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Notes: For most foods, true standardization is not yet feasible because there is insufficient information on the relative importance of individual allergens and their variants. Standardization without sufficient information may easily be counterproductive because improvements are less likely to be implemented. In the analysis of natural test material, the following principles apply:〈list xml:id="l1" style="custom"〉1) RAST inhibition is usually inferior to other means of allergen quantitation.2) Immunoblot is inefficient for some “important” allergens and over-efficient for some “unimportant” allergens and may therefore be deceptive.3) Single-component assays are the only satisfactory way to describe complex mixtures.Improving the actual food-testing procedure is important, but will not alone result in a reliable diagnostic procedure. Tests for measuring “effect modifiers” will have to be developed in order to predict in vivo reactions to foods.

Notes: To obtain more information on IgE cross-reactivity between bumblebee venom and honeybee venom, we tested sera from venom-sensitized patients for specific IgE against venoms from the European bumblebee (Bombus terrestris), the North American bumblebee (Megabombm pennsylvanicus), and the honeybee (Apis mellifera). RAST, RAST-inhibition, and immunoblotting experiments indicate that bumblebee venom and honeybee venom contain venom-specific IgE-binding epitopes. These results suggest that immunotherapy using honeybee venom may not be effective in all bumblebee venom-allergic patients. Our experiments also revealed differences in IgE binding for venom from European and American bumblebees.

Notes: Crossreactivity to Dactylis glomerata, Festuca rubra, Phleum pratense, Anthoxanthum odoratum, Secale cereale, Zen mays, and Phragmites communis of IgE antibodies against Lol p I or Lol p V was investigated by means of RAST-inhibition. Within a group of sera the degree of crossreactivity was demonstrated to be highly variable. Individual sera were not always equally crossreactive to all pollen species. A high degree of crossreactivity for Group I allergens did not necessarily implicate the same for Group V. Group I and Group V representatives were found to be present in all eight species. It was demonstrated that within this group of grass species significant quantitative and qualitative differences exist, with respect to Group I and Group V allergens. Species with a low phylogenetic affinity to Lolium perenne, like Zea mays and Phragmites communis showed a very low degree of reactivity, even when measured with the most crossreactive sera. A higher taxonomic relationship however, did not always implicate a closer antigenic resemblance. Antigenically both allergens from Zea mays are more similar to Lol p I and Lol p V, than the analogues in Secale cereale.

Notes: We report on the relation between the month of birth and the chance of developing an IgE antibody response as found in a study sample of 150000 subjects. Our results confirm that for the three seasonal allergens birch pollen, grass pollen and house dust mite, an increased relative risk was found for subjects born up to 3 months before the main season for that allergen in The Netherlands. For cat and dog allergy an increased relative risk was found from November to January, perhaps reflecting increased exposure to these pets during the winter. Surprisingly, however, also for egg white and cow's milk a clearly increased relative risk was found from November to January and a decreased relative risk in May. These data support the hypothesis of a ‘sensitive’ period in the first months of life during which allergen exposure is more likely to prime for an allergy later in life. The results with the non-seasonal allergens suggest that another seasonal factor exists which early in life assists (or prevents) priming by allergen.

Notes: Background: Fluctuations in the level of mite allergens in domestic house dust are the result of changes in the balance between synthesis, removal and decay. Purely physical forces as well as enzymatic degradation, mediated by house dust inhabiting microbes, may contribute to the decay of allergens in domestic dust. Knowledge about the speed of decay is essential for an understanding of the dynamics of allergen levels.Objective: The present study is a quantitative assessment of the speed of decay at nine combinations of temperature (15°C, 20°C and 25°C) and relative humidity (33%, 55% and 75%).Methods: Samples of mite infested material of an old rug were stored at these temperature/relative humidity-combinations for 6, 12 or 18 months, after the mites were killed by cither a freezing treatment or an acaricide (lindane). The microbes living in the rug presumably survive these treatments. Concentrations of Der p I and Der p II + Der f II. in extracts of the rug material, were measured by a radio immunoassay.Results: No significant changes in the levels of Der p I and Der p II +Der f II, could be detected even after 11/2 year at a high temperature and humidity.Conclusion: These findings incidate that mite allergens can be extremely stable under normal domestic circumstances.

Notes: Background In children at high risk of inhalation allergy, food sensitization is associated with an increased risk for sensitization to inhalant allergens. Furthermore, this association was also found in a cross-sectional study.Objective To examine in a prospective study, whether levels of IgG to foods (i.e. mixture of wheat and rice, mixture of soy bean and peanut, egg white, cow's milk, meat, orange and potato) indicate an increased risk for the future development of IgE antibodies to inhalant allergens in a low-risk population and whether they can be used as predictors of the subsequent development of IgE antibodies in young, initially IgE-negative children.Methods Coughing children, aged 1–5, visiting their GPs, were tested for IgE antibodies to mite, dog and cat (RAST) and IgG (ELISA) to foods. All IgE-negative children were retested for IgE antibodies after two years. The IgG results (66 percentiles) of the first blood sample were compared to the RAST-scores of the second blood sample.Results After two years, 51 out of 397 (12.8%) originally IgE-negative children, had become IgE-positive for cat, dog and/or mite. An increased IgG antibody level to wheat-rice (OR = 2.2) and to orange (OR = 2.0) indicated an increased risk of developing IgE to cat, dog or mite allergens. In addition to IgG to a mixture of wheat-rice and orange; total IgE, breastfeeding, eczema as a baby and age were the most important predictors for the subsequent development of IgE to inhalant allergens.Discussion An increased IgG antibody level to a mixture of wheat-rice or orange, indicates an increased risk of developing IgE to cat, dog or mite allergens. This indicates that excessive activity of the mucosal immune system is present before IgE antibodies to airborne allergens can be demonstrated. Nevertheless, IgG to foods is not very helpful (with a positive predictive value of 16.5%, and negative predictive value of 90.6%) in identifying individual children at risk in clinical practice. However, besides other risk factors, IgG to wheat-rice and to orange could be useful as a screening test for studies in the early identification, i.e. before IgE antibodies can be detected, of children with an increased risk of developing IgE antibodies in the future.

Notes: Background IgE titres tend to rise early after the start of immunotherapy, followed by a decline to pre-immunotherapy levels or lower.Objectives We were interested to ktiow whether the early increase in IgE antibodies includes new specificities of IgE, and whether these responses persist.Methods Sera of 64 patients undergoing grass pollen immunotherapy were tested for IgE against four purified grass pollen allergens: Lol p 1. 2, 3, and 5. At least two serum samples were taken, one before the start of therapy and one between 5 and 18 months after the first immunization (mean: 10 months).Results The mean IgE responses to Lol p 1, 2 and 3 showed a moderate but not significant increase. In contrast, the mean IgE response to Lol p 5 showed a significant decrease of 〉30%. IgE against total Lolium perenne pollen extract moderately increased (〉20%), showing that a RAST for total pollen is not always indicative for the development of IgE against its major allergens. For 〉40% of the patients it was found that IgE against one or more of the four allergens increased, while IgE against the remaining allergen(s) decreased. Eor 10 sera the ratio of IgE titres against at least two allergens changed by at least a factor of 5. The changes in specific IgE also included conversions from negative (〈 0.1 RU) to positive (0.6 to 5.0 RU) for five patients. For two patients, the induction of these ‘new’ IgE antibodies against major allergens was shown to result in a response that was persistent over several years.Conclusion Although active induction of new IgE specificities by immunotherapy was not really proven, the observations in this study indicate that monitoring of IgE against purified (major) allergens is necessary to evaluate changes in specific IgE in a reliable way.

Notes: Six monoclonal antibodies against Bet v I, the major cross-reactive allergen of birch pollen (Betula verrucosa), were obtained. Four did not react with fruits, but two monoclonal antibodies (mAbs) (5H8 and 9C11) were reactive with apple and other fruits. These two cross-reactive antibodies reacted with identical or overlapping sites, but differed in their relative degree of cross-reactivity toward various fruits and hazelnut. Cross-reactive human IgE antibodies reacted with a nonoverlapping epitope, as indicated by results of a two-site radioimmunoassay (RIA) with the fruit-reactive mAb 9C11. By isoelectric focusing (IEF) in conjunction with immunoblotting, a maximum of seven isoforms could be distinguished. Depletion of birch-pollen extract for Bet v I with the most reactive mAb (7F7) removed approximately 95% of the IgE cross-reactivity between birch pollen and apple extract. The remaining 5% cross-reactive material was still capable of inhibiting the binding of IgE to apple allergen completely, and was reactive with mAbs 5H8 and 3C4. By means of IEF/immunoblot, it was shown that these mAbs recognize an isoform of Bet v I that is poorly, if at all, recognized by mAb 7F7. These results illustrate the heterogeneity of Bet v I, both with respect to the cross-reactive sites as well as to the backbone structure. This type of heterogeneity has possible implications for the use of monoclonal antibodies in allergen standardization.

Notes: Background: Bovine serum albumin (BSA) is widely used to block nonspecific binding in immunochemical assays. Whereas a previous study had indicated that soluble allergen present during the incubation with anti-IgE in the RAST did not affect bound IgE, we reinvestigated this in the current study, using IgE elution from BSA by soluble BSA as a test system. Methods: Sepharose-coupled BSA (0.08, 0.4, 2, or 10 µg BSA/test) was incubated overnight with serum and washed. The Sepharose was then incubated with different concentrations of soluble BSA (0, 12, 60, 300, or 1500 µg/test), washed again, and incubated with radioactive anti-IgE. The effect on IgE binding was investigated for various incubation periods (t=0, 1, 2, 4, and 20 h). Results: Incubation in buffer without BSA did not change IgE binding. Soluble BSA eluted IgE antibodies from immobilized BSA by up to 85%. If the BSA density on the solid phase was ≥2 µg/test, the elution efficiency was dependent on the levels of both immobilized BSA and soluble BSA. At lower densities, the dissociation was dependent only on the concentration of soluble BSA. The time needed to obtain 50% IgE elution (t½) was less if the density of immobilized BSA decreased. Below the critical density (0.8 µg BSA/mg solid phase), t½ was independent of the coating density (45 min). Probably all IgE antibodies are monovalently bound below this density. Conclusions: Dissociation of IgE from immobilized protein in the presence of soluble protein should be taken into account, particularly when IgE to mammalian serum albumin is involved (milk, meat, or animal dander).