Purification and characterization of two new allergens from the salivary glands of the horsefly, Tabanus yao

P>Background: Horsefly bite can cause allergic reactions in humans. There is no information about allergenic horsefly proteins. Objectives: The current work aims to purify and characterize IgE-binding proteins from horsefly salivary glands. Methods: Two IgE-binding proteins, Tab a 1 and Tab a 2 with molecular weight of 26 and 35 kd, respectively, were purified and characterized from 60 000 pairs of horsefly salivary glands of Tabanus yao, respectively. Their primary sequences were determined by Edman degradation and cDNA cloning. Their allergenicity was examined using enzyme-linked immunosorbent assay (ELISA), ELISA inhibition tests, and immunoblots. Results: Immunoblotting demonstrated IgE binding by 32 and 34 of 37 (86.5% and 91.8%) subjects' sera to Tab a 1 and Tab a 2, respectively. They were identified as an antigen 5-related (Ag 5) protein and hyaluronidase, respectively. ELISA inhibitions of serum IgE reactivity to the horsefly salivary gland extract (SGE) using purified Tab a 1 and Tab a 2 were significant (about 45%). In addition, these proteins showed some IgE-binding capacity to sera of subjects with wasp sting allergy. Conclusions: We have first identified and characterized two IgE-binding proteins, Tab a 1, an Ag 5-like protein and Tab a 2, a hyaluronidase, from the horsefly salivary glands. They appear to be of importance for the allergic reactions induced by horsefly bite. These allergens are thus not only found in stinging but also found in hematophagous insects. These results also provided support for the presence of the so-called wasp-horsefly syndrome (WHS).