A Radioimmunoassay for Evaluation of the IgE and IgG Antibody Responses in the Rat

An assay, the paper radioallergosorbent test (PRAST), for the measurement of specific serum IgE antibody in the rat is described in detail. This assay has been used, in conjunction with a modified PRAST for the determination of relative specific serum IgG antibody and the PRIST assay for total serum IgE [13], to measure specific IgE and IgG and total IgE immune responses in normal parasite infected rats immunized using various protocols. The results indicate that there is a relationship between the basic IgE level and the immune response, i.e. a rat strain with a low constitutive IgE level demonstrates a weak response whereas a high level strain reacts strongly. When PRAST and passive cutaneous anaphylaxis (PCA) were compared, using standardized IgE antibody containing sera, the results were in good agreement. However, PRAST is the preferable assay as it shows less intrinsic variation, is more sensitive than PCA, and is not influenced by high serum IgE levels in the recipient animal.     

Reduction of interference by specific IgG with a modified microtiter solid-phase radioimmunoassay to measure honeybee venom IgE

By means of monoclonal mouse anti-human IgE antibody, the microtiter solid-phase radioimmunoassay (MSPRIA) was modified to measure honeybee venom (HBV) IgE. HBV IgG was measured by MSPRIA(G) using affinity column purified goat anti-human IgG antibody and correlated with results obtained with staphylococcal protein A-SPRIA. Nonlinear specific IgE dilution curves were associated with high HBV IgG content. By serial transfer and assay of the supernatant, the interference of IgE by specific IgG could be reduced. The degree of interference was correlated with the specific IgG content. The principle of serial supernatant transfer and assay was applied to the standardization of reference HBV IgE serum, and the quantitation of HBV IgE contents in patients and controls. HBV IgE levels in patients allergic to honeybee stings correlated with the skin-test endpoint titrations. While 75% of the allergic patients were skin-test positive, 83% of them were IgE positive by the MSPRIA and 67% were IgE positive by the RAST.     

An investigation of human immune response to perennial ryegrass (Lolium perenne) pollen cytochrome c (Lol p X)

Cytochrome c (Lol p X) was purified from the pollen of perennial ryegrass (Lolium perenne). It was used in a sensitive double-antibody radioimmunoassay to determine specific IgG and IgE antibody (Ab) levels in the sera of grass-allergic human subjects and the relationship between immune responsiveness and HLA type. A total of 139 subjects were studied, including 63 subjects treated by grass immunotherapy and 76 untreated subjects. The sera of only four subjects were found to contain anticytochrome c IgG Ab; there was no evidence of cytochrome c-specific IgE Ab in any of these patients. Based on these data, we conclude that the prevalence of IgE and IgG Ab responses to this protein is low. Because of the small number of responders, it was not possible to demonstrate a significant association between any HLA-D specificity and immune responsiveness to ryegrass cytochrome c.     

Effect of specific IgG on IgE-induced systemic anaphylaxis in the rabbit

Immunization of rabbits as neonates and periodically thereafter has been shown to induce the long-term preferential production of specific IgE antibodies. Specific IgG antibodies are not detected in the majority (greater than 70%) of rabbits when classical immunological detection techniques are used, including heterologous PCA in guinea-pig skin. Nevertheless, in this study we demonstrate that all rabbits neonatally immunized to the antigen horseradish peroxidase (HRP) do produce low levels of specific IgG antibody detectable by an ELISA technique. Serum levels of anti-HRP IgG were found to be log normally distributed, with a geometric mean for the heterologous PCA-negative sera of 31.6 X/divided by 2.69 micrograms/ml. Serum anti-HRP IgE levels (log2 homologous PCA titres) are bimodally distributed. Specific IgG and IgE levels in individual rabbits have a significant direct relationship. Six heterologous PCA-negative and seven heterologous PCA-positive rabbits were challenged intravenously with HRP. All of the respiratory and circulatory alterations typical of IgE anaphylaxis occurred in every challenged rabbit. Regression analysis of percentage changes in the physiological variables vs log specific IgE level indicated that none of the changes was either directly or inversely related to the specific IgG levels. Also the mean changes of the heterologous PCA-positive vs negative rabbits did not differ significantly. Thus, we could find no evidence for either a blocking or enhancing effect of the specific IgG antibodies (range 10-529 micrograms/ml serum) on the IgE-induced anaphylactic reaction.     

Comparative study of IgG1 and IgE antibody mediated homologous PCA in the mouse ear. Lack of cross-desensitization of IgG1 antibody mediated PCA to IgE antibody mediated PCA

To characterize IgG1 antibody and IgE antibody mediated homologous passive cutaneous anaphylaxis (PCA) in the mouse ear, cross-desensitization was studied using a double-sensitizing technique. Mice were sensitized by injecting into their ears a mixture of two kinds of antibodies with distinct antigen specificities, and PCAs were elicited twice with specific antigens at an appropriate interval. The first PCA was elicited with one antigen free from Evans blue dye and the second with the other antigen in the presence of Evans blue to visualize the reaction. The amount of extravasated dye caused by the latter was determined colorimetrically and compared to that of control which did not receive the first challenge. In PCA mediated by IgG1 antibodies, elicitation of the first reaction significantly inhibited the second reaction evoked after 6, 12, or 24 h. Using IgE antibodies, elicitation of the first PCA also significantly inhibited the following reaction. Inhibition of the second reaction was observed 6 h to 12 days after the first reaction. Conversely, IgG1 antibody mediated PCA did not affect the following reaction caused by IgE antibody. These results strongly suggest that alteration of skin mast cells is smaller in IgG1 antibody than in IgE antibody mediated PCA, when reactions with a similar intensity are evoked. Therefore, it is also suggested that the slight reaction of mast cells initiated by IgG1 antibody might be potentiated by another mechanism in IgG1 antibody mediated PCA.     

Detection of Allergen-Specific IgE Antibody Responses

Allergen-specific IgE production is the central event in the pathogenesis of atopic disorders and increases in specific IgE serum antibodies are an indicator of immediate hypersensitivity responses in humans and in animal models of allergy. Consequently, accurate and user-friendly methods are needed to measure serum levels of allergen-specific IgE. This review examines historical and recent developments in in vivo and in vitro methods for the detection of allergen-specific IgE in humans and in animal models. Routinely, in vitro methods such as enzyme-linked immunosorbant assays or radioallergosorbant tests and in vivo methods such as the skin prick test (SPT) for humans and the passive cutaneous anaphylaxis assay (PCA) used in animals are utilized to detect allergen-specific IgE. While in vivo assays are usually more accurate than in vitro assays since they provide a functional readout of IgE activity, they are relatively costly and require considerable expertise. On the other hand in vitro assays are limited by the fact that the amount of allergen-specific serum IgG exceeds IgE antibody by several orders of magnitude, resulting in competition for allergen binding. Consequently, methods that use allergen as a direct capture step are limited by the availability of free allergen binding sites for IgE. In order to circumvent this problem, in vitro methods usually require prior depletion of IgG or use high amounts of allergen in order to facilitate availability of free binding sites for IgE detection. Clearly, these approaches are limited for small sample volumes and allergens that are in short supply. New methods such as protein microarray could potentially overcome this problem by providing high allergen concentrations in a relatively small reaction volume. Currently, in vitro methods are rarely used in isolation for prognosis but are used primarily to complement the information obtained from in vivo assays. With the emergence of new technologies it is conceivable that in vitro assays may in the future replace in vivo assays, however until then in vivo assays remain the gold standard of allergen-specific IgE detection.     

Homologous passive cutaneous anaphylaxis (PCA) in mice and heterologous PCA induced in rats with mouse IgE.

A study was made of the effect of anit-histamine, antiserotonin and of different anti-anaphylactic drugs on PCA reactions induced in mice with IgG1 or IgE. Further, using the ability of mouse IgE to sensitize rat mast cells, a comparative study was also made of PCA reactions induced in mice and rats with mouse IgE. Antihistamines produced a partial inhibition of PCA reactions induced in mice with mouse IgG1 or IgE and in rats with mouse IgE whereas antiserotonin inhibited PCA reactions induced in rats with mouse IgE, but had no effect on PCA reactions induced in mice with mouse IgG1 or IgE. The simultaneous use of antihistamine and antiserotonin resulted in a total inhibition of PCA reactions induced in mice with IgG1 and in a marked but not total inhibition of PCA reaction due to IgE; PCA reactions induced in rats with mouse IgE were totally inhibited. Compounds known to change the intracellular level of cyclic AMP were found to have little or no effect on PCA reactions induced in mice with either IgG1 or IgE in spite of producing a complete marked inhibition of PCA reactions induced with mouse IgE in rats. Diethylcarbamazine or disodium cromoglycate were also very effective inhibitors of rat PCA reactions induced with mouse IgE although having no effect on PCA reaction induced in mice with this same antibody or with IgG1. Thus, in spite of sharing common mediators released from the same type of target cell sensitized with the same type of antibody, PCA reactions induced in mice and rats with mouse IgE reacted very differently to the pharmacological effect of most of the drugs tested. This fact seems to indicate that the physiological mechanism operating in mouse mast cells are different from those operating in rat mast cells.     

The role of major olive pollen allergens Ole e 1, Ole e 9, and Ole e 10 on mice sensitization.

BACKGROUND: Olive pollen is an important cause of allergy in Mediterranean countries. To date, 10 allergens (Ole e 1 to Ole e 10) have been isolated and characterized. Animal models of olive pollen allergy are suitable tools for testing the efficacy and safety of new forms of immunotherapy. OBJECTIVES: To characterize the immune response in mice sensitized with olive pollen extract and to compare it with that of allergic patients. METHODS: BALB/c mice were sensitized by 4 intraperitoneal injections of olive pollen extract in aluminum hydroxide. The allergic state was proved by measuring serum specific IgG1 and total IgE antibody levels. The IgG1 responses to olive pollen allergens were assayed by immunoblotting and enzyme-linked immunosorbent assay. Competition experiments between human IgE and mouse IgG1 binding to olive pollen allergens were performed. RESULTS: Sensitization with olive pollen extract induced high levels of specific IgG1 and total IgE in all tested animals. Immunoblotting experiments showed that the mouse IgG1 binding pattern to pollen extract was complex and heterogeneous, as occurs with human IgE. High IgG1 antibody levels to the major olive pollen allergens described for humans were detected in serum samples from sensitized mice, whereas minor olive pollen allergens induced no significant IgG1 response. Coincubation of mouse serum samples with a cocktail of Ole e 1, Ole e 9, and Ole e 10 resulted in a significant decrease (60%) in IgG1 binding to olive pollen extract. Specific mouse IgG1 strongly inhibited human IgE binding to olive pollen allergens. CONCLUSIONS: This mouse model of olive pollen sensitization mimics immunologic features of human pollinosis and could be a useful tool for designing novel forms of immunotherapy for olive pollen allergy based on allergen cocktails.     

Familial human short-term sensitizing (IgG S-TS) antibody

Skin tests on a middle-aged English housewife (the initial case) with a history of seasonal grass pollen rhinitis and asthma showed strong immediate and late reactions to grass pollens. Early in the grass pollen season, 1976, RAST showed normal levels of IgE antibodies in the serum. Passive transfer in Rhesus monkey skin was also negative for heat-labile IgE but gave a very vigorous reaction for short-term sensitizing heat-stable IgG antibody (IgG S-TS Ab). Skin tests and studies on sera from other members of the family showed that another five also formed IgG S-TS Ab, and indicated that the ability to form this antibody was familial. One member of the family had only an immediate reaction on skin testing, and much IgE to grass pollen. Towards the end of the grass pollen season the IgE titre in the initial case had been tripled and the IgG S-TS Ab had disappeared. By September the specific IgE titre had risen even further.     

Enhancement of murine IgE antibody detection by IgG removal

RATIONALE: Although animal models for the study of allergic reactions are desirable, the use of mice has been hindered by the lack of sufficiently sensitive in vitro immunoglobulin epsilon (IgE) antibody assays. The aim of this study was to enhance IgE antibody measurements by immunoglobulin gamma (IgG) depletion. METHODS: Seven- to eight-week-old female mice of four strains (C3H/HeJ, CBA/J, C57Bl/6J, and Balb/c) were immunized (20 mice/group) with shrimp or peanut extracts using Al(OH)(3) as adjuvant. Following immunization, animals were sacrificed by exsanguination and the sera of each group pooled. Initial measurements of IgE antibody levels by enzyme-linked immunosorbent assay (ELISA) were relatively low; IgG and IgE reactivity patterns by immunoblot were similar. Thus, sera from shrimp or peanut immunized mice were depleted of IgG (absorbed 3-6 times with immobilized protein G) and then tested for IgE antibody to shrimp or peanut allergen. RESULTS: A 3- to 5-fold increase in IgE antibody reactivity as measured by ELISA was demonstrated when >80-90% of the IgG was removed. This increase in detection of allergen-specific IgE occurred in sera from all mouse strains and to all allergens tested. In addition, reactivity of IgE antibodies to peanut or shrimp allergens by immunoblot increased visually approximately 4- to 10-fold. CONCLUSIONS: These studies indicate that allergen-specific IgG antibodies, which may be in more than 100-fold excess to IgE antibodies, interferes with detection of allergen-specific IgE, probably by competitive binding to allergenic epitopes. Substantial depletion of IgG antibodies (>80%) result in a significant increase in the sensitivity of the antibody measurements.     

A study of the human immune response to Lolium perenne (Rye) pollen and its components, Lol p I and Lol p II (Rye I and Rye II) : I. Prevalence of reactivity to the allergens and correlations among skin test, IgE antibody, and IgG antibody data

In a stratified random sample of 320 white adults, the prevalence of puncture skin test positivity (ST +) to Lolium perenne (rye grass)-pollen extract (LPE) was 16%. Fifteen percent of all subjects (or 84% of subjects classified LPE IgE antibody positive [Ab +]) was classified IgE Ab + to highly purified Lol p I (Rye I), and 4% of all subjects (or 26% of subjects classified LPE IgE Ab +) was classified IgE Ab + to highly purified Lol p II (Rye II). These data and similar results obtained in an allergy-enriched group of 361 subjects are consistent with previous studies that Lol I is a major allergen and Lol II is a minor allergen of LPE. Whether we studied LPE, Lol I, or Lol II, responder subjects were younger than nonresponder subjects and more male than female subjects were responders. We then investigated the quantitative interrelationships among ST, IgE, and IgG Ab responsiveness to LPE, Lol I, and Lol II in the allergy-enriched group. For each allergen, log-log correlations were strong and significant for ST versus IgE Ab and for IgE Ab versus IgG Ab. All subjects IgE Ab + to Lol I or Lol II were IgG Ab + to that allergen, supporting other evidence for a commonality in the genetic control influencing the production of IgE and IgG Abs to a given allergen. Log-log correlations among ST end points, IgE Ab levels, or IgG Ab levels were strong for LPE versus either Lol I or Lol II but weak between Lol I and Lol II, consistent with the reported lack of cross-reactivity between Lol I and Lol II. Despite these findings, almost all Lol II + subjects were Lol I + by ST (98%), IgE Ab (91%), and IgG Ab (83%), suggesting that the Ia-restricted immune recognition of both these molecules is at least in part under a common genetic control.     

Properties of mouse homocytotropic and heterocytotropic antibodies.

Mouse antisera were analyzed for the presence of homocytotropic and heterocytotropic antibodies. Two distinct populations of antibodies were detected by the homologous passive cutaneous anaphylaxis (PCA) reaction. The first was active 2 hr after injection, heat-stable, and partially reactive with antisera to mouse IgG1. The second was active 48 hr after injection, was heat-labile, and probably belonged to the IgE class of mouse immunoglobulins. In addition, it was demonstrated that there were also two antibodies active in the heterologous PCA reaction in rats, a heat-stable antibody and a heat=labile antibody. Contrary to results obtained with homocytotropic antibodies, none of the heterocytotropic antibodies detected reacted with antisera to mouse IgG1 or IgG2. These studies suggest that in addition to IgE there may exist another heterocytotropic antibody in mouse antisera and that caution should be employed when using the 2-hr PCA reaction in the rat as a sole criterion for detection of mouse IgE.     

Bicistronic expression plasmid encoding allergen and anti-IgE single chain variable fragment antibody as a novel DNA vaccine for allergy therapy and prevention

Several approaches have been applied in order to alleviate the difficulties allergic patients are suffering from. Among them DNA vaccination and anti-IgE antibody have shown promising results. Herewith, a combination of both strategies is proposed to minimize IgE production while inducing high levels of blocking IgG and strong Th1 immune responses. A bicistronic expression plasmid including an internal ribosomal entry site (IRES) can express both, allergen and a single chain variable fragment (scFv) antibody against human IgE within antigen presenting cells (APCs) including B cells. Presentation of allergen derived peptides via MHC I and MHC II stimulates specific Th1 responses resulting in high levels of IFN-gamma and IgG. Anti-IgE scFv antibody binds to newly synthesized IgE molecules within B cell cytoplasm and also to free serum IgE, thereby inhibiting attachment of IgE to its receptors on basophils and mast cells. Also, IgE-anti-IgE complex functions as blocking antibody and neutralizes allergens entering the body. Additionally, anti-IgE scFv antibody binds to membrane bound IgE (mIgE) on B cells and interferes with IgE expression. Using assays, such as enzyme linked immunosorbent assay (ELISA), IgG and IgE production in response to this expression system can be evaluated. Also, rat basophil leukemia cell assay (using RBL-2H3 cells) can show the amount of functional IgE in sera as basophil mediator release is regarded as an indicator of the allergic hypersensitive reactions. The proposed approach may result in high levels of blocking IgG and low levels of IgE secretion from B cells. Additionally, it can inhibit activity of IgE in degranulation of basophils and mast cells.     

Immunological safety assessments of anti-IgE antibody which is detached from therapeutic immunoadsorbents for removing IgE

Assessments were made of the safety of antibodies which might be detached from a therapeutic immunoadsorbent (IA) during extracorporeal circulation, with respect to possible immunological responses to such antibodies. The IA used was antihuman IgE antibody (a-IgE Ab) immobilized on a carrier, for removal of IgE from patients' plasma. The antibody was raised in goats and isolated to give an IgG fraction. This fraction was either used without further purification or was subjected to immunoaffinity purification. The active anaphylaxis test in guinea pigs indicated that positive responses were not observed at doses of less than 0.1 micrograms of goat IgG per animal. Rabbits given goat IgG intravenously 3 times a week for 8 weeks did not produce the specific antibody against goat IgG at doses of less than 0.05 micrograms/kg, which corresponds to less than 3 micrograms for an adult with a body weight of 60 kg. However, none of the rabbits given goat IgG at 2.5 mg/kg showed any toxic reactions and different patterns of the body weight growth from these in the control group. In addition, we tested whether immunoaffinity purified a-IgE Ab could trigger Type I hypersensitivity in a monkey model. Anaphylactic reactions were not observed after a single intravenous injection of a-IgE Ab at less than 10 micrograms/kg. These in vivo results are useful to judge whether the amount of antibody that leaks from a therapeutic IA is acceptable or not in a clinical situation.     

Suppression of specific IgE antibody responses by liposome-conjugated ovalbumin in mice sensitized with ovalbumin via the respiratory tract

BACKGROUND: Previously we have shown that intranasal administration of ovalbumin (OVA) together with cholera toxin (CT) abrogates nasal tolerance to OVA, resulting in the induction of specific IgE antibody (Ab) responses, and that intraperitoneal injection of OVA coupled with liposomes (OVA-liposomes) induces a selective suppression of IgE Ab responses to OVA. Whether OVA-liposomes suppress anti-OVA IgE Ab responses in mice sensitized with CT-combined OVA via the respiratory tract remains to be clarified. METHODS: In some experiments, mice were given OVA, liposomes or OVA-liposomes with or without CT intranasally three times, at 2-week intervals (weeks 0, 2 and 4). In other experiments, mice were given OVA-liposomes intranasally 2 days before or 1 and 3 weeks after CT-combined OVA (week 0), which was administered intranasally three times, at 2-week intervals (weeks 0, 2 and 4). Two weeks after the third administration of CT-combined OVA (week 0), nasal wash and serum IgA, IgG and IgE Ab responses were assayed. RESULTS: Pretreatment with OVA-liposomes suppressed IgE Ab responses to CT-combined OVA, with a significantly high production of both nasal IgA and serum IgG Abs. Moreover, treatment with OVA-liposomes 1 and 3 weeks after CT-combined OVA administration also suppressed IgE Ab responses. The suppression of anti-OVA IgE Ab production by OVA-liposomes was accompanied by a simultaneous enhancement of specific IgA and IgG (IgG1, and especially IgG2a) Ab production. CONCLUSIONS: Postimmunization treatment with OVA-liposomes, as well as preimmunization treatment, suppressed specific IgE Ab responses in mice sensitized intranasally with CT-combined OVA. Allergens conjugated to liposomes may be appropriate for preventing the development of allergies to inhaled or dietary antigens in humans.     

化学偶联双特异性抗体的制备及鉴定

用化学偶联的方法,制备了抗鼠IgM-抗破伤风类毒素双特异性抗体复合物,以此作为研究B细胞抗原递呈作用的工具及模拟抗原特异性B细胞的高效抗原递呈作用。经ELISA鉴定以及通过ELISA竞争抑制试验和中和抑制试验证实,此抗体复合物确有识别两种不同抗原的双特异性。     

Hymenoptera sting hypersensitivity: IgE, IgG and haemagglutinating antibodies to bee venom constituents in relation to exposure and clinical reaction to bee stings

High levels of IgE antibodies (Ab) to bee venom constituents were mainly found in patients with bee sting hypersensitivity, whereas the sera of bee keepers usually contained high levels of IgG Ab and haemagglutinins, representing blocking antibody activity. In bee keepers there was a positive correlation between the degree of severity of an adverse reaction to bee stings and IgE Ab, a negative one between the severity of reaction and IgG Ab and haemagglutinins, a positive one between the number of stings per season and IgG Ab and haemagglutinins, and a negative one between the number of stings and IgE Ab. This suggests a mutual dependence of the production of IgE Ab and blocking Ab in a regularly exposed population. No such correlation was found in bee sting-allergic patients, indicating that other factors influence the severity of reaction in this only sporadically exposed group. The values of IgG Ab and haemagglutinins correlated well with each other, with IgG Ab giving a better correlation with the severity of reaction and number of stings than the haemagglutinins.     

Antigen-specific versus total immunoglobulin synthesis: total IgE and IgG1, but not IgG2a levels predict murine antigen-specific responses

BACKGROUND: Induction of an effective antibody (Ab) response requires delivery of multiple signals to B cells. Cross-linking of the B cell antigen receptor (BCR), signaling through CD40 and CD80/86 and cytokine signals combine to induce class switching and expression of specific isotypes. These signals are principally derived from activated, antigen (Ag)-specific T cells. In contrast, IFNgamma, the only cytokine known to induce class switch to IgG2a, can be produced systemically by activated NK or NKT cells, suggesting that Ag-nonspecific signals may also regulate IgG2a production. METHODS: Given the potential differences in regulation between IgE/IgG1 versus IgG2a, we immunized mice on day 0 with ovalbumin (OVA) in the presence of strong type-1- or type-2-immunity-inducing adjuvants and boosted mice 4 weeks later. Mice were bled during the primary immune response and after boost to assess primary and recall Ab responses. RESULTS: Regardless of strain of mice used, phenotype (type 1 versus type 2 dominated) or nature of the immune response induced (primary versus recall), strong correlations between OVA-specific and total IgE and IgG1 were demonstrated. In contrast, a consistent lack of correlation between OVA-specific and total IgG2a levels was observed in all but BALB/c mice. CONCLUSION: These data indicate that the increase in total levels of IgE/IgG1 isotypes is primarily a result of increased levels of OVA-specific Ab. In contrast, the lack of correlation between total and OVA-specific IgG2a suggests broader activation of IgG2a-producing B cells routinely occurs following exogenous Ag immunization.     

Detection of auto-anti-idiotypic antibodies to Lol p I (rye I) IgE antibodies in human sera by the use of murine idiotypes: levels in atopic and non-atopic subjects and effects of immunotherapy.

Anti-idiotypic antibodies (anti-Id Abs) are involved in the regulation of a number of immune responses including the IgE antibody production. In atopic patients, the increased synthesis of IgE antibodies could be related to a defective production of regulatory anti-Id Abs. In the present study, we first developed a sensitive assay for measuring the levels of anti-Id Abs directed against antibodies specific for Lol p I, the major allergenic determinant of Lolium perenne (rye grass). In this assay, we used previously described murine monoclonal anti-Lol p I antibodies that were shown to share epitopic specificities with human anti-Lol p I IgE and IgG antibodies, thus short-cutting the need for purification of F(ab')2 fragments of human IgG Abs and insuring optimal specificity and sensitivity. Levels of anti-Id Abs against two anti-Lol p I monoclonal antibodies (290A-167, 348A-6) were higher in normal volunteers than in untreated atopic patients. Specific immunotherapy increased the levels of anti-Id Abs to those of normal volunteers. These observations suggest a role for the Id-anti-Id network in the regulation of IgE antibody production.     

Influence of plant lipids on immune responses in mice to the major Brazil nut allergen Ber e 1

BACKGROUND: Lipids, particularly bacterial lipopolysaccharide, can impact on immune responses to proteins, with low doses enhancing type 2 responses. OBJECTIVE: We have examined the influence of natural plant lipid extracts on antibody responses provoked in mice by recombinant Ber e 1, the major allergen in Brazil nuts. METHODS: BALB/c strain mice were immunized (by intraperitoneal injection) with natural or recombinant Ber e l produced in Pichia pastoris and admixed with various lipid fractions isolated from Brazil nuts. Serum samples were analysed for specific IgE antibody by homologous passive cutaneous anaphylaxis assay and for IgG by enzyme-linked immunosorbant assay. RESULTS: Exposure to recombinant (lipid-free) Ber e 1 alone failed to induce detectable IgG or IgE antibody. Co-administration of the total lipid fraction (with reduced triglyceride levels), sterol-rich, or polar lipid fractions, resulted in marked adjuvant effects on IgG and IgE. However, the beta-sitosterol and glycolipid-rich fractions were associated with only low-level IgG antibody, and had little impact on IgE antibody production. Natural Ber e 1 containing endogenous lipids also provoked IgG and IgE antibody responses. Identical IgE and IgG antibody responses were detected regardless of whether natural or recombinant Ber e 1 was used as substrates for analyses. CONCLUSION: Endogenous Brazil nut lipids are required for the induction of optimal antibody responses to Ber e 1 in the BALB/c strain mouse. Appropriate antibody binding sites are present on both natural and recombinant forms of Ber e 1, suggesting that the impact of lipid is at the induction phase, rather than antibody recognition, and is possibly required for efficient antigen presentation.