Three distinct epitopes within the loop region of hen egg lysozyme defined with monoclonal antibodies.

Five monoclonal antibodies specific for the loop region of hen egg lysozyme were prepared by immunisation with a synthetic conjugate of a proteolytic fragment of lysozyme coupled to bovine serum albumin. Their fine specificities were investigated using a panel of variant lysozymes and peptide fragments of lysozyme in a quantitative radio-immunoassay procedure. Knowledge of the structure of hen lysozyme to high resolution and the use of computer graphics enables the localisation of the epitopes recognised by the antibodies with some precision. The antibodies were shown to define three distinct, overlapping epitopes within what was previously considered to be a single antigenic site. These results are discussed in relation to current ideas of the antigenic nature of proteins and other recent studies in which anti-protein antibodies have been elicited by immunisation with small peptides.     

Epitopic analysis by anti-I-Ak monoclonal antibodies of I-Ak-restricted presentation of lysozyme peptides

Anti-I-A mAb were used as probes of functional epitopes for both the presentation of hen egg lysozyme (HEL) peptides to I-Ak-restricted T cell hybridomas and the direct binding of the HEL (46-61) peptide. When mAb directed to polymorphic regions of I-Ak were used as inhibitors of Ag presentation, several different patterns of inhibition were observed among T cells specific for the same HEL peptide as well as among T cells specific for different fragments of HEL. Although there appears to be a conserved usage of some TCR V beta gene segments among the T cell hybrids specific for the same HEL peptide, no correlation is evident between a single V gene usage and susceptibility to blocking of Ag presentation by a particular anti-I-Ak mAb. Several of the mAb demonstrated T cell "clonotypic blocking" of Ag presentation, whereas others blocked presentation to every T cell hybrid tested, regardless of the peptide specificity. When mAb directed to nonpolymorphic regions of the I-A molecule were tested for their ability to block Ag presentation, little or no inhibition was observed. In addition, Fab' fragments of inhibitory mAb functioned identically to their intact homologous counterparts in their ability to block Ag presentation indicating that "nonspecific" steric hindrance was not playing a major role in the inhibitions observed. When the polymorphic region-directed anti-I-A mAb were tested for their ability to block the direct binding of the lysozyme peptide HEL(46-61) to I-Ak, those mAb that block HEL presentation to all T cell hybrids were found to block the binding of this peptide. However, anti-I-A mAb that demonstrate selective inhibition of T cell hybrid stimulation during Ag presentation, i.e., those directed to polymorphic serologic specificities Ia.15 and Ia.19, do not block the binding of HEL(46-61) to I-Ak. These data indicate that functionally independent epitopes exist on the I-Ak molecule for the binding of antigenic peptides and for interaction with the TCR.     

Protective efficacy of IgM monoclonal antibodies in experimental group B streptococcal infection is a function of antibody avidity

We have produced and characterized six mAb directed against group B streptococci (GBS). All antibodies are IgM. We have previously shown that some of these antibodies are highly protective in the treatment of experimental infections in neonatal rats, whereas others do not appear to have any protective efficacy. Using an ELISA, we demonstrate the specificity of both protective and nonprotective antibodies. Two antibodies, binding different epitopes, are directed against antigenic structures present on all GBS; two are specific for type III carbohydrate determinants; one binds to a protein Ag present on all type I and II GBS; and one appears to bind to type Ia GBS only. Quantitative absorption assays provide evidence that the difference between protective antibodies and nonprotective antibodies is the avidity that the antibody demonstrates for the epitope recognized on the surface of the bacteria; 10 to 15 times as much protective antibody binds to GBS as does nonprotective antibody. Direct binding experiments with radiolabeled antibody confirm this conclusion.     

A set of closely related antibodies dominates the primary antibody response to the antigenic site CB of the A/PR/8/34 influenza virus hemagglutinin

Approximately 50% of the primary antibody response of BALB/c mice to the A/PR/8/34 influenza virus hemagglutinin is directed to the Cb site, one of the four major antigenic regions of the molecule. To determine the structural basis of the anti-Cb site response, we have examined the paratypic and genetic diversity exhibited by a panel of 24 primary and 4 secondary response mAb specific for this antigenic region. Reactivity pattern analysis demonstrated 20 distinct fine specificities among these antibodies, and V region gene sequence analysis showed that they are encoded by 17 different VH gene segments from 6 VH gene families and 14 different VK gene segments from 6 VK gene groups. Despite this overall diversity, many of the antibodies can be placed in a limited number of sets based on the shared expression of VH and/or VK genes. One set contains antibodies encoded by a single gene of the VK4/5 group in combination with one of two closely related genes from the J558 VH family. This set accounts for half of the Cb site-specific primary response hybridomas, indicating that the representation of the various anti-Cb site B cell specificities during the primary response to A/PR/8/34 influenza virus is not uniform. The preferential participation of B cells expressing this VH/VK combination is largely responsible for the dominance of anti-Cb site antibodies in the primary anti-hemagglutinin response.     

Epitope mapping of antigenic determinants of hepatitis C virus proteins by phage display

Study of individual hepatitis C (HCV) proteins could help to find a molecular structure and conformation, localization of antigenic and immunogenic determinants, to reveal of protective epitopes. It is necessary for practical medicine - development of diagnostic test-systems, vaccines and therapeutics. Linear and conformation dependent epitopes of HCV proteins was localized in this work and immunogenic properties of phage displayed peptides screened on monoclonal antibodies to HCV proteins have been investigated. Eleven epitopes of four HCV proteins have been studied. Three epitopes was found as linear, two epitopes were dependent on secondary structure of proteins and one epitope was dependent on tertiary structure of NS3 protein. Aminoacid sequences of other determinants have been determined and the distinct localization of these determinants will be continued after discovering of tertiary structure of HCV proteins. It was shown, that phage mimotope 3f4 is immunogenic and could induce specific hu- moral immune response to NS5A HCV protein. The data obtained could be useful for improving of HCV diagnostic test-systems, studying of amino acid substitutions and its influence on antigenic properties of the HCV proteins. The results could help to study an immune response in patients infected with different genotypes of HCV. Phage displayed peptides mimicking the antigenic epitopes of HCV proteins could be applied to development of HCV vaccine.     

Immunoregulation by monoclonal sheep erythrocyte-specific IgG antibodies: suppression is correlated to level of antigen binding and not to isotype

Six different monoclonal IgG antibodies with specificities for sheep erythrocytes (SRBC) were tested for immunosuppressive ability. Four of them, one IgG3, two IgG2a, and one IgG1, could yield suppression of more than 90% of the anti-SRBC response. The remaining two antibodies, which were both IgG2a, were found to have no significant effect. The degree of suppression correlated well with the amount of antibodies used that could bind to SRBC, as measured by an ELISA assay. High avidity for SRBC was also a factor making the monoclonal antibody more efficient as an immunosuppressor. The response against antigenic determinants on the SRBC other than those for which the monoclonals were specific, was suppressed to an equal degree. This was established by immunizing mice with SRBC using monoclonal anti-SRBC antibodies that did or did not bind to goat RBC (GRBC). The PFC responses against both SRBC and GRBC were then measured. The anti-SRBC and GRBC responses were suppressed in parallel regardless of whether or not the monoclonal reacted with GRBC. None of the tested antibodies displayed any significant ability to enhance the anti-SRBC response. Thus, IgG1 is not the only murine isotype that can efficiently suppress the immune response against SRBC, but IgG2a and IgG3 can also exert this capacity. The mechanism of IgG-mediated suppression is not one of merely blocking single epitopes but involves the immunogenicity of the entire SRBC.     

Autoantibodies to thyroglobulin are encoded by diverse V-gene segments and recognize restricted epitopes.

Murine experimental autoimmune thyroiditis has been used as a model for human autoimmune thyroiditis. Experimental autoimmune thyroiditis is induced in mice by immunization with mouse thyroglobulin (Tg) in CFA. To characterize the antibodies to this autoantigen, we have studied the binding specificities and determined the nucleotide sequences of monoclonal anti-Tg antibodies. The specificities of the mAb for determinants on Tg varied extensively. Seven of 16 mAb showed reactivity to only mTg, 4 reacted to Tg from more than one species and four reacted to a variety of Ag. Many of the mAb were competitively inhibited by thyroid hormones, suggesting that they recognize the hormonogenic sites on the Tg molecule. The mAb could be divided into at least seven reactivity patterns based on reciprocal competitive inhibition studies, indicating that mTg contains at least seven antigenic regions. DNA sequence analysis of the mAb showed that a large number of V region gene segments encoded the H and L chains. No evidence for preferential use of any V region family or gene segment was found. Gene segments from the VH 7183, Q52, J558, and VH10 families were used by heavy chains, and the V kappa 1, 4, 8, 9, 19, and 21 families were used by kappa-chains. The results indicate that the antigenic epitopes on mTg elicit a very diverse autoantibody response that is derived from a large number of V region gene segments. Many of these autoantibodies show specific reactivity with mTg indicating they recognize species specific epitopes. The results suggest that clonal deletion of autoreactive Ab to certain self-epitopes may not occur.     

Multiple overlapping epitopes in the three antigenic regions of horse cytochrome c1

To gain a better understanding of the diversity of epitopes on a protein, the specificities of 103 monoclonal antibodies to a model antigen, horse cytochrome c(cyt c), were analyzed. The antibodies were generated in in vitro monoclonal, secondary antibody responses against horse cyt c coupled to hemocyanin in splenic fragment cultures. For this assay, horse cyt c-primed murine B lymphocytes were transferred to irradiated, hemocyanin-primed recipients. A panel of seven mammalian cyts c differing at one to six residues out of 104 and cyanogen bromide-cleaved fragments of horse cyt c containing residues 1-65, 1-80, and 66-104 was used to examine the specificities of the antibodies. Twenty-two distinct reactivity patterns were observed, even though the majority of the monoclonal antibodies were found to bind in the three previously identified antigenic regions of the molecule about residues 44-47, 60-62, and 89-92. The results indicate that each of the three antigenic regions consists of multiple overlapping epitopes. Few of the antibodies directed to any given antigenic region bound polypeptide fragments inclusive of the epitope sequences, demonstrating that some antibodies were more conformationally dependent than others. Only 13% of the antibodies bound to cyanogen bromide-cleaved polypeptide fragments that together encompassed the entire length of the protein. Considering the large number of antibodies analyzed and the reoccurrence of 13 of the 22 clonotypes in different lymphocyte donors, it is likely that the antibody specificities tabulated herein approach yet do not completely enumerate the total inventory of the horse cyt c-specific B cell repertoire. The remarkable diversity for epitope recognition within antigenic regions observed here is likely to pertain to protein antigens in general, and strongly supports the widely held notion that the entire surface of a protein is potentially antigenic. The restriction of the epitopes of horse cyt c to three antigenic regions where the amino acid sequences of the mammalian cyts c differ probably results from tolerance of the mice to their own cyt c.     

Immunization with antigen bound to bispecific antibody induces antibody that is restricted in epitope specificity and contains antiidiotype.

Mice can be efficiently immunized in the absence of adjuvant using chemically cross-linked bispecific antibody (biAb) that bind to both class II MHC molecules and a protein Ag of interest. In our experiments, mice were immunized with the protein Ag hen egg lysozyme (HEL) bound to several different biAb, each of which contained a different mAb specific for a distinct (nonoverlapping) epitope of HEL. Primary and secondary serum antibody responses of the immunized mice were analyzed for their specificity for different epitopes of HEL. The results show that immunization with each HEL-biAb complex produced a bias in the epitope specificity of the primary antibody response. This bias was determined by the individual specificity of the anti-HEL mAb used in each biAb. The primary response was dominated by antibody reacting with epitopes distinct from that bound by the mAb in the immunizing complex, and was deficient in antibody that recognized the epitope bound by the biAb during immunization. This bias in antibody specificity was maintained during the secondary antibody response that followed a single challenge with soluble HEL alone. However, an additional challenge with HEL induced a switch in the specificity pattern, with increased amounts of antibody against the epitope that was previously ignored. In addition, immunization with Ag bound to biAb resulted in a substantial primary anti-Id response, detected by serum antibody specific only for the Fab'2 fragment of the mAb used in the biAb. These studies illustrate two unique features of immunization using biAb that allow for fine manipulation of the epitope specificity and anti-Id repertoires of the antibody response to whole protein Ag.     

The regulation of resistance to Schistosoma mansoni by auto-anti-idiotypic immunity. III. An analysis of effects on epitopic recognition, idiotypic expression, and anti-idiotypic reactivity at the clonal level.

Auto-anti-idiotypic mechanisms can regulate the protective immune response against Schistosoma mansoni. Anti-idiotypic responses were stimulated by immunization of mice either with nonspecifically induced lymphoblasts, produced with Con A, or with Ag-induced lymphoblasts bearing specific idiotypic receptors. The effect of the induced anti-idiotypic response upon clonotypic cellular reactivity was assessed in vitro through the suppression of antigen-mediated blast transformation by cloned T cells and in vivo by suppression of resistance to S. mansoni and delayed-type hypersensitivity responses against specific Ag. Differential regulation of humoral immune responses was studied at the levels of specific epitopic recognition, the expression of specific Id, and the production of anti-idiotypic responses directed against mAb bearing specific Id. Anti-idiotypic sensitization resulted in variable (10 to 90%) suppression of the immune response to discrete antigenic epitopes, the expression of specific idiotypic phenotypes, and anti-idiotypic, antiparatopic responses against T cell clonotypes and antibody idiotypic phenotypes. In vitro admixture and in vivo challenge studies resulted in consonant differential suppression. Thus idiotypic regulation can mold the fine specificities of the protective immune response to S. mansoni at the clonal level and may provide an approach to optimize the expression and assessment of resistance.     

Mapping of antigenic determinants of hepatitis C virus proteins using phage display

Analysis of the properties for individual hepatitis C virus (HCV) proteins makes it possible to establish their molecular structure and conformation, to localize antigenic and immunogenic determinants, to identify protective epitopes, and to solve applied problems (e.g., design of diagnostic tests, vaccines, and drugs). Linear and conformational epitopes of HCV proteins were localized using the phage display technique, and the peptides exposed on the phages selected with monoclonal antibodies against HCV proteins were tested for immunogenicity. Of the 11 epitopes revealed, three were strongly linear; two depended on the secondary; and one on the tertiary structure of the corresponding protein (conformational epitopes). Amino acid sequences involved in the other epitopes were established. The results can be used to improve the diagnosis of hepatitis C, to study the effect of amino acid substitutions on the antigenic properties of HCV proteins, and to analyze the immune response in patients infected with genotypically different HCV. It was shown with the example of the NS5A epitope that phage particles with epitope-mimicking peptides (mimotopes) induce production of antibodies against the corresponding HCV proteins.     

Specificity of monoclonal antibodies against human thyroglobulin; comparison with autoimmune antibodies.

Ten monoclonal antibodies (mAb) directed against human thyroglobulin (hTgb) were produced, purified and characterized. The mAb avidity for hTgb ranged from 10(-10) to 10(-6) M. The species specificity of the mAb was as follows: eight mAb reacted with monkey Tgb, three with dog Tgb and one with pig Tgb; none with bovine and ovine Tgb. The binding of mAb to hTgb was not significantly inhibited in the presence of Tgb carbohydrate moieties, tyrosine, iodotyrosines and iodothyronines. The topology of the antigenic determinants recognized by the 10 mAb on hTgb was explored by inhibition of Tgb binding of radiolabeled mAb by the other antibodies. Six distinct clusters of reactivity were described. Localization of the antigenic determinants recognized by mAb on hTgb was attempted using tryptic fragments of hTgb to inhibit the binding of mAb to hTgb. The inhibitory effect of hydrolysis products was different for each mAb but exhibited partial analogies between mAb of the same cluster of reactivity. Anti-hTgb autoimmune antibodies (aAb) purified from sera of Graves patients cross-reacted essentially with mAb of one out of the six clusters. These results demonstrate that the large number of antigenic determinants presented by the hTgb are not disseminated on the molecule but are clustered in antigenic regions. Furthermore, from the six antigenic regions evidenced in this paper, only one is involved in autoimmune antibody production in Grave's disease.     

Eimeria tenella: B-cell epitope mapping following primary and secondary infections

Immunisation against coccidiosis has become more reliable and effective with improved administration techniques for new vaccines. On the other hand, an ideal coccidial vaccine should contain both B- and T-cell immunogenic epitopes. Fine specificity of B-cell epitopes recognised by antibodies prepared following primary and secondary infections with Eimeria tenella were studied using "PepScan" techniques. Mapping of B-cell epitopes within an antigenic sequence from E. tenella showed that four distinct types of epitopes were recognised by the host immune system during the primary and secondary infections with the parasite. These observations demonstrated that new epitopes are also involved in induction of antibody responses following the secondary infection.     

A rabbit antiserum detects a VH J558 subgroup marker highly expressed among anti-alprenolol antibodies

A rabbit antiserum raised against anti-alprenolol mAb 14C3 detects common antigenic determinants (ADC3) in 10 out of 14 anti-alprenolol mAb that use different germ-line VH and/or Vk genes. The anti-14C3 antiserum binds only to H chains in immunoblots, therefore suggesting that at least part of the ADC3 determinants may be encoded by H chain V region genes. Analysis of VH gene family usage among the anti-alprenolol mAb reveals that the expression of ADC3 correlates with utilization of VH genes that belong to the J558 gene family, regardless of the JH, Vk, and Jk genes. To determine whether the ADC3 determinants are general V region markers or whether they are unique to anti-alprenolol antibodies, we have extended our analysis to a random panel of antibodies that also use VH genes of the J558 family. Among 23 mAb of various specificities, 14 react with the anti-14C3 antiserum in immunoblot and in ELISA, irrespective of antibody specificity. Adsorption of the antiserum on one of these positive antibodies results in a loss of reactivity toward both anti-alprenolol and unrelated antibodies. Therefore, several but not all antibodies that use a J558 VH gene also express the complete set of epitopes defining ADC3. These results strongly suggest that ADC3 are markers of a subset of J558 VH gene products. The anti-14C3 antiserum may thus constitute a "serologic probe" for identification of a VH gene subgroup from the J558 gene family.     

Masking of antigenic epitopes by antibodies shapes the humoral immune response to influenza

The immune responses to influenza, a virus that exhibits strain variation, show complex dynamics where prior immunity shapes the response to the subsequent infecting strains. Original antigenic sin (OAS) describes the observation that antibodies to the first encountered influenza strain, specifically antibodies to the epitopes on the head of influenza''s main surface glycoprotein, haemagglutinin (HA), dominate following infection with new drifted strains. OAS suggests that responses to the original strain are preferentially boosted. Recent studies also show limited boosting of the antibodies to conserved epitopes on the stem of HA, which are attractive targets for a ‘universal vaccine’. We develop multi-epitope models to explore how pre-existing immunity modulates the immune response to new strains following immunization. Our models suggest that the masking of antigenic epitopes by antibodies may play an important role in describing the complex dynamics of OAS and limited boosting of antibodies to the stem of HA. Analysis of recently published data confirms model predictions for how pre-existing antibodies to an epitope on HA decrease the magnitude of boosting of the antibody response to this epitope following immunization. We explore strategies for boosting of antibodies to conserved epitopes and generating broadly protective immunity to multiple strains.     

The concept and operational definition of protein epitopes

The antigenic determinants or epitopes of a protein correspond to those parts of the molecule that are specifically recognized by the binding sites or paratopes of certain immunoglobulin molecules. Epitopes are thus relational entities that require complementary paratopes for their operational recognition. Some authors consider that the concept of epitope necessarily involves the two properties of antigenic reactivity (ability to bind to a paratope) and immunogenicity (ability to induce an immune response). Such a view creates difficulties because it makes the existence of epitopes in a protein depend on immunogenetic and regulatory mechanisms of the immunized host. The delineation of epitopes can be achieved by antigenic cross-reactivity studies or by X-ray crystallography. Both approaches require specific criteria for deciding which residues of the antigen are in contact with the paratope and are functionally part of the epitope. The relative contribution of static accessibility, segmental mobility and induced fit to immune recognition remains controversial. Each of the methods used for analysing antigenic specificity is subject to various operational constraints originating from the type of experimental probe and from the format sensitivity and specificity of the immunoassay used. If a protein is assumed to contain as many epitopes as the number of different monoclonal antibodies that can be raised against it, the delineation of epitopes corresponds to the summation in various hosts of the immune repertoire specific for the antigen. Neutralization epitopes are a special subclass of the epitopes of infectious agents and toxins that are specifically recognized by antibody molecules able to neutralize the biological activity of the antigen. The identification of neutralization epitopes is important for the development of synthetic vaccines because it is this type of epitope that should be mimicked by synthesis and used as a vaccine for eliciting protective immunity. The first demonstration that synthetic peptides could elicit antibodies that neutralized viral infectivity was made by Anderer and his colleagues in the 1960s in their work with tobacco mosaic virus. Nearly 20 years passed before it was shown that antibodies to synthetic peptides were also able to neutralize the infectivity of other viruses such as foot-and-mouth disease, polio and hepatitis B viruses.     

Mechanisms of allergen-antibody interaction of cockroach allergen Bla g 2 with monoclonal antibodies that inhibit IgE antibody binding

Background

Cockroach allergy is strongly associated with asthma, and involves the production of IgE antibodies against inhaled allergens. Reports of conformational epitopes on inhaled allergens are limited. The conformational epitopes for two specific monoclonal antibodies (mAb) that interfere with IgE antibody binding were identified by X-ray crystallography on opposite sites of the quasi-symmetrical cockroach allergen Bla g 2.

Methodology/Principal Findings

Mutational analysis of selected residues in both epitopes was performed based on the X-ray crystal structures of the allergen with mAb Fab/Fab′ fragments, to investigate the structural basis of allergen-antibody interactions. The epitopes of Bla g 2 for the mAb 7C11 or 4C3 were mutated, and the mutants were analyzed by SDS-PAGE, circular dichroism, and/or mass spectrometry. Mutants were tested for mAb and IgE antibody binding by ELISA and fluorescent multiplex array. Single or multiple mutations of five residues from both epitopes resulted in almost complete loss of mAb binding, without affecting the overall folding of the allergen. Preventing glycosylation by mutation N268Q reduced IgE binding, indicating a role of carbohydrates in the interaction. Cation-π interactions, as well as electrostatic and hydrophobic interactions, were important for mAb and IgE antibody binding. Quantitative differences in the effects of mutations on IgE antibody binding were observed, suggesting heterogeneity in epitope recognition among cockroach allergic patients.

Conclusions/Significance

Analysis by site-directed mutagenesis of epitopes identified by X-ray crystallography revealed an overlap between monoclonal and IgE antibody binding sites and provided insight into the B cell repertoire to Bla g 2 and the mechanisms of allergen-antibody recognition, including involvement of carbohydrates.     

Epitope-specific antibody response to murine hepatitis virus-4 (strain JHM)

Monoclonal hybridoma antibodies to the structural proteins of murine hepatitis virus-4, strain JHM (MHV-4) were used in a competition binding enzyme immunoassay to analyze at the epitope level the antibody response of mice after infection with MHV-4. Colonized mice often had pre-existing MHV antibodies directed against epitopes on the E2 glycoprotein, the E1 glycoprotein, and the nucleocapsid protein. These mice generated a secondary antibody response after virus inoculation, reaching peak levels 7 days after infection. In contrast, Nude/+ mice raised in a pathogen-free colony had no detectable circulating MHV antibodies and generated a primary antibody response which gradually increased to peak levels 14 to 28 days after infection. Kinetics of antibody responses against specific epitopes usually correlated well with measured total virus-specific antibody responses, but variation was observed. Mice injected with three antigenically distinct strains of MHV made antibody responses to conserved epitopes but not to an antigenic determinant absent in these strains. Measurement of epitope-specific responses in a polyclonal population of viral specific antibodies is feasible and a valuable adjunct in understanding viral immunity.     

B细胞表位作图研究进展

抗原-抗体的特异性结合是由抗体表面的抗原决定簇与抗原表面的表位基序间的特异性互补识别决定的。B细胞表位作图既包括B细胞抗原表位基序的鉴定(即确定抗原分子上被B细胞表面受体或抗体特异性识别并结合的氨基酸基序),也包括绘制抗原蛋白的全部或接近全部的B细胞表位基序在其一级或高级结构上的分布图谱的过程。B细胞表位作图是研发表位疫苗、治疗性表位抗体药物和建立疾病免疫诊断方法的重要前提。目前,已经建立了多种B细胞表位鉴定或绘制抗原蛋白B细胞表位图谱的实验方法。基于抗原-单抗复合物晶体结构的X-射线晶体学分析的B细胞表位作图和基于抗原蛋白或抗原片段的突变体库筛选技术的B细胞表位作图可以在氨基酸水平,甚至原子水平上揭示抗原分子上与单抗特异性结合的关键基序;其它B细胞表位作图方法(如基于ELISA的肽库筛选技术)常常只能获得包含B细胞表位的抗原性肽段,因而,很少用于最小表位基序的鉴定;而改良的生物合成肽法多用于B细胞表位的最小基序鉴定和精细作图。鉴于每种B细胞作图方法都存在各自的优势与不足,B细胞表位作图往往需要多种作图方法的有机结合。本文对目前常用的B细胞表位作图的实验方法及其在动物疫病防控中的应用进行综述,以期为研究者设计最佳的表位作图方案提供参考。     

Immunogenic properties of multiple antigen peptide systems containing defined T and B epitopes.

Immunization with chemically defined synthetic polymers, multiple Ag peptide (MAP) systems, containing T and B epitopes of the circumsporozoite protein of P. berghei induce high levels of circulating antibodies that are detectable several months after boosting. The anti-MAP secondary antibody response is characterized by an increase in the levels of circulating IgG and a concomitant decrease in the IgM levels. In vitro and in vivo experiments indicated that Th epitopes included in the MAP are recognized by T cells induced after immunization with the native protein and, also, that MAP-induced T cells can recognize the native protein. In addition to high levels of anti-B epitope antibodies, MAP immunization also induces antibodies against the T epitope. This anti-T epitope immune response does not affect the generation of the anti-B epitope antibodies. Immunization of different strains of mice revealed that the antibody response is consistent with the genetically restricted pattern of recognition of the T epitope. There are, however, significant differences in the levels of antibody responses observed among responder strains. The findings of this study indicate that MAP are potent immunogens capable of inducing immunologic memory and are, thus, good candidates for the development of subunit vaccines designed to induce high levels of circulating antibodies.