Antigenic determinants of adenovirus capsids. II. Homogeneity of hexons, and accessibility of their determinants, in the virion.

We have tested the two principal theories which explain the previous finding that small amounts of type-specific antibody to the adenovirus hexon can neutralize infectivity, whereas even large amounts of cross-reactive antibody do not. a) It has been suggested that the type-specific determinants are especially prominent in the virion. We have therefore measured the capacity of whole virus to bind appropriate antibodies, using a sensitive radioimmunoprecipitation (RIP) system. In fact, virions bound type-specific and cross-reactive antibodies impartially. Moreover, they bound both much less effectively than did free hexon or disrupted virus, suggesting that many of each kind of determinant are inaccessible in virions. b) It has been suggested that the type-specific determinants are confined to those hexons located next to the pentons, and that they are the targets for neutralizing antibody. We have therefore studied the antigenicity of peripentonal and nonamer hexons isolated from virions, and found that each possessed both kinds of determinants. Furthermore, these were present in the same proportion as in hexons purified from the soluble antigens in infected cells ("free hexons"). We concluded that the mechanism of neutralization by antibody is complicated, and that the type-specific determinants exposed on the virion must play a crucial role.     

Immunoenzyme analysis of adenovirus hexons. Indication and characteristics of rabbit and mouse antibodies against hexons

An enzyme-linked immunosorbent assay (ELISA) was developed for analysis of rabbit and mouse IgG antibodies specific to adenoviral hexon. The anti-hexon antibodies were detected by capture with purified hexon coated onto polystyrene microtiter plates and visualizing them by respective anti-IgG horseradish peroxidase conjugates. In the sera from hyperimmunized rabbits and mice as well as in the mouse ascite fluids the ELISA procedure revealed primarily type-specific (epsilon) and genus-specific (alpha) antigenic determinants in hexon but not those of intermediate specificities.     

Antigenic composition of adenovirus hexons]

Quantitative relations between the group-specific and the type-specific components of the hexons of adenovirus type 2 and 5 were studied by means of FITC-conjugated Fab-fragments of antibodies directed against type 2 and type 5 hexons. From the sedimentation constant of the complexes of hexons and Fab in the region of excess of Fab we conclude that there are at least 20 determinants on the hexon. Half of these are type-specific and the others are group-specific. Both components of the type 2 hexon consist of equal parts of carbodiimide sensitive and carbodiimide resistent determinants.     

Antibodies to envelope glycoprotein of dengue virus during the natural course of infection are predominantly cross-reactive and recognize epitopes containing highly conserved residues at the fusion loop of domain II

The antibody response to the envelope (E) glycoprotein of dengue virus (DENV) is known to play a critical role in both protection from and enhancement of disease, especially after primary infection. However, the relative amounts of homologous and heterologous anti-E antibodies and their epitopes remain unclear. In this study, we examined the antibody responses to E protein as well as to precursor membrane (PrM), capsid, and nonstructural protein 1 (NS1) of four serotypes of DENV by Western blot analysis of DENV serotype 2-infected patients with different disease severity and immune status during an outbreak in southern Taiwan in 2002. Based on the early-convalescent-phase sera tested, the rates of antibody responses to PrM and NS1 proteins were significantly higher in patients with secondary infection than in those with primary infection. A blocking experiment and neutralization assay showed that more than 90% of anti-E antibodies after primary infection were cross-reactive and nonneutralizing against heterologous serotypes and that only a minor proportion were type specific, which may account for the type-specific neutralization activity. Moreover, the E-binding activity in sera of 10 patients with primary infection was greatly reduced by amino acid replacements of three fusion loop residues, tryptophan at position 101, leucine at position 107, and phenylalanine at position 108, but not by replacements of those outside the fusion loop of domain II, suggesting that the predominantly cross-reactive anti-E antibodies recognized epitopes involving the highly conserved residues at the fusion loop of domain II. These findings have implications for our understanding of the pathogenesis of dengue and for the future design of subunit vaccine against DENV as well.     

Stability of the structure and antigenic determinants of adenovirus type 1 native hexon to proteases

Hexon capsomers of human adenovirus type 1 (h1) labeled by iodine 125 were digested in a native state (trimers) by trypsin, chymotrypsin or papain, and the resulting hydrolysates were analyzed by SDS-PAGE. In each case, a discrete and temporally stable pattern of relatively large fragments was revealed. The degree of hexon polypeptide hydrolysis was maximal for papain, intermediate for chymotrypsin and minimal for trypsin, the largest fragments in the digest being 32, 40 and 80 kD, respectively. At room temperature, all the electrophoretically discernible hexon proteolytical fragments were held together in structures resembling intact hexon trimers and could be regarded as "hexon cores", of which papain hexon cores were the most stable during SDS-PAGE. Radioimmunoprecipitation analysis revealed a complete absence of native hexon antigenicity in thermodenaturated fragments of hexon protease digests, while native trypsin, chymotrypsin and papain hexon cores could be precipitated by hexon-specific antibodies. The immunoprecipitated material contained all of the hexon fragments found in appropriate hexon cores and retained the structure of the original cores. Trypsin, chymotrypsin and papain hexon cores were shown to possess at least part of native Ad h1 hexon antigenic determinants of each of the following specificities: species-specific (epsilon), cross-reactive with hexon of human adenoviruses (h3 and h6), simian adenovirus (sim 16), bovine adenoviruses (bos 3 and bos 7) and avian adenovirus (Aviadenovirus gal 1 or CELO). Thus, the full spectrum of known hexon antigenic determinants (species-specific to intergenus-crossreactive) is at least portly stable against protease attack of native hexon capsomers.     

Antibodies against the two serotypes of vesicular stomatitis virus measured by enzyme-linked immunosorbent assay: immunodominance of serotype-specific determinants and induction of asymmetrically cross-reactive antibodies.

The serological relationship between the two vesicular stomatitis virus (VSV) strains Indiana (VSV-Ind) and New Jersey (VSV-NJ) were analyzed by using an enzyme-linked immunosorbent assay (ELISA). Immunoglobulin G responses, defined by their resistance to treatment with 2-mercaptoethanol, were assessed by ELISA by using sucrose gradient-purified VSV or purified VSV glycoproteins (G) as antigens. When low doses (10(6) PFU) of live VSV or 10(8) PFU of UV-inactivated virus were given intraperitoneally (i.p.), only non-cross-reactive antibody responses were observed in a primary immune response. However, when 10(6) PFU of live VSV were injected intravenously (i.v.), cross-reactive antibodies were generated; anti-VSV-NJ antibodies cross-reacted more against VSV-Ind than did anti-VSV-Ind antibodies against VSV-NJ. When 10(8) PFU of live VSV or UV-inactivated VSV mixed with complete Freund adjuvant was given i.p., high levels of cross-reactive antibodies detectable by ELISA were induced in primary and secondary responses. When purified G protein was used instead of purified whole virus in the ELISA, the cross-reactivity was found to be asymmetrical after immunization with live VSV given i.v. but not after i.p. inoculation; anti-VSV-NJ sera bound almost equally well to VSV-Ind G protein, whereas anti-VSV-Ind sera bound virtually exclusively to the G protein of the homologous serotype. The data suggest that immunization with VSV given i.p. results in a more specific, i.e., less cross-reactive, response than that either after i.v. infection or after the virus antigen is made available in great amounts or if it persists for prolonged periods when given i.p. together with complete Freund adjuvant. The unique determinants were immunodominant because they induced antibodies preferentially, whereas partially shared determinants induced antibody responses asymmetrically, more slowly, and with lower titers. Interestingly, the asymmetric cross-reactivity of anti-VSV antibodies, as measured by ELISA, against purified VSV G was opposite that observed for cytotoxic T cells.     

Analysis of 15 adenovirus hexon proteins reveals the location and structure of seven hypervariable regions containing serotype-specific residues.

The first full-length hexon protein DNA and deduced amino acid sequences of a subgenus D adenovirus (AV) were determined from candidate AV48 (85-0844). Comprehensive comparison of this sequence with hexon protein sequences from human subgenera A, B, C, D, F, bovine AV3, and mouse AV1 revealed seven discrete hypervariable regions (HVRs) among the 250 variable residues in loops 1 and 2. These regions differed in length between serotypes, from 2 to 38 residues, and contained > 00% of hexon serotype-specific residues among human serotypes. Alignment with the published crystal structure of AV2 established the location and structure of the type-specific regions. Five HVRs were shown to be part of linear loops on the exposed surfaces of the protein, analogous to the serotype-specific loops or "puffs" in picornavirus capsid proteins. The HVRs were supported by a common framework of conserved residues, of which 68 to 75% were hydrophobic. Unique sequences were limited to the seven HVRs, so that one or more of these regions contain the type-specific neutralization epitopes. A neutralizing AV48 hexon-specific antiserum recognized linear peptides that corresponded to six HVRs by enzyme immunoassay. Affinity-purification removal of all peptide-reactive antibodies did not significantly decrease the neutralization titer. Eluted peptide-reactive antibodies did not neutralize. Human antisera that neutralized AV48 did not recognize linear peptides. Purified trimeric native hexon inhibited neutralization, but monomeric heat-denatured hexon did not. We conclude that the AV48 neutralization epitope(s) is complex and conformational.     

Construction and Characterization of Hexon-Chimeric Adenoviruses: Specification of Adenovirus Serotype

This study has used the strategy of gene replacement to characterize the contribution of the adenovirus (Ad) capsid protein hexon to serotype definition. By replacing the Ad type 5 (Ad5) hexon gene with sequences from Ad2, we have changed the type specificity of the chimeric virus. The type-determining epitopes are primarily associated with loop 1 of hexon and, to a much lesser degree, with loop 2. In spite of the serotype distinctiveness of the chimeric hexon viruses, epitope similarity between the vectors resulted in a low level of cross-reactive neutralizing antibody, which in combination with activated cellular and innate arms of the immune system is sufficient to suppress gene transduction following readministration in vivo.     

Inferring Epitopes of a Polymorphic Antigen Amidst Broadly Cross-Reactive Antibodies Using Protein Microarrays: A Study of OspC Proteins of Borrelia burgdorferi

Epitope mapping studies aim to identify the binding sites of antibody-antigen interactions to enhance the development of vaccines, diagnostics and immunotherapeutic compounds. However, mapping is a laborious process employing time- and resource-consuming ‘wet bench’ techniques or epitope prediction software that are still in their infancy. For polymorphic antigens, another challenge is characterizing cross-reactivity between epitopes, teasing out distinctions between broadly cross-reactive responses, limited cross-reactions among variants and the truly type-specific responses. A refined understanding of cross-reactive antibody binding could guide the selection of the most informative subsets of variants for diagnostics and multivalent subunit vaccines. We explored the antibody binding reactivity of sera from human patients and Peromyscus leucopus rodents infected with Borrelia burgdorferi to the polymorphic outer surface protein C (OspC), an attractive candidate antigen for vaccine and improved diagnostics for Lyme disease. We constructed a protein microarray displaying 23 natural variants of OspC and quantified the degree of cross-reactive antibody binding between all pairs of variants, using Pearson correlation calculated on the reactivity values using three independent transforms of the raw data: (1) logarithmic, (2) rank, and (3) binary indicators. We observed that the global amino acid sequence identity between OspC pairs was a poor predictor of cross-reactive antibody binding. Then we asked if specific regions of the protein would better explain the observed cross-reactive binding and performed in silico screening of the linear sequence and 3-dimensional structure of OspC. This analysis pointed to residues 179 through 188 the fifth C-terminal helix of the structure as a major determinant of type-specific cross-reactive antibody binding. We developed bioinformatics methods to systematically analyze the relationship between local sequence/structure variation and cross-reactive antibody binding patterns among variants of a polymorphic antigen, and this method can be applied to other polymorphic antigens for which immune response data is available for multiple variants.     

Human antibodies recognize multiple distinct type-specific and cross-reactive regions of the minor capsid proteins of human papillomavirus types 6 and 11.

Human serum samples derived from a case-control study of patients with cervical carcinoma (n = 174) or condyloma acuminatum (n = 25) were tested for the presence of immunoglobulin G antibodies to human papillomavirus type 6 (HPV6) L2 and HPV11 L2 recombinant proteins in a Western immunoblot assay. Thirty-six samples (18%) were positive for HPV6 L2 antibodies alone, 25 (13%) were positive for HPV11 L2 antibodies alone, and 34 (17%) were positive for both HPV6 L2 and HPV11 L2 antibodies. Thirty samples that were positive for both antibodies were tested for the presence of HPV6-HPV11 L2 cross-reactive antibodies. Fifteen (50%) serum samples contained HPV6-HPV11 L2 cross-reactive antibodies, and 15 (50%) contained independent, type-specific HPV6 L2 and HPV11 L2 antibodies. Altogether, 82% of the HPV6 L2 and HPV11 L2 antibody reactivities were type specific and 18% were HPV6-HPV11 cross-reactive. There was no significant difference in the prevalence of antibody reactivities between samples from patients with cervical carcinoma and those with condyloma acuminatum. Deletion mapping identified five HPV6 L2 regions that reacted with HPV6 type-specific antibodies: 6U1 (amino acids [aa] 152 to 173), 6U2 (aa 175 to 191), 6U3 (aa 187 to 199), 6U4 (aa 201 to 217), and 6U5 (aa 351 to 367). Five HPV11 L2 regions that reacted with HPV11 type-specific antibodies were identified: 11U1 (aa 49 to 84), 11U2 (aa 147 to 162), 11U3 (aa 179 to 188), 11U4 (aa 180 to 200), and 11U5 (aa 355 to 367). Two HPV6-HPV11 cross-reactive regions were identified: 6CR1 (HPV6 L2 aa 106 to 128)/11CR1 (HPV11 L2 aa 103 to 127) and 6CR2 (HPV6 L2 aa 187 to 199)/11CR2 (HPV11 L2 aa 180 to 200).     

Isolation of a nucleocapsid polypeptide of herpes simplex virus types 1 and 2 possessing immunologically type-specific and cross-reactive determinants.

A polypeptide (p40) of approximately 40,000 molecular weight was isolated from herpes simplex virus type 1 and 2 nucleocapsids by gel filtration and ion exchange chromatography. This protein appears to be the same as protein 22a described previously (Gibson and Roizman, J. Virol. 10:1044--1052, 1972). Competition immunoassays were developed by using purified p40 and antisera prepared in guinea pigs. The assays indicated that the p40's from herpes simplex virus types 1 and 2 possess both type-specific and cross-reactive antigenic determinants. Antibodies to the p40 cross-reactive determinant reacted with antigens in simian herpes virus SA8-infected cells, but not with antigens induced by pseudorabies virus. Preliminary results indicated that a radioimmunoprecipitation test can be used to detect type-specific herpes simplex virus p40 antibodies in human sera.     

Structural and immunochemical analysis of the products of proteolysis of simian adenovirus hexons

Hexon capsomers of simian adenovirus sim16 (SA7) and of human adenoviruses h5 (Ad5) and h6 (Ad6) were proteolytically digested and the resulting products studied by SDS-polyacrylamide gel electrophoresis and by radioimmunoprecipitation analysis. The trypsinolysis of native SA7 hexon leads to a stable molecular "core" containing 4-5 fragment species of 10 to 65 kDa and resembling the intact capsomer in quarternary structure (trimer). Similar cores but consisting of smaller fragments (less than 40 kDa) were obtained after chymotryptic digestion of native SA7, Ad5 and Ad6 hexons. The chymotryptic hexon fragments were also held together in pseudotrimeric structures. The similarity of proteolytic hexon fragment patterns between different primate adenoviral hexons suggested a homology to exist in localisation of the exposed tryptic and chymotryptic cleavage sites in their respective hexon polypeptide chains. Papain caused a complete hydrolysis of native SA7 hexon (trimer) yielding small peptides, but at first stage of digestion a stable papain hexon core containing small fragments (less than 10 kDa) was observed. The tryptic SA7 hexon cores in native state retained their antigenicity in reactions with homo- and heterologous antibodies, but after core denaturation the resulting fragments had no antigenic activity of native capsomer. In contrast to the data previously published, chymotryptic cores of SA7, Ad5 and Ad6 hexons not only reacted with respective homologous antibodies but also retained (at least in part) cross-reactive antigenic determinants. The questions of formation and stability of native adenoviral hexon conformation are discussed as well as the possible nature of hexon antigenic determinants.     

Effect of monoclonal antibodies on limited proteolysis of native glycoprotein gD of herpes simplex virus type 1.

We examined the properties of 17 monoclonal antibodies to glycoprotein gD of herpes simplex type 1 (HSV-1) (gD-1) and HSV-2 (gD-2). The antibodies recognized eight separate determinants of gD, based on differences in radioimmuno-precipitation and neutralization assays. The determinants were distributed as follows: three were gD-1 specific, one was gD-2 specific, and four were type common. Several type-specific and type-common determinants appeared to be involved in neutralization. We developed a procedure for examining the effect that binding of monoclonal antibody has on proteolysis of native gD-1 by Staphylococcus aureus protease V8. We showed that several different patterns of protease V8 cleavage were obtained, depending on the monoclonal antibody used. The proteolysis patterns were generally consistent with the immunological groupings. With four groups of antibodies, we found that fragments of gD-1 remained bound to antibody after V8 treatment. A 38,000-dalton fragment remained bound to antibodies in three different groups of monoclonal antibodies. This fragment appeared to contain one type-common and two type-specific determinants. A 12,000-dalton fragment remained bound to antibodies belonging to one type-common group of monoclonal antibodies. Tryptic peptide analysis revealed that the 12,000-dalton fragment represented a portion of the 38,000-dalton fragment and was enriched in a type-common arginine tryptic peptide.     

Idiotype-specific T lymphocytes. I. Regulation of antibody production by idiotype-specific H-2-restricted T lymphocytes

Cytotoxic T lymphocytes (CTL) specific for MOPC-104E myeloma cells of BALB/c origin could be induced in BALB/c, (BALB/c X BALB.B)F1, and (BALB/c X BALB.K)F1 mice. (BALB/c X BALB.B)F1 CTL activity specific for MOPC-104E was effectively inhibited by anti-H-2d but not by anti-H-2b alloantiserum. However, the activity was hardly blocked by specific anti-idiotypic antibodies to MOPC-104E. For further analysis of the recognition of idiotype on target cells by CTL, the effect of those lymphocytes on anti-dextran B1355S antibody-producing B lymphocytes, which have a cross-reactive idiotype to MOPC-104E, was investigated. Lymphocytes from the CTL population did inhibit antibody production by dextran-immune spleen cells, but those from the CTL population specific for irrelevant myeloma cells (MOPC-167) did not. The (BALB/c X BALB.K)F1 CTL population suppressed the antibody production of BALB/c but not of BALB.K. This indicates that F1 cells can preferentially see H-2 antigens of immunizing myeloma cells on target B lymphocytes. The inhibition of antibody production was antigen specific and was only restricted to the PFC that were inhibitable by anti-idiotypic antibodies. The surface phenotypes of the cells that inhibited the antibody production were Thy-1+, Lyt-1-, Lyt-2+, and I-J-. These results strongly suggest that CTL specific for MOPC-104E recognize self H-2 antigens simultaneously with idiotypic determinants on B lymphocytes. Possible immunoregulatory roles of idiotype-specific CTL on antibody production systems are also suggested.     

Serological cross-reactivity of murine A.TH anti-A.TL antibody (anti-l-Ek antibody) with La-like molecules of human antigen-presenting cells

The purified protein derivative (PPD)-specific proliferative responses of T cells from human peripheral blood are shown to be dependent on antigen-presenting cells (APC) which bear HLA-DR antigen detected by the monoclonal anti-HLA-DR antibody. The serological cross-reactivity of murine A.TH anti-A.TL antibody was observed in human APC. By absorption experiments using H-2 congenic mice, the serological cross-reactivity of A.TH anti-A.TL antibody with human APC is mapped in the I-E subregion. Thus, anti-I-Ek antibody reacts with the Ia-like molecule(s) on human APC. Murine allo-anti-I-Ek antibody does not always react with determinants of Ia-like molecule(s) on human APC, since this antibody did not eliminate PPD-specific proliferative responses in one particular case. Thus, anti-I-Ek antibody seems to react some type of the polymorphic determinants but not of the shared determinants of human Ia-like molecule(s) on APC. The relationship between the cross-reactive molecule detected by murine allo-anti-I-Ek antibody and the HLA-DR antigen remains to be analyzed.     

Analysis of the diversity of murine antibodies to dextran B1355. I. Generation of a larger, pauci-clonal response by a bacterial vaccine.

Mice immunized with a combination of dextran B1355 in adjuvant followed by three injections of 2 x 10(9) Escherichia coli B organisms produced an average of 14.5 mg/ml of anti-dextran antibodies. It was demonstrated that the stimulating effect of E. coli B was due to antigenic determinants cross-reactive with B1355 and not solely because of adjuvant properties of the organism. The anti-dextran antibodies were distributed among both 7S and 19S components. Isoelectric focusing of the 7S antibodies showed several spectrotypes of antibody, most of which were shared by the majority of the individual sera. The limited spectrotypic heterogeneity of the 7S antibodies was supported by idiotypic studies. Thus, a heterologous, anti-idiotypic serum, rabbit anti-M104, was prepared which distinguished between two closely related myeloma proteins, M104 and J558,with specificity for alpha-(1 leads to 3) dextran. This antiserum demonstrated that some, but not all, of the 7S and 19S anti-dextran antibodies possessed variable region determinants cross-reactive with M104.     

Human peripheral blood lymphocytes from recently vaccinated individuals produce both type-specific and intertypic cross-reacting neutralizing antibody on in vitro stimulation with one type of poliovirus

An in vitro system of poliovirus-specific antibody production by peripheral blood B cells on stimulation by the virus has been developed. Virus-neutralizing antibodies in culture supernatant fluids, or virus-specific antibody-secreting cells (ASC) were detected by microneutralization assay and ELISA-SPOT test, respectively. After booster immunization with polio vaccine, anti-poliovirus-neutralizing ASC were present in circulation. This response was measurable between 5 and 12 days after booster vaccination. At between 12 and 90 days, another subset of B cells was found in peripheral blood that only produced poliovirus-specific neutralizing antibody after in vitro antigenic stimulation. The in vitro virus-induced response required B cells, monocytes, and T4+ (T helper) cells, and was shown to result from de novo protein synthesis. The anti-poliovirus-neutralizing response in vitro could be dissected in a type-specific and intertypic cross-reactive response by using various antigen concentrations for in vitro stimulation. Evidence was obtained by absorption studies for the existence of intertypic cross-reactive neutralization-inducing epitopes.     

Immunoenzyme analysis of adenovirus hexon. Determination of antibodies from hyperimmunized chickens

We have elaborated three systems of enzyme-linked immunosorbent assay (ELISA) for detection of chicken IgG antibodies specific for hexon antigens of three immunologically distinct adenovirus groups: those of mammalian adenoviruses (Mastadenovira), typical avian adenoviruses (Aviadenovira) and of egg-drop syndrome-76 (EDS-76) virus. In each system the antibodies against respective hexons were specifically detected. In mammalian adenovirus hexons the ELISA detects primarily the type-specific (epsilon) and genus-specific (alpha) antigenic determinants. The time course of anti-hexon antibodies content was followed during immunization. The level of anti-hexon antibodies in egg yolk reflects adequately their content in blood serum. The technique is suitable for serological diagnosis of chicken adenoviral infections as well as for characterization of egg-yolk antibodies obtained by preparative hyperimmunization of hens.     

Glycosylation of immunodominant linear epitopes in the carboxy-terminal region of the caprine arthritis-encephalitis virus surface envelope enhances vaccine-induced type-specific and cross-reactive neutralizing antibody responses

This study evaluated type-specific and cross-reactive neutralizing antibodies induced by immunization with modified surface glycoproteins (SU) of the 63 isolate of caprine arthritis-encephalitis lentivirus (CAEV-63). Epitope mapping of sera from CAEV-infected goats localized immunodominant linear epitopes in the carboxy terminus of SU. Two modified SU (SU-M and SU-T) and wild-type CAEV-63 SU (SU-W) were produced in vaccinia virus and utilized to evaluate the effects of glycosylation or the deletion of immunodominant linear epitopes on neutralizing antibody responses induced by immunization. SU-M contained two N-linked glycosylation sites inserted into the target epitopes by R539S and E542N mutations. SU-T was truncated at 518A, upstream from the target epitopes, by introduction of termination codons at 519Y and 521Y. Six yearling Saanen goats were immunized subcutaneously with 30 microg of SU-W, SU-M, or SU-T in Quil A adjuvant and boosted at 3, 7, and 16 weeks. SU antibody titers determined by indirect enzyme-linked immunosorbent assay demonstrated anamnestic responses after each boost. Wild-type and modified SU-induced type-specific CAEV-63 neutralizing antibodies and cross-reactive neutralizing antibodies against CAEV-Co, a virus isolate closely related to CAEV-63, and CAEV-1g5, an isolate geographically distinct from CAEV-63, were determined. Immunization with SU-T resulted in altered recognition of SU linear epitopes and a 2.8- to 4.6-fold decrease in neutralizing antibody titers against CAEV-63, CAEV-Co, and CAEV-1g5 compared to titers of SU-W-immunized goats. In contrast, immunization with SU-M resulted in reduced recognition of glycosylated epitopes and a 2.4- to 2.7-fold increase in neutralizing antibody titers compared to titers of SU-W-immunized goats. Thus, the glycosylation of linear immunodominant nonneutralization epitopes, but not epitope deletion, is an effective strategy to enhance neutralizing antibody responses by immunization.     

Structural studies on induced antibodies with defined idiotypic specificities. V. The complete amino acid sequence of the light chain variable regions of anti-p-azophenylarsonate antibodies from A/J mice bearing a cross-reactive idiotype.

The complete amino acid sequence of the variable regions of light chains derived from anti-p-azophenylarsonate antibodies from A/J mice bearing a cross-reactive idiotype is reported. At least two and probably more than three distinct light chains are associated with this idiotypically characterized antibody. The antibodies have several differences in their "framework" structures but evidence is presented indicating that all three light chain hypervariable regions have a homogeneous sequence. The data are discussed in relation to the various theories of antibody diversity. In addition, the findings support the view that hypervariable regions, idiotypic determinants, and the antibody-combining site involve, to a large extent, the same molecular structures.