It is still unclear whether allergen-specific immunotherapy (AIT) with birch pollen improves birch pollen-related food allergy. One reason for this may be the lack of standardized tests to assess clinical reactions to birch pollen-related foods, for example apple. We tested the applicability of recombinant (r) Mal d 1, the Bet v 1-homolog in apple, for oral challenge tests. Increasing concentrations of rMal d 1 in 0.9% NaCl were sublingually administered to 72 birch pollen-allergic patients with apple allergy. The dose of 1.6 g induced oral allergy syndromes in 26.4%, 3.2 g in 15.3%, 6.3 g in 27.8%, 12.5 g in 8.3%, 25 g in 11.1%, and 50 g in 4.2% of the patients. No severe reactions occurred. None of the patients reacted to 0.9% NaCl alone. Sublingual administration of 50 g of rMal d 1 induced no reactions in three nonallergic individuals. Our approach allows straight forward, dose-defined sublingual challenge tests in a high number of birch pollen-allergic patients that inter alia can be applied to evaluate the therapeutic efficacy of birch pollen AIT on birch pollen-related food allergy.