Ailanthus altissima pollen (AAP) is considered as an emerging cause of respiratory allergy in United States, Italy and Iran. However, the allergenic composition of AAP is still unknown and has yet to be characterized. The present study aimed to identify AAP allergens using a proteomics-based approach. For this purpose, optimized AAP protein extracts were analyzed using 1D- and 2D- gel electrophoresis and confronted to twenty sera from individuals with respiratory allergy during the AAP season. Candidate allergens were detected using the serum from an allergic patient with clinical history of AAP pollinosis. IgE-binding spots were identified using MALDI-TOF/OF mass spectrometry and database searching. According to our results, AAP extracts were rich in proteins (up to 16.25 mg/ml) with a molecular-weight distribution ranging from 10 to 175 kDa. Two-D electrophoresis of AAP extracts revealed 125 protein spots from which 13 were IgE reactive. These IgE-binding proteins were identified as enolase, calreticulin, probable pectate lyase 6, conserved hypothetical protein and ras-related protein RHN1-like. By our knowledge, this study is the first report identifying AAP allergens. These findings will open up further avenues for the diagnosis and immunotherapy of the AAP allergy as well as for the cloning and molecular characterization of relevant allergens. Biological significance: Ailanthus altissima colonizes new areas every year in Iran and is spreading aggressively worldwide. According to USDA, the tree of heaven is now present as an invasive plant in 30 states in US (www.invasivespeciesinfo.gov/plants/treeheaven.shtml) and come to dominate large areas in many regions. Up to now, several cases of allergy to A. altissima pollen have been reported in United States, Italy and Iran [14]. However, there is still no information on the sensitizing allergens and the molecular origin of these clinical symptoms, which constitutes a serious threat to patients suffering from respiratory allergies in these regions. To our knowledge, the current study describes, therefore, the first panel of proteins responsible for IgE-mediated A. altissima pollinosis by using a gel-based proteomic approach. This work represents the pioneer proteomic investigation on Simaroubaceae spp. and provides useful insights for further studies on the allergens of this widely distributed plant family. (C) 2016 Elsevier B.V. All rights reserved.