Molecular evaluation of IgE reactivity in Hymenoptera venom allergy

The background of this PhD-thesis is based on the Hymenoptera venom allergy and the associated complex of problems of severe anaphylactic reactions. In general around 0.3 to 8.9% of the population suffer from systemic reactions after a Hymenoptera sting. Allergic reactions are frequently provoked by stings of honeybee (Apis mellifera) and yellow jacket (Vespula vulgaris). In this context 10 to 20 fatal causalities are reported in Germany per annum; in other countries the rate is between 0.1 to 0.5 per million inhabitants and it can be assumed that the estimated number of unreported cases is much higher. As permanent therapy of allergies the so called “specific immunotherapy” (SIT) has been established. In the case of Hymenoptera venom allergy a proper diagnosis and identification of the allergy provoking insect is a prerequisite for such therapy. In vitro diagnostic is based i.a on determination of IgE reactivities against the venom; however this is hampered by the complexity of the venoms. A molecular, component resolved analysis of IgE reactivities offers an opportunity to distinguish relevant IgE reactivities from clinical irrelevant cross-reactivities. Therefore in the context of this work several different honeybee venom allergens were expressed in insect cells. The choice of the expression system offers the opportunity to avoid the establishment of cross-reactive carbohydrate determinants (CCDs). Beside biochemical and allergogenic characterisation of the allergens, coupling on a diagnostic surface allowed analyses of IgE reactivities to the allergens by the use of a great cohort of patient sera. Thereby for the first time insights into the recognition patterns of allergic patients and the relevance of particular allergens could be obtained. Additional allergens relevant for therapy could be identified and the diagnostic sensitivity could be enhanced and improved from 72 to 94% by this component resolved diagnostic. Another aspect of this work is represented by the phenomena of IgE reactivities to CCDs. In this case IgE antibodies are binding specific to the α-1,3-core fucose epitope, which is present on glycosylated Hymenoptera venom allergens. The circumvention of the epitope formation offers diagnostic benefits, however the relevance of this reactivity is still unclear. For the detection of such reactivities marker proteins which bear different CCD epitopes are widely used rendering a differentiation and fine analysis difficult. Therefore a set of