antibodies utilize V kappa 1-3 gene(s).

The anti-phosphocholine (PC) memory response of BALB/c mice to PC-KLH contains two groups of antibodies distinguished by fine specificity and by expression of the T15 idiotype that dominates Group I but not Group II anti-PC antibodies. The contribution of V kappa genes to this diversity was investigated by the analysis of L chains from PC-binding hybridoma proteins (PCBHP) representative of Group I and Group II. N-terminal amino acid sequence analysis was performed on the L chains of three independently derived Group II PCBHP up to residue 23 (PCG1-1) or 21 (aPC-111-1 and aPC-12-3). These three sequences differed from each other by only one or two residues, but differed by approximately 50% from the L chains of the Group I-like PC-binding myeloma proteins (PCBMP); the Group II sequences are closely related to V kappa 1-3. Isoelectric focusing analysis was also performed on the L chain of PCG1-1, as well as on L chains from PCBHP typical of Group I antibodies, and from an atypical PCBHP differing from Groups I and II in fine specificity. A Group I PCBHP and the atypical PCBHP expressed L chains related to V kappa 8 and V kappa 24, respectively. The L chains of another Group I PCBHP and of the Group II protein, PCG1-1, appeared different from those found in the PCBMP and from each other. The results indicate a more diverse expression of L chains in the memory anti-PC response than is represented by the PCBMP; both V kappa 8- and V kappa 24-derived L chains (and, presumably, somatic variants), as well as products of additional V kappa genes (V kappa 1-3), appear to be present in the anti-PC memory pool.

Rheumatoid factors in 129XB recombinant inbred strains. Igh-1-linked control of allotypic and isotypic specificities.

To examine the role of autologous IgG in the induction of murine rheumatoid factors (RF) we have analyzed the allotypic specificity of anti-IgG2a RF in recombinant inbred strains derived from 129/Sv (Igh-1a) and C57BL/6 (Igh-1b) mice. In five of six Igh-1a strains, anti-IgG2a RF reacted with IgG2aa but failed to react with IgG2ab. In contrast, isotype-specific RF, which reacted equally well with a and b allotypes of IgG2a, represented the major RF species in one Igh-1a and all five Igh-1b strains tested. An additional form of RF specific for IgG2ab and not reactive with IgG2aa was detected in one Igh-1b strain. RF specific for a give allotype was thus only found in the presence of that allotype, which strongly suggests the involvement of autologous IgG in the induction of mouse RF synthesis. The specificity of RF was apparently further controlled by genes linked to but different from the Igh-C locus, as indicated by the absence of IgG2aa-specific RF in one of the 6 Igh-1a strains tested. Because this strain, 129XBG, has been shown to express idiotypic markers characteristic of Igh-1b mice, it is likely that the genes, which in the presence of a given allotype induce the production of isotype rather than allotype-specific RF, are identical to those that control the expression of idiotypes. Evidence was also obtained to indicate that Igh-1-linked genes influence the isotypic specificity and the isotype of RF itself: IgA anti-IgG2a predominated in Igh-1a strains and IgM anti-IgG1 in Igh-1b strains. Interestingly enough, total IgA and IgG2a levels also were higher in Igh-1a than in Igh-1b strains.

The D segment defines the T15 idiotype: the immunoresponse of A/J mice to Pneumococcus pneumoniae.

In the immune response of BALB/c mice (Igha) to Pneumococcus the majority of antibodies express the idiotype of the myeloma protein TEPC 15 (T15). In contrast mice of the A/J strain (Ighe) do not express this idiotype. Using (BALB/c X A/J)F1, F2 or backcross mice it could be shown that in allotype heterozygous animals (Igha/e) Pneumococcus pneumoniae preferentially stimulates B cells expressing a heavy chain (H) encoded by genes in the BALB/c H chain gene complex. Phosphorylcholine (PC)-specific hybridoma lines were established from BALB/c and A/J spleen cells and idiotypically analyzed using monoclonal antibodies (mAb) specific for the T15 idiotopes 32/65, 10/13, 16/13 or 21A5. Whereas the majority of the BALB/c PC-binding mAb express these idiotopes, only some of the A/J mAb are positive for one or the other of the idiotopes formed by the variable (V) regions of the H and the light chain of the myeloma protein T15. However, 80% of the A/J PC-binding hybridoma proteins were bound by the anti-idiotopic mAb 21A5. This mAb is specific for a determinant partially formed by the C alpha and partially by the V regions of the myeloma protein T15. The mRNA of one of these T15- A/J PC-binding hybridoma lines was sequenced. VH and V kappa were identical with sequences found for BALB/c T15-like antibodies. The sequence of the D segment was structurally very different. The importance of the D segment in the dominant expression of the T15 idiotype is discussed.

Analysis of the anti-azobenzenearsonate response at the molecular level

We have analysed, at the molecular level, the antibodies associated with the anti-azobenzenearsonate response of A/J and BALB/c mice as well as the genes that encode them. All antibodies expressing the predominant idiotype are derived by somatic mutation from a single VH gene. Antibody molecules express the idiotype regardless of their light chains or the JH segments which encode them. The D segment, however, does seem to be important for idiotype expression. The anti-azobenzenearsonate antibodies possessing the minor idiotype (36-60) are encoded by single, highly homologous VH genes in A/J and BALB/c mice. The nucleotide sequences of the VH genes from these two strains show only two nucleotide disparities. However, as opposed to A/J mice, the 36-60 idiotype in BALB/c appears to be the dominant idiotype among anti-azobenzenearsonate antibodies.

Idiotypic analysis of monoclonal anti-fluorescyl antibodies: Localization and characterization of idiotypic determinants

Nine monoclonal IgG anti-fluorescyl antibodies, which exhibit diverse affinities for fluorescein (Fl) ( K a values ranging from 5 × 10 6 to 10 10 M −) were analyzed idiotypically. Each of the BALB/c hybridoma proteins (γ, κ) exhibited unique idiotypic determinants although two clones (6-10-6 and 20-19-1) were partially (15–20%) cross-reactive. Of two other clones (4-6-9 and 4-6-10) derived from the same cell line, 4-6-9 contained γl heavy (H) chains and 4-6-10 contained both γl and γ2b H-chains. In addition, 4-6-9 shared idiotypic determinants with 4-6-10 although the latter also displayed unique idiotypic specificities. Collectively, the nine clones demonstrated structural diversity analogous to previous studies which denned binding mechanism diversity. The location of determinants recognized by antiidiotype reagents directed against each of the monoclonal antibodies was examined by binding inhibition with free Fl and fluorescein-BSA (Fl-BSA). All clones contained determinants both within the active site (Fl-inhibitable) and in close proximity to it (Fl-BSA-inhibitable), although the relative proportions of these determinants varied among the clones. Inhibitor concns required for 50% inhibition varied independently of ligand binding affinity, and therefore were more likely influenced by the heterogeneous nature and affinity of the anti-idiotype reagents toward the individual determinants. Idiotypic analysis of H- and light (L) chains derived from five monoclonal antibodies of diverse affinities was performed. Fl binding and expression of idiotypic determinants by all clones required both H- and L-chains. Restoration of the idiotye by reassociated H- and L-chains was found to be highly restricted to homologous H- and L-chain pairs, as heterologous combinations did not result in the expression of either parental idiotype. The latter was true whether the heterologous pairs were derived from clones of the same isotype or the heterologous combination associated to form an intact molecule with greater affinity than the parental H- and L-chain combination. Heterologous recombinants from the two clones (6-10-6 and 20-19-1) exhibiting partial idiotypic cross-reactivity were able to restore a fraction (~25%) of their idiotypic determinants. Results demonstrated the extensive conformational requirements of ligand binding and idiotype expression and indicated that a high degree of specificity in the V H- and V L-chain interaction must exist for the expression of these idiotypes.

The role of B-cell I region encoded antigens in T-cell dependent B-cell activation: I region encoded antigen density correlates with idiotype expression.

The role of B-cell I region encoded antigens in T-cell dependent B-cell activation: I region encoded antigen density correlates with idiotype expression.

Idiotypes of anti-major histocompatibility complex antibodies.

Our studies to date indicate that treatment with anti-idiotype to monoclonal anti-MHC antibodies can markedly influence the repertoire of anti-MHC antibodies expressed. The antibodies discussed here appear to represent two classes, one of which is public, probably representing expression of a germ line gene, and the second of which probably represents either a somatic variant of a germ line gene or one of a very large number of germ line genes devoted to the same specificity. In either case, this class of idiotype arises only rarely following antigen, but is readily induced by anti-idiotype treatment. There may indeed exist a third class of anti-MHC monoclonal antibodies representing distant somatic diversification from a germ line gene. Our only indication of this so far is that certain idiotypes are only induced after multiple boosts with anti-idiotypes rather than a single treatment. This finding, however, may reflect again the enormous number of different ways in which anti-MHC antibodies to the same nominal specificity can be produced. Finally, our results in adoptive transfer systems indicate that manipulation of idiotype expression by anti-idiotype treatment probably involves a complex pathway of cellular interactions. If, as we expect, these intercellular interactions involve idiotype and/or anti-idiotypic receptors, they should provide a model for mechanistic studies of the in vivo immune network.

Cellular and molecular mechanisms that regulate idiotype expression in myeloma cells.

Cellular and molecular mechanisms that regulate idiotype expression in myeloma cells.

Galactan-binding antibodies. Diversity and structure of idiotypes.

A group of eight IgM hybridoma proteins induced with beta(1,6)-D-galactan-containing antigens has been characterized in terms of primary amino acid sequence and idiotype expression. The H chain amino acid sequences reveal very strong homology in the VH segment although several substitutions are seen that suggest the occurrence of somatic mutation in these IgM molecules. Significant sequence variation was observed in CDR-3, the region generated by the D segment, and the two recombination events, VH-D and D-JH. The number of amino acids in this region contributed by the D segment was found to vary from two to six, yet the overall length of CDR-3 was precisely maintained by the addition of amino acids on either side of D during the recombination processes. These additional amino acids are suggested to result from nucleotide addition by repair enzymes. Idiotypic analysis of these proteins, in conjunction with an assessment of the H chain sequences, has permitted an identification of the molecular basis of both cross-reacting and unique idiotypic determinants expressed by these molecules.