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Pru p 3 is a lipid transfer protein (LTP) that has been identified as the major peach (Prunus persica) allergen. However, little is known about the amount present in both raw and processed foodstuffs. Moreover, the in vivo release upon consumption of peach-containing foods remains unclear. We have developed a sensitive monoclonal antibody-based ELISA for Pru p 3. The method has been applied to measure the allergen levels in foodstuffs and the allergen release under different physiological conditions. A significant variability in all raw peaches and peach-containing foods tested has been detected. The allergen was extracted more efficiently at a low pH, and it was highly resistant to pepsin. This ELISA will be very useful in controlling the allergen concentration in diagnostics, in evaluating threshold levels in provocation tests, and in detecting hidden allergens in processed foods and cosmetics.

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Background: Group 1 allergens elicit a specific IgE response in about 90% of grass pollen-allergic patients. The aim of this work was to study the antigenic similarity among group 1 allergens from different grasses and to develop a monoclonal antibody (MAb)-based quantitation ELISA. Methods: Twenty specific MAbs were produced from BALB/c mice immunized with natural Phl p 1. These MAbs were tested for specificity with thirteen different grass pollen extracts from the Poaceae family and in cross-inhibition experiments for the binding of Phl p 1. Purified group 1 allergens from Poeae grasses (Dactylis glomerata, Lolium perenne, Festuca pratensis and Poa pratensis) were tested for parallelism in quantitation ELISA. Results: Eighteen to nineteen anti-Phl p 1 MAbs recognized the homologous allergen in pollen extracts from grasses of the Poeae tribe. In contrast, only four MAbs recognized group 1 from Cynodon dactylon and Phragmites communis. Four groups of MAbs with different epitope specificity were identified. A grass group 1 quantitation ELISA was developed using a mix of three MAbs on the solid phase and a polyclonal rabbit antibody as the second antibody. The group 1 content could be measured in different batches of Phleum pratense as well as in pollen extracts from Poeae grasses, since they showed parallel dose-response curves. Conclusions: MAbs produced in this work enabled us to show the high antigenic similarity between group 1 allergens from temperate grasses. The results prove the usefulness of the ELISA method developed for standardization of grass allergen products.

A solid-phase, monoclonal antibody-based ELISA was set up to quantitate group 5 allergens in pollen extracts of wild and cultivated Pooideae grasses. The method was able to evaluate group 5 concentration in mass units with a sensitivity in the ng/ml range and a practical working range of 1-100 ng/ml. The group 5 ELISA was compared with rocket immunoelectrophoresis for determination of allergen levels in several Phleum pratense extracts, and a very good quantitative correlation was found (r = 0.98; P < 0.0001). A highly significant correlation (r > 0.8) was also obtained in comparing allergenic potency determined by RAST inhibition to group 5 content in several wild and cultivated grass species. The results proved the usefulness of the method in the standardization of Pooideae pollen extracts employed in diagnosis and treatment.

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Background: Clustered severe adverse reactions to immunotherapy with olive pollen extracts have been occasionally reported in areas where olive trees are extensively grown. Allergic patients from these areas, in addition to the major olive pollen allergen Ole e 1, frequently recognize a recently described allergen, Ole e 9. Objective: We aimed to develop an immunoassay to measure Ole e 9 concentration and to study the variability of this allergen in olive pollen extracts. Methods: Monoclonal antibodies (mAb) to Ole e 9 were produced from mice immunized with the pure allergen. One of these mAbs was used to develop a sandwich ELISA with an anti-olive pollen extract rabbit serum as the tracer. Olive pollen batches from several suppliers were analyzed using this method. These batches were also analyzed for Ole e 1 content and biological activity. Results: A 10-fold variation between the extreme values was found for the biological activity of the batches analyzed. Ole e 1 concentration showed a 25-fold variation. Variability of Ole e 9 concentration was extremely high, up to 161 times. The ratio Ole e 1/Ole e 9 varied in a range from 0.6 to 390.4. Conclusion: The availability of a mAb-based ELISA for Ole e 9 made it possible for us to detect an important source of variability in olive pollen batches. This variability may be the cause of outbreaks of adverse reactions in the course of immunotherapy treatments, which have sometimes been observed among olive-allergic patients living in areas with very high levels of airborne olive pollen.