Detection of trace amounts of hidden allergens: hazelnut and almond proteins in chocolate

Many patients with immediate type allergy to tree pollen also suffer from intolerance to hazelnuts and almonds. Since rather low levels of hazelnut and almond proteins can provoke an allergic reaction in sensitized individuals, an immunoblot technique has been developed for the detection of potentially allergenic hazelnut and almond proteins in chocolate. Initially, IgE binding hazelnut and almond proteins were detected by immunoprobing with allergic patients’ sera. For routine analysis, patients’ sera were substituted with polyclonal rabbit antisera, and sensitivity was enhanced by the use of a chemiluminescent detection method. This technique allowed the detection of less than 0.5 mg of hazelnut or almond proteins per 100 g of chocolate (=5 ppm). It was applied for routine screening purposes in product quality control as well as for optimization of cleaning steps of filling facilities to minimize cross contamination during production.